家蚕含染色质域基因的鉴定与MSL-3同源体和Bm-MOF基因的功能研究
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摘要
家蚕是一种重要的经济昆虫，在生产上的主要应用为泌丝结茧。目前我国共有种桑养蚕农户两千多万户，是在国际市场上处于绝对优势的特色产业。但与日本等发达国家相比，由于蚕茧的经济性状差造成我国的生丝质量普遍不高，严重影响了蚕农和国家的经济利益。其中，茧丝生产上最重要的经济性状产丝量和丝质容易受外因遗传即环境条件的影响。染色质域参与外因遗传密码的遗传，同时也是外因遗传密码的识别者，对于它在外因遗传及染色质形态上的调控作用，将有助于家蚕数量性状遗传的研究，对进一步巩固我国蚕丝业的世界地位，提高其竞争力和农民收入也具有重大意义。本研究基于家蚕全基因组框架图和EST数据，对家蚕含染色质域基因进行了分析，并且有机结合双杂交系统的反应环境和共纯化技术，对MSL-3同源基因和MOF基因进行了克隆、表达和功能研究。获得的主要结果如下：
1．家蚕基因组中含染色质域同源基因的鉴定
将酵母、果蝇和人的所有的含染色质域基因在家蚕基因组中进行同源性分析，结果显示，11个含染色质域基因在家蚕基因组中具有同源体。应用EST证据和RT-PCR鉴定基因在家蚕中都有表达，此乃首次从家蚕基因组水平鉴定家蚕含染色质域基因。
比较分析家蚕与果蝇的含染色质域基因，几乎所有的果蝇的含染色质域基因在家蚕中都具有同源体。家蚕有三个CHD家族基因，较果蝇少一个，而较人等哺乳动物中有明显的减少。CHD家族的三个基因的表达情况也有较大差异，BmCHD1在所检测的所有时期中均没有表达；而BmCHD3除滞育卵中为微弱可见的表达量，其它三个时期有明显的表达；BmCHD6在二龄起蚕和蛹期较达量较高，其它两个时期表达量较低。
果蝇SUV39、HP1家族和PC家族基因在家蚕中具有同等数目的同源基因。RT-PCR鉴定表达情况有较大差异，BmHP1α在所检测的时期中都有表达：BmHP1γ在所检测的时期中有显著差异的表达，蛾期表达量较高，其次为蛹期，二龄起蚕和滞育卵中相对较低；BmHP1β在二龄起蚕有明显表达，滞育卵中为微弱表达，而蛹期和蛾期完全没有表达。BmSu(var)39与BmCHD6较相似，在二龄起蚕和蛹期有表达，其它两个时期无表达；BmPC在蛹期表达量较高，蛾期、二龄起蚕依次降低，滞育卵中没有表达。
果蝇的MOF基因和MSL-3基因是在果蝇的剂量补偿中起作用。对于剂量补偿，到目前为止，还没有证据表明家蚕具有剂量补偿机制。在家蚕中具有ms13和mof同源体是一个有趣的现象。据报道哺乳动物的MOF和MSL3的同源基因组成一个转录调控复合物而非剂量补偿复合物，暗示家蚕可能也有这种机制，利用这些蛋白调控基因的转录。
2．家蚕MSL-3同源体的克隆与原核表达
通过RT-PCR技术，克隆了家蚕MSL-3同源基因：Bm-ms13和Bm-mrg15，序列已在国际核酸数据库GenBank上登录，登录号分别为DQ887343和DQ888173，基因的cDNA长度分别为：2356bp和1113bp。克隆的开放阅读框长度分别为1665bp和1020bp，分别编码554和339个氨基酸残基，均含有一个N-末端染色质域和一个MRG结构域，无信号肽序列。同源序列的多序列比对结果显示两基因编码的蛋白在两个结构域区非常保守，尤其是MRG结构域，而部分基因的编码蛋白发生了染色质域的缺失。Bm-ms13基因的编码蛋白与果蝇MSL-3蛋白质序列高度相似，有61％的一致性，而Bm-mrg15与人的MRG15蛋白序列的相似性更高，有47％的一致性。在家蚕基因组中的BLASTN程序检索，发现两基因在基因组均只有一个拷贝，cDNA与基因组序列比对，发现Bm-ms13具有13个外显子、12个内含子，除一处的外显子／内含子边界不符GT-AG规则外，其它均符合。Bm-mrg15基因在家蚕基因组中也为单拷贝，且有6个外显子、5个内含子，各外显子／内含子边界均符合GT-AG规则。RT-PCR结果显示两基因在所检测的家蚕时期和组织中均有表达，说明其可能在家蚕的发育过程中起着重要的作用。
分别将Bm-ms13和Bm-mrg15亚克隆到pGEX-4t-2原核表达载体，诱导表达产物的SDS-PAGE电泳表明pGEX-4t-2-BmMSL3仅在低温18℃培养时在70KDa的位置比对照多出一条明显的特异带，可溶组分具有GST酶活性，可能为Bm-ms13重组蛋白的截短表达形式；pGEX-4t-2-BmMRG15在65KDa的位置比对照多出一条明显的表达带，低温诱导其可溶性增加，且具有GST酶活性，表明目的蛋白得到了高效表达。
3．pET-50b-BmMOF的优化表达及其与家蚕MSL-3同源体重组蛋白的相互作用
采用不同的培养基、IPTG浓度和温度对家蚕pET50b-BmMOF重组质粒的表达条件进行优化，得到具有高效且可溶的表达条件为：TB培养基，0.4mM IPTG，25℃。
分别共转化pET50b-BmMOF和两个MSL-3同源基因的重组质粒至BL21(DE3)菌株，应用同时具有氨苄抗性和卡那霉素抗性的培养基筛选后，进行培养和融合蛋白的诱导表达，以及表达蛋白的His．Bind树脂的共纯化，并通过SDS-PAGE电泳、共纯化及测定纯化前后蛋白的GST酶活性，检测目的蛋白的表达及Bm-MOF和MSL-3同源体间的相互作用情况。SDS-PAGE电泳结果表明目的蛋白均得到了较好的诱导和表达，但共纯化后的蛋白仅观察到有pET50b-BmMOF融合蛋白，没发现有MSL-3重组蛋白的存在。GST酶活性检测也表明目的蛋白得到了诱导表达，纯化后蛋白的弱的GST活性，说明其中也有少量的MSL-3重组蛋白，家蚕的两基因间可能存在弱相互作用。
4．家蚕含CD域基因的序列分析与CD域的内含子的插入情况和进化分析
大部分家蚕含染色质域基因的结构域与其它物种同源基因一样，除含有典型的CD域外，还都有其典型的所属家族的结构域，只有BmHP1γ为两个CD域，不含CSD域。虽然CD域和CSD域具有相似的结构和氨基酸组成，即都由三个β折叠和一个α螺旋组成，但其功能存在明显的差异。BmHP1γ特殊的结构组成可能与家蚕的特殊的染色体形态相关，其在家蚕中的功能也有待于进一步的研究。
家蚕的基因与人的同源基因具有较高的相似性，尤其BmCHD1、BmCHD6、BmHP1α、BmHP1β、Bm-mof，而它们与果蝇基因的相似性稍差。这是一个比较有趣的问题，从常规进化论的角度来说，鳞翅目昆虫家蚕和双翅目昆虫果蝇间的进化距离和基因的差异应该小于家蚕和人之间，但现在却刚好相反。
含CD域基因及CD域的内含子插入情况的分析结果表明，虽然家蚕的Su(Var)3-9和MOF基因的内含子插入情况与果蝇同源基因相似，都没有内含子，而人的同源基因有。但家蚕与人的CHD家族和HP1家族的两个CD域，大多都有内含子的插入。而果蝇的两个家族的基因中，只有HP1α的第一个CD域内有内含子的插入，其它的CD域内都没有内含子，甚至HP1β和HP1γ的整个基因中都不存在内含子。
The silkworm, Bombyx mori, an important economic insect, can secrete silk to form cocoon. There are all 20,000,000 sericulturists now in our country and it is an characteristic industry in advantage preponderance in international market. But because of the low economy character, the quality of most of the raw silk is poor in our country compared to developed countries such as Japan, which decreases the economic profit of sericulturists and our country. The foremost character in silk manufacture, output and quality of silk, can be easily affected by environment, epigenetic factor. Chromodomain, a recognizer of epigenetic codon, participates in the heredity of epigenetic codon. The role of chromodomain in epigenetic and in chromatin conformation could redound to the study of quantitive heredity and it also has great significance in reinforcing the world station of our silk industry, improving the competition and increasing the income of farmer. Based on the silkworm genome draft and EST(expressed sequence tag), we analyzed the chromodomain-containing(CD gene) proteins of silkworm and studied on the cloning and function of MSL-3 homologs and MOF genes in them. The main results present as follows.
1. Identification of homologs of CD genes in silkworm genome
Homology analysis between the silkworm genome and all the known genes of yeast, Drosophila and human CD genes shown that 11 homologs were found in silkworm and all of them expressed in silkworm by EST proof and RT-PCR. Homologs of CD genes were reported firstly in silkworm genome.
Analysis of the CD genes between silkworm and Drosophila indicated that almost all the CD genes of Drosophila has homologs in silkworm. There are 3 CHD family genes, one less than in Drosophila, but obvious decreased to in mammal such as human. The expression of the 3 CHD family genes has obvious difference by RT-PCR. BmCHD1 has no expression in all the tested stages, BmCHD3 has visible expression except in dispaused eggs, faint expression in it, and the expression of BmCHD6 is high in 1st day of 2nd instar larva and pupa,low in the other stages.
The amount of SUV39, HP1 and PC family genes between silkworm and Drosophila is equal. The expression detected by RT-PCR indicate that there are distinct difference among them. BmHP1αexpresses in all the tested periods, BmHP1γis notably difference, high in moth, pupa is second and relative low in 1st day of 2nd instar larva and dispaused eggs, while BmHP1βhas no expression in pupa and moth, although it expresses in the other two stages. Similar to BmCHD6, BmSu(var)39 is expressed in 1st day of 2nd instar larva and pupa, no expression in the other two stages. The expression of BmPC is decreasing in pupa, moth, 1st day of 2nd instar larva and even no in dispaused eggs.
The msl3 and mof gene of Drosophila function in dosage compensation. It is no evidence to make shown that there are dosage compensation in silkworm now. The being of msl3 and mof in silkworm is interesting. The report that MOF and MSL3 in mammals compose a transcriptional complex and not involved in dosage compensation indicates that their function in silkworm maybe the same with that in mammals. 2. The cloning and prokaryotic expression of MSL-3 homologs in silkworm
The MSL-3 homologs in silkworm Bm-msl3 and Bm-mrg15 were cloned by RT-PCR and the accession number of them in GenBank are DQ887343 and DQ888173. The cDNA length of them are 2356 bp and 1113 bp, respectively, with an ORF of 1665 bp and 1020 bp coding 554 and 339 residues, which contain a N-ter CD and an MRG domain, no signal peptide. Multiple sequence alignment of homologs indicate that the two structure domains of coded proteins are conserved, especially the MRG domain, while the CD of some genes are lack. The Bm-msl3 has high similarity with drosophila homologs, 61% identity between them. While the similarity of MRG15 gene between silkworm and human is high, 47% identity between them. The results of BLASTN search of the two genes in silkworm genome shown that they both have only one copy in genome. The comparison between cDNA and genome indicates that 13 exons and 12 introns are in Bm-msl3 and all of the boundaries of exon/intron is GT-AG except one. Similarly, gene Bm-mrg15 has 6 exons and 5 introns and the boundaries of them are all GT-AG. RT-PCR experiment indicated that the two genes were expressed in all tested tissues and stages, which seem to imply its key role in silkworm development.
Prokaryotic expression of the two genes was carried out through sub-cloning into pGEX-4t-2 vector. SDS-PAGE electrophoresis of the product after inducement indicated that pGEX-4t-2-BmMSL3 had no expression except in 18℃, low incubation temperature and the specific band is about 70 KDa and the solute composition has GST enzyme activity, which is postulated the cut short expression of recombinant Bm-msl3. pGEX-4t-2-BmMRG15 is successfully expressed with a 65 KDa specific band, and the solute composition of expressed protein is increased and has GST enzyme acitivy when inducement in low temperature.
3. Expression optimization of pET50b-BmMOF and its interaction with recombinant MSL-3
Different culture medium, IPTG concentration, and temperature were applied to optimize the expression of recombinate plasmid pET50b-BmMOF and the optimum condition was TB culture medium, 0.4 mM IPTG and 25℃.
The co-expression of Bm-mof and MSL-3 homologs came true by transformation of recombinant pET50b-BmMOF and MSL-3 homologs to the same BL21(DE3) and screen by the culture medium with ampicillin and kanamycin sulfate. Expression proteins were copurification by His.Bind. SDS-PAGE, co-purification and detection of GST enzyme activity of the protein before and after purification were all carried out to identify the interaction between them. All the proteins has effective inducement and efficient expression by SDS-PAGE. But there is only recombinant protein pET50b-BmMOF and no recombinant MSL-3 homologs in the proteins after purification. The results of GST enzyme activity is the same except after purification, which has weak GST enzyme activity, too. All the results indicatd that there may be weak interaction between the two gene of silkworm.
4. The analysis of silkworm CD genes and the intron and phylogenesis of CD
Same to the homologs, most of silkworm CD genes has typical domain of the family which it is affiliated besides CD, except one, BmHP1γ, two CD contained and no CSD. Although the structure and residues composition between CD and CSD is similar, and both composed by 3β-sheets and oneα-helix, the function of them has distinct difference. The special structure of BmHP1γmay be related to chromosome shape of silkworm and the function of it needs further studies.
The results of homologous search indicate that they are more similar to human homologs, especially BmCHD1、BmCHD6、BmHP1α、BmHP1β、Bm-mof, and the similarity to drosophila is lower, which is an interesting thing for the distance in evolution between silkworm(lepidopteran) and drosophila(dipteran) should be less than silkworm and human in general evolutionism, while it is the contrary in this event.
The analysis results of intron of CD genes and CD domain shown that although Su(Var)3-9 and MOF gene of silkworm and drosophila are similar, no intron in them, while homologs of human has. To the CD domain of CHD family and HP1 family of silkworm and human, most of them have. While in drosophila, there are no except HP1α, one intron in the first CD domain, and no intron in the HP1βand HP1γgenes.
1．家蚕基因组中含染色质域同源基因的鉴定
将酵母、果蝇和人的所有的含染色质域基因在家蚕基因组中进行同源性分析，结果显示，11个含染色质域基因在家蚕基因组中具有同源体。应用EST证据和RT-PCR鉴定基因在家蚕中都有表达，此乃首次从家蚕基因组水平鉴定家蚕含染色质域基因。
比较分析家蚕与果蝇的含染色质域基因，几乎所有的果蝇的含染色质域基因在家蚕中都具有同源体。家蚕有三个CHD家族基因，较果蝇少一个，而较人等哺乳动物中有明显的减少。CHD家族的三个基因的表达情况也有较大差异，BmCHD1在所检测的所有时期中均没有表达；而BmCHD3除滞育卵中为微弱可见的表达量，其它三个时期有明显的表达；BmCHD6在二龄起蚕和蛹期较达量较高，其它两个时期表达量较低。
果蝇SUV39、HP1家族和PC家族基因在家蚕中具有同等数目的同源基因。RT-PCR鉴定表达情况有较大差异，BmHP1α在所检测的时期中都有表达：BmHP1γ在所检测的时期中有显著差异的表达，蛾期表达量较高，其次为蛹期，二龄起蚕和滞育卵中相对较低；BmHP1β在二龄起蚕有明显表达，滞育卵中为微弱表达，而蛹期和蛾期完全没有表达。BmSu(var)39与BmCHD6较相似，在二龄起蚕和蛹期有表达，其它两个时期无表达；BmPC在蛹期表达量较高，蛾期、二龄起蚕依次降低，滞育卵中没有表达。
果蝇的MOF基因和MSL-3基因是在果蝇的剂量补偿中起作用。对于剂量补偿，到目前为止，还没有证据表明家蚕具有剂量补偿机制。在家蚕中具有ms13和mof同源体是一个有趣的现象。据报道哺乳动物的MOF和MSL3的同源基因组成一个转录调控复合物而非剂量补偿复合物，暗示家蚕可能也有这种机制，利用这些蛋白调控基因的转录。
2．家蚕MSL-3同源体的克隆与原核表达
通过RT-PCR技术，克隆了家蚕MSL-3同源基因：Bm-ms13和Bm-mrg15，序列已在国际核酸数据库GenBank上登录，登录号分别为DQ887343和DQ888173，基因的cDNA长度分别为：2356bp和1113bp。克隆的开放阅读框长度分别为1665bp和1020bp，分别编码554和339个氨基酸残基，均含有一个N-末端染色质域和一个MRG结构域，无信号肽序列。同源序列的多序列比对结果显示两基因编码的蛋白在两个结构域区非常保守，尤其是MRG结构域，而部分基因的编码蛋白发生了染色质域的缺失。Bm-ms13基因的编码蛋白与果蝇MSL-3蛋白质序列高度相似，有61％的一致性，而Bm-mrg15与人的MRG15蛋白序列的相似性更高，有47％的一致性。在家蚕基因组中的BLASTN程序检索，发现两基因在基因组均只有一个拷贝，cDNA与基因组序列比对，发现Bm-ms13具有13个外显子、12个内含子，除一处的外显子／内含子边界不符GT-AG规则外，其它均符合。Bm-mrg15基因在家蚕基因组中也为单拷贝，且有6个外显子、5个内含子，各外显子／内含子边界均符合GT-AG规则。RT-PCR结果显示两基因在所检测的家蚕时期和组织中均有表达，说明其可能在家蚕的发育过程中起着重要的作用。
分别将Bm-ms13和Bm-mrg15亚克隆到pGEX-4t-2原核表达载体，诱导表达产物的SDS-PAGE电泳表明pGEX-4t-2-BmMSL3仅在低温18℃培养时在70KDa的位置比对照多出一条明显的特异带，可溶组分具有GST酶活性，可能为Bm-ms13重组蛋白的截短表达形式；pGEX-4t-2-BmMRG15在65KDa的位置比对照多出一条明显的表达带，低温诱导其可溶性增加，且具有GST酶活性，表明目的蛋白得到了高效表达。
3．pET-50b-BmMOF的优化表达及其与家蚕MSL-3同源体重组蛋白的相互作用
采用不同的培养基、IPTG浓度和温度对家蚕pET50b-BmMOF重组质粒的表达条件进行优化，得到具有高效且可溶的表达条件为：TB培养基，0.4mM IPTG，25℃。
分别共转化pET50b-BmMOF和两个MSL-3同源基因的重组质粒至BL21(DE3)菌株，应用同时具有氨苄抗性和卡那霉素抗性的培养基筛选后，进行培养和融合蛋白的诱导表达，以及表达蛋白的His．Bind树脂的共纯化，并通过SDS-PAGE电泳、共纯化及测定纯化前后蛋白的GST酶活性，检测目的蛋白的表达及Bm-MOF和MSL-3同源体间的相互作用情况。SDS-PAGE电泳结果表明目的蛋白均得到了较好的诱导和表达，但共纯化后的蛋白仅观察到有pET50b-BmMOF融合蛋白，没发现有MSL-3重组蛋白的存在。GST酶活性检测也表明目的蛋白得到了诱导表达，纯化后蛋白的弱的GST活性，说明其中也有少量的MSL-3重组蛋白，家蚕的两基因间可能存在弱相互作用。
4．家蚕含CD域基因的序列分析与CD域的内含子的插入情况和进化分析
大部分家蚕含染色质域基因的结构域与其它物种同源基因一样，除含有典型的CD域外，还都有其典型的所属家族的结构域，只有BmHP1γ为两个CD域，不含CSD域。虽然CD域和CSD域具有相似的结构和氨基酸组成，即都由三个β折叠和一个α螺旋组成，但其功能存在明显的差异。BmHP1γ特殊的结构组成可能与家蚕的特殊的染色体形态相关，其在家蚕中的功能也有待于进一步的研究。
家蚕的基因与人的同源基因具有较高的相似性，尤其BmCHD1、BmCHD6、BmHP1α、BmHP1β、Bm-mof，而它们与果蝇基因的相似性稍差。这是一个比较有趣的问题，从常规进化论的角度来说，鳞翅目昆虫家蚕和双翅目昆虫果蝇间的进化距离和基因的差异应该小于家蚕和人之间，但现在却刚好相反。
含CD域基因及CD域的内含子插入情况的分析结果表明，虽然家蚕的Su(Var)3-9和MOF基因的内含子插入情况与果蝇同源基因相似，都没有内含子，而人的同源基因有。但家蚕与人的CHD家族和HP1家族的两个CD域，大多都有内含子的插入。而果蝇的两个家族的基因中，只有HP1α的第一个CD域内有内含子的插入，其它的CD域内都没有内含子，甚至HP1β和HP1γ的整个基因中都不存在内含子。
The silkworm, Bombyx mori, an important economic insect, can secrete silk to form cocoon. There are all 20,000,000 sericulturists now in our country and it is an characteristic industry in advantage preponderance in international market. But because of the low economy character, the quality of most of the raw silk is poor in our country compared to developed countries such as Japan, which decreases the economic profit of sericulturists and our country. The foremost character in silk manufacture, output and quality of silk, can be easily affected by environment, epigenetic factor. Chromodomain, a recognizer of epigenetic codon, participates in the heredity of epigenetic codon. The role of chromodomain in epigenetic and in chromatin conformation could redound to the study of quantitive heredity and it also has great significance in reinforcing the world station of our silk industry, improving the competition and increasing the income of farmer. Based on the silkworm genome draft and EST(expressed sequence tag), we analyzed the chromodomain-containing(CD gene) proteins of silkworm and studied on the cloning and function of MSL-3 homologs and MOF genes in them. The main results present as follows.
1. Identification of homologs of CD genes in silkworm genome
Homology analysis between the silkworm genome and all the known genes of yeast, Drosophila and human CD genes shown that 11 homologs were found in silkworm and all of them expressed in silkworm by EST proof and RT-PCR. Homologs of CD genes were reported firstly in silkworm genome.
Analysis of the CD genes between silkworm and Drosophila indicated that almost all the CD genes of Drosophila has homologs in silkworm. There are 3 CHD family genes, one less than in Drosophila, but obvious decreased to in mammal such as human. The expression of the 3 CHD family genes has obvious difference by RT-PCR. BmCHD1 has no expression in all the tested stages, BmCHD3 has visible expression except in dispaused eggs, faint expression in it, and the expression of BmCHD6 is high in 1st day of 2nd instar larva and pupa,low in the other stages.
The amount of SUV39, HP1 and PC family genes between silkworm and Drosophila is equal. The expression detected by RT-PCR indicate that there are distinct difference among them. BmHP1αexpresses in all the tested periods, BmHP1γis notably difference, high in moth, pupa is second and relative low in 1st day of 2nd instar larva and dispaused eggs, while BmHP1βhas no expression in pupa and moth, although it expresses in the other two stages. Similar to BmCHD6, BmSu(var)39 is expressed in 1st day of 2nd instar larva and pupa, no expression in the other two stages. The expression of BmPC is decreasing in pupa, moth, 1st day of 2nd instar larva and even no in dispaused eggs.
The msl3 and mof gene of Drosophila function in dosage compensation. It is no evidence to make shown that there are dosage compensation in silkworm now. The being of msl3 and mof in silkworm is interesting. The report that MOF and MSL3 in mammals compose a transcriptional complex and not involved in dosage compensation indicates that their function in silkworm maybe the same with that in mammals. 2. The cloning and prokaryotic expression of MSL-3 homologs in silkworm
The MSL-3 homologs in silkworm Bm-msl3 and Bm-mrg15 were cloned by RT-PCR and the accession number of them in GenBank are DQ887343 and DQ888173. The cDNA length of them are 2356 bp and 1113 bp, respectively, with an ORF of 1665 bp and 1020 bp coding 554 and 339 residues, which contain a N-ter CD and an MRG domain, no signal peptide. Multiple sequence alignment of homologs indicate that the two structure domains of coded proteins are conserved, especially the MRG domain, while the CD of some genes are lack. The Bm-msl3 has high similarity with drosophila homologs, 61% identity between them. While the similarity of MRG15 gene between silkworm and human is high, 47% identity between them. The results of BLASTN search of the two genes in silkworm genome shown that they both have only one copy in genome. The comparison between cDNA and genome indicates that 13 exons and 12 introns are in Bm-msl3 and all of the boundaries of exon/intron is GT-AG except one. Similarly, gene Bm-mrg15 has 6 exons and 5 introns and the boundaries of them are all GT-AG. RT-PCR experiment indicated that the two genes were expressed in all tested tissues and stages, which seem to imply its key role in silkworm development.
Prokaryotic expression of the two genes was carried out through sub-cloning into pGEX-4t-2 vector. SDS-PAGE electrophoresis of the product after inducement indicated that pGEX-4t-2-BmMSL3 had no expression except in 18℃, low incubation temperature and the specific band is about 70 KDa and the solute composition has GST enzyme activity, which is postulated the cut short expression of recombinant Bm-msl3. pGEX-4t-2-BmMRG15 is successfully expressed with a 65 KDa specific band, and the solute composition of expressed protein is increased and has GST enzyme acitivy when inducement in low temperature.
3. Expression optimization of pET50b-BmMOF and its interaction with recombinant MSL-3
Different culture medium, IPTG concentration, and temperature were applied to optimize the expression of recombinate plasmid pET50b-BmMOF and the optimum condition was TB culture medium, 0.4 mM IPTG and 25℃.
The co-expression of Bm-mof and MSL-3 homologs came true by transformation of recombinant pET50b-BmMOF and MSL-3 homologs to the same BL21(DE3) and screen by the culture medium with ampicillin and kanamycin sulfate. Expression proteins were copurification by His.Bind. SDS-PAGE, co-purification and detection of GST enzyme activity of the protein before and after purification were all carried out to identify the interaction between them. All the proteins has effective inducement and efficient expression by SDS-PAGE. But there is only recombinant protein pET50b-BmMOF and no recombinant MSL-3 homologs in the proteins after purification. The results of GST enzyme activity is the same except after purification, which has weak GST enzyme activity, too. All the results indicatd that there may be weak interaction between the two gene of silkworm.
4. The analysis of silkworm CD genes and the intron and phylogenesis of CD
Same to the homologs, most of silkworm CD genes has typical domain of the family which it is affiliated besides CD, except one, BmHP1γ, two CD contained and no CSD. Although the structure and residues composition between CD and CSD is similar, and both composed by 3β-sheets and oneα-helix, the function of them has distinct difference. The special structure of BmHP1γmay be related to chromosome shape of silkworm and the function of it needs further studies.
The results of homologous search indicate that they are more similar to human homologs, especially BmCHD1、BmCHD6、BmHP1α、BmHP1β、Bm-mof, and the similarity to drosophila is lower, which is an interesting thing for the distance in evolution between silkworm(lepidopteran) and drosophila(dipteran) should be less than silkworm and human in general evolutionism, while it is the contrary in this event.
The analysis results of intron of CD genes and CD domain shown that although Su(Var)3-9 and MOF gene of silkworm and drosophila are similar, no intron in them, while homologs of human has. To the CD domain of CHD family and HP1 family of silkworm and human, most of them have. While in drosophila, there are no except HP1α, one intron in the first CD domain, and no intron in the HP1βand HP1γgenes.
引文
[1] Eissenberg J.C. Molecular biology of the chromo domain: an ancient chromatin module comes of age. Gene. 2001, 275: 19-29
[2] Cowell I.G. and Austin C.A. Self-association of chromo domain peptides. Biochim. Biophys. Acta. 1997,1337:198-206
[3] Strutt H. and Paro R. The polycomb group protein complex of Drosophila melanogaster has different compositions at different target genes. Mol. Cell. Biol. 1997,17:6773-6783
[4] Akhtar A., Zink D., and Becker P.B. Chromodomains are protein-RNA interaction modules. Nature. 2000,407:405-409
[5] Bouazoune K., Mitterweger A., Langst G., et al. The dMi-2 chromodomains are DNA binding modules important for ATP-dependent nucleosome mobilization. EMBO J. 2002,21: 2430-2440
[6] Paro R., Hogness D. The polycomb protein shares a homologous domain with a heterochromatin associated protein of Drosophila. Proc. Natl. Acad. Sci. USA. 1991,88:263-267
[7] Breiling A., Bonte E., Ferrari S., et al. The drosophila polycomb protein interacts with nucleosomal core particles in vitro via its repression domain. Mol. Cell. Biol. 1999,19: 8451-8460
[8] Nielsen A.L., Oulad-Abdelghani M., Ortiz J.A., et al. Heterochromatin formation in mammalian cells: interaction between histories and HP1 proteins. Mol. Cell.. 2001,7:729-739
[9] James T.C., Elgin S.C.R. Identification of a nonhistone chromosomal protein associated with heterochromatin in Drosophila and its gene. Mol. Cell. Biol. 1986,6:3862-3872
[10] Eissenberg J.C., James T.C., Foster-Harmett D.M., et al. Mutation in a heterochromatinspecific chromosomal protein is associated with suppression of position-effect variegation in Drosophila melanogaster. Proc. Natl. Acad. Sci. USA. 1990,87:9923-9927
[11] Paro R. Imprinting a determined state into the chromatin of Drosophila. Trends Genet. 1990,6: 416-421
[12] Paro R. Mechanisms of heritable gene repression during development in Drosophila. Curr. Opin. Cell Biol. 1993,5:999-1005
[13] Singh P.B., Miller J.R., Pearce J., et al. Asequece motif found in a drosophila heterochromatin protein is conserved in animals and plants. Nucleic Acids Res. 1991,19:789-794
[14] Epstein H., James T.C., and Singh P.B. Cloning and expression of Drosophila HP1 homologs from a mealybug, Planococcus citri. J. Cell Sci. 1992,101:463-474
[15] Pearce J.J.H., Singh P.B., and Gaunt S.J. The mouce has a polycomb-like chromobox gene. Development. 1992,114:921-929
[16] Saunders W.S., Chue C., Goebl M., et al. Molecular cloning of a human homologue of Drosophila heterochromatin protein HP1 using anti-centromere autoantibodies with anti-chromo specificity. J. Cell Sci. 1993,104:573-582
[17] Lorentz A., Ostermann K., Fleck O., et al. Switching gene swi6 involved in repression of silent mating-type loci in fission yeast, encodes a homologue of chromatin-assoeiated proteins from Drosophila and mammals. Gene. 1994,143:139-143
[18] Aasland R. and Stewart A.F. The chromo shadow domain, a second chromo domain in heterochromatin-binding protein-1, HP-1. Nucleic Acids. Res. 1995,23: 3168-3173
[19] Woodage T., Basrai M. A., Baxevanis A. D., et al. Characterization of the CHD family of proteins. Proc. Natl Acad. Sci. USA 1997,94:11472-11477
[20] Asifa A., Peter B.B. Activation of Transcription through Histone H4 Acetylation by MOF, an Acetyltransferase Essential for Dosage Compensation in Drosophila. Molecular Cell. 2000,2: 367-375
[21] Alessia B., Thomas K., Jop H.K., et al. MOF-Regulated Acetylation of MSL-3 in the Drosophila Dosage Compensation Complex. Molecular Cell. 2003,11:1265-1277
[22] Qi C., Zhu Y., Pan J., et al. Absence of spontaneous peroxisome proliferation in enoyl-CoA Hydratase/L-3-hydroxyacyl-CoA dehydrogenase-deficient mouse liver. J. Biol. Chem. 1999,274: 15775-15780
[23] Lai A., Marcellus R.C., Corbeil H.B. et al. RBP1 induces growth arrest by repression of E2F-dependent transcription. Oncogene. 1999,18:2091-2100
[24] Kim J.K., Huh S.O., Choi H., et al. Srg3, a mouse homolog of yeast SWI3, is essential for early embryogenesis and involved in brain development. Mol. Cell. Biol. 2001,21:7787-7795
[25] Matsumoto-Taniura N., Pirollet F., Monroe R., et al. Identification of novel M phase phosphoproteins by expression cloning. Mol. Biol. Cell. 1996,7:1455-1469
[26] Henikoff S. and Comai L. A DNA methyltransferase homolog with a chromodomain exists in multiple polymorphic forms in Arabidopsis. Genetics. 1998,149:307-318
[27] Marin I. and Llorens C. Ty3/Gypsy retrotransposom: Description of new Arabidopsis thaliana elements and evolutionary perspectives derived from comparative genomic data. Mol. Biol. Evol. 2000,7:1040-1049
[28] Kozak L., Gopal G., Yoon J.H., et al. Elflp, a member of the ABC class of ATPases, functions as a mRNA export factor in Schizosacchromyces pombe. J. Biol. Chem. 2002,277:33580-33589
[29] Hamvas R.M., Reik W., Gaunt S.J., et al. Mapping of a mouse homolog of a heterochromatin protein gene to the X chromosome. Mamm. Genome. 1992,2:72-75
[30] Horseley D., Hutching A., Butcher G.W., et al. M32, a murine homologue of Drosophila heterochromatin protein 1(HP1), localises to euchromatin within interphase nuclei and is largely excluded from constitutive heterochromatin. Cytogenet. Cell Genet. 1996,73:308-311
[31] Le Douarin B., Bour E.V., Zechel C., et al. Ligand-dependent interaction of nuclear receptors with potential transcriptional intermediary factors(mediators) Philos. Trans. R. Soc. Lond. Ser.B 1996,351: 569-578
[32] Ekwall K., Nimmo E.R., Javerzat J.P., et al. Mutation in the fission yeast silencing factors clr4+ and rikl+disrupt the localization of the chromo domain protein Swi6p and impair centromere function, J. Cell Sci. 1996,109:2637-2648
[33] James T.C., Eissenberg J.C., Craig C., et al. 1989. Distribution patterns of HP1, a heterochromatin-associaged nonhistone chromosomal protein of Drosophila. Eur. J. Cell Biol. 1989,50:170-180
[34] Clark R.F., and Elgin S.C.R. heterochromatin protein 1, a known suppressor of position-effect variegation, is highly conserved in drosophila. Nucleic Acids Res. 1992,20:6067-6074
[35] Singh P.B. and Klar A.J. Actie genes in budding yeast display enhanced in vivo probe for chromatin structure of yeast. Genes Dev. 1992,6:186-196
[36] Ekwall K., Javerzat J.P., Lorentz A., et aL The chromodomam protein Swi6: a key component at fission yeast centromeres. Science. 1955,269: 1429-1431
[37] Eissenberg J.C., Morris G.D., Reuter G., et al. The heterochromatin-associated protein HP1 is an essential protein in Drosophila with a dosage-dependent effect on position-effect variegation. Genetics. 1992,131: 345-352
[38] Pak D.T., Pflumm M., Chesnokov I., et al., Association of the origin recognition complex with heterochromatin and HP1 in higher eukaryotes. Cell. 1997, 91: 311-323.
[39] Fanti L., Giovinazzo G., Berloco M., et al. The heteroehromatin protein 1 prevents telomere fusions in Drosophila. Mol. Cell. 1998, 2:527-538
[40] Kellum R. and Alberts B.M. Heterochromatin protein 1 is required for correct chromosome segregation in Drosophila embryos. J. Cell Sci. 1995,108:1419-1431
[41] Zhao T., Heyduk T., Allis C.D. et al. Heterochromatin protein 1 binds to nucleosomes and DNA in vitro. J. Biol. Chem. 2000, 275:28332-28338
[42] Delattre M., Spierer A., Tonka C.H. et al. The genomic silencing of position-effect variegation in Drosophila melanogaster: interaction between the heterochromatin-associated proteins Su(var)3-7 and HP1. J. Cell Sci.2000,113: 4253-4261
[43] Wallrath L. Unfolding the mysteries of heterochromatin. Curr. Opin. Genet.Dev. 1998,8: 147-153
[44] Eissenberg J.C. and Elgin S.C.R. The HP1 protein family: getting a grip on chromatin. Curr. Opin. Genet. Dev. 2000,10:204-210
[45] Pflumm M.E and Botchan M.R. Orc mutants arrest in metaphase with abnormally condensed chromosomes. Development. 2001,128:1697-1707
[46] Vassalo M.F., Tanese N. Isoform-specific interaction of HP1 with human TAFII130. Proc. Natl. Acad. Sci. USA. 2002,99:5919-5924
[47] Zhao T., Heyduk T., Eissenberg J.C. Phosphorylation site mutations in Heterochromatin Protein 1 (HP1) reduce or eliminate silencing activity. J. Biol. Chem. 2001,276:9512-9518
[48] Fanti L., Dorer D.R., Berloco M., et al. Heterochromatin protein 1 binds transgene arrays. Chromosoma. 1998,107:286-292
[49] Bannister A.J., Zegerman P., Partridge J.F., et al. Selective recognition of methylated lysine 9 on histone H3 by the HP1 chromo domain. Nature. 2001,410:120-124
[50] Lachner M., O' Carrol, D., Rea S., et al. Methylation of histone H3 lysine 9 creates a binding site for HP1 proteins. Nature. 2001,410:116-120
[51] Nakayama J., Rice J.C., Strahl B.D., et al. Role of histone H3 lysine 9 methylation in epigenetic control of heterochromatin assembly. Science. 2001,292:110-113
[52] Jacobs S.A., Taverna S.D., Zhang Y., et al. Specificity of the HP1 chromo domain for the methylated N-terminus of historic H3. EMBO J. 2001,20:5232-5241
[53] Platero J.S., Hartnett T., Eissenberg J.C. Functional analysis of the chromo domain of HP1. EMBO J. 1995,14: 3977-3986.
[54] Nielsen P.R., Nietlispach D., Mott H.R., et al. Structure of the HP1 chromodomain bound to histone H3 methylated at lysine 9. Nature. 2002,416:103.
[55] Furuta K., Chart E.K.L., Kiyosawa K., et al. Heterochromatin protein HP1 Hsbeta(p25beta)and its localization with centromeres in mitosis. Chromosoma. 1997,106:11-19
[56] Nicol L. and Jeppesen P. Human autoimmune sera recognize a conserved 26kD protein associated with mammalian heterochromatin that is homologus to heterochromatin protein 1 of Drosophila. Chromosome Res. 1994,2:245-23
[57] Ye Q., and Worman H.J. Interaction between an integral protein of the nuclear envelop inner membrane and human chromodomain proteins homologous to Drosophila HP1. J. Biol. Chem. 1996,271:14653-14656
[58] Kirschmann D.A., Lininger R.A., Gardner L.M., et al. Down-regulation of HP1hs{alpha} expression is associated with the metastatic phenotype in breast cancer. Cancer Res 2000, 60: 3359-63
[59] Tschiersch B.A. Hofmann V. Krauss R. et al. The protein encoded by the Drosophila position-effect variegation suppressor gene Su(var)3-9 combines domains of antagonistic regulators of homeotic gene complexes. EMBO J. 1994, 13: 3822-3831
[60] Ivanova A.V., Bonaduce M.J., Ivanov S.V. et al. The chromo and SET domains of the Clr4 protein are essential for silencing in fission yeast. Nat. Genet. 1998, 9: 192-195.
[61] Aagaard L., Laible G., Selenko P., et al. Functional mammalian homologues of the Drosophila PEV-modifier Su(var)3-9 encode centromere-associated proteins which complex with the heterochromatin component M31.EMBO J. 1999,18:1923-1938
[62] Reuter. Central role of Drosophila SU(VAR)3-9 in histone H3-K9 methylation and heterochromatic gene silencing. EMBO J. 2002,21:1121-1131
[63] Schotta G., Ebert A., Reuter G. SU(VAR)3-9 is a conserved key function in heterochromatic gene silencing. Genetica. 2003,117:149-158
[64] Cleard F., Delattre M., and Spierer P. SU(VAR)3-7, a Drosophila heterochromatin-associated protein and companion of HP1 in the genomic silencing of position-effect variegation. EMBO J. 1997, 16: 5280-5288
[65] Krauss V. and Reuter G. Two Genes Become One: The Genes Encoding Heterochromatin Protein SU(VAR)3-9 and Translation Initiation Factor Subunit eIF-2g Are Joined to a Dicistronic Unit in Holometabolic Insects. Genetics. 2000,156:1157-1167
[66] Melcher M., Schmid M., Aagaard P., et al. Structure-Function Analysis of SUV39H1 Reveals a Dominant Role in Heterochromatin Organization, Chromosome Segregation, and Mitotic Progression Mol. Cell Biol. 2000, 20:3728-3741
[67] Messmer S., Franke A., and Paro R. Analysis of the functional role of the Polycomb chromo domain in Drosophila melanogaster. Genes & Dev. 1992,6:1241-1254
[68] Cao R., Wang L., Wang H., et al. Role of histone H3 lysine 27 methylation in Polycomb-group silencing. Science. 2002,298:1039-1043
[69] Min J., Zhang Y., Xu R.M. Structural basis for specific binding of Polycomb chromodomain to historic H3 methylated at Lys 27. Genes & Dev. 2003,17:1823-1828
[70] Fischle W., Wang Y., Jacobs S.A., et al. Molecular basis for the discrimination of repressive methyl-lysine marks in histone H3 by Polycomb and HP1 chromodomains Genes & Dev. 2003,17:1870-1881
[71] Schwartz, Y.B., Kahn T.G., Nix D.A., et al. Genome-wide analysis of Polycomb targets in Drosophila melanogaster. Nat. Genet. 2006,38:700-705
[72] Rea S., Eisenhaber F., O'Carroll D., et al. Regulation of chromatin structure by site-specific histone H3 methyltransferases. Nature. 2000,406:593-599
[73] Sengupta A.K., Kuhrs A. and Muller J. General transcriptional silencing by a Polycomb response element in Drosophila. Development. 2004,131: 1959-1965
[74] Czermin B., Melfi R., McCabe D., et al. Drosophila enhancer of Zeste/ESC complexes have a histone H3 methyltransferase activity that marks chromosomal Polycomb sites. Cell. 2002,111: 185-196
[75] Kuzmichev A., Nishioka K., Erdjument-Bromage H., et al. Histone methyltransferase activity associated with a human multiprotein complex containing the Enhancer of Zeste protein. Genes & Dev. 2002, 16: 2893-2905
[76] Muller J., Hart C.M., Francis N.J., et al. Histone methyltransferase activity of a Drosophila Polycomb group repressor complex. Cell. 2002, 111: 197-208
[77] Tai H.H., Geisterfer M., Bell J.C., et al. CHD1 associates with NcoR and histone deacetylase as well as with RNA splicing proteins. Biochem Biophy Res. Com. 2003,308:170-176
[78] Hosoi Y., Miyachi H., Matsumoto Y., et al. Induction of interleukin-1 beta and intefleukin-6 mRNA by low doses of ionizing radiation in macrophages. Int J Cancer. 2001,96:270-276
[79] Robson T., Joiner M.C., McCullough W., et al. A novel human stress-related gene with a potential role in induced resisance. Radiat Res. 1999,152:451-461
[80] Tran H.G., Steger D.J., Iyer V.R., et al. The chromo domain protein chdlp from budding yeast is an ATP-dependent chromatin modifying factor. EMBO J. 2000,19:2323-2331
[81] Kelley D.E., Stokes D.G., Perry R.P. CHD1 interacts with SSRP1 and depends on both its chromodamain and its ATPase/helicase-like domain for proper association with chromatin. Chromosoma. 1999,108: 10-25.
[82] Dellaire G., Makarov E.M., Cowger J.J., et al, Mammalian PRP4 kinase copurifies and interacts with components of both the U5 snRNP and the N-CoR deacetylase complexes. Mol. Cell Biol. 2002,22:5141-5156
[83] Thompson P.M., Gotoh T., Kok M., et al. CHD5, a new member of the chromodomain gene family, is preferentially expressed in the nervous system. Oncogene. 2003,22:1002-1011
[84] Krogan N.J., Kim M., Ahn S.H., et al. RNA polymerase Ⅱ elongation factors of Saccharomyces cerevisiae: a targeted proteomics approach. Mol. Cell. Biol. 2002,22:6979-6992
[85] Simic R., Lindstrom D.L., Tran H.G., et al. Chromatin remodeling protein Chd1 interacts with transcription elongation factors and localizes to transcribed genes. EMBO J. 2003,22:1846-1856
[86] Pray-Grant M.G., Daniel J.A., Schieltz D., et al. Chd1 chromodomain links histone H3 methylation with SAGA-and SLIK-dependent acetylation. Nature. 2005,433:434-438
[87] Flanagan J.F., Mi L.Z., Chruszcz M., et al. Double chromodomains cooperate to recognize the methylated histone H3 tail. Nature, 2005,438:1181-1185
[88] Okuda M, Horikoshi M, Nishimura Y. 2007. Structural Polymorphism of Chromodomains in Chd1. J Mol Biol. 365:1047-1062
[89] Kehle J., Beuchle D., Treuheit S., et al. dMi-2, a hunchback-interacting protein that functions in polycomb repression. Science. 1998,282:1897-1900
[90] Seelig H.P., Moosbrugger I., Ehrfeld H., et al. The major dermatomyositis-specific Mi-2 autoantigen is a presumed helicase involved in transcriptional activation. Arthritis Rheum. 1995,38:1389-1399
[91] Nilasena D.S., Trieu E.P., Targoff I.N. Analysis of the Mi-2 autoantigen of dermatomyositis. Arthritis Rheum. 1995,38:123-128
[92] Tong J.K., Hassig C.A., Schnitzler G.R., et al. Chromatin deacetylation by an ATP-dependent nucleosome remodelling complex. Nature. 1998,395:917-921
[93] Bowen N.J., Fujita N., Kajita M., et al. Mi-2/NuRD: multiple complexes for many purposes. Biochem Biiphys Acta. 2004,15:52-57
[94] Wade P.A., Gegonne A., Jones P.J., et al. Mi-2 complex couples DNA methylation to chromatin remodelling and histone deacetylation. Nat Genet. 1999,23:62-66
[95] Zhang Y., Ng H.H., Erdjument-Bromage H., et al. Analysis of the NuRD subunits reveals a histone deacetylase core complex and a connection with DNA methylation. Genes Dev. 1999,13: 1924-1935
[96] Guschin D., Wade P.A., Kikyo N., et al. ATP-Dependent histone octamer mobilization and histone deacetylation mediated by the Mi-2 chromatin remodeling complex. Biochemistry. 2000, 39:5238-5245
[97] Shimono Y., Murakami H., Kawai K., et al. Mi-2 beta associates with BRG1 and RET finger protein at the distinct regions with transcriptional activating and repressing abilities. J Biol. Chem. 2003, 278:51638-51645
[98] Khorasanizadeh S. The nucleosome: from genomic organization to genomic regulation. Cell. 2004, 116: 259-272
[99] 孙志增，徐勤枝，隋建丽，等．辐射诱导转录因子RIGb cDNA对Hela细胞增殖的抑制作用．中国生物化学与分子生物学报．2005，21：384-389
[100] 周平坤，隋建丽，耿煜，等．辐射诱导基因LRIGx的细胞周期特异性及编码产物同源性分析表达．中国生物化学与分子生物学报．2002，18：272-276
[101] Loidl P. Historic acetylation: facts and questions. Chromosoma. 1994.103:441-449
[102] Fletcher T.M., Hansen J.C. Core histone tail domains mediate oligonucleosome folding and nucleosomal DNA organization through distinct molecular mechanisms. J. Biol. Chem. 1995,270:25356-25362
[103] Luger K., Mader A.W., Richmond R.K., et al. Crystal structure of the nucleosome core particle at 2.8 A resolution. Nature. 1997,389:251-260
[104] Hong L., Sehroth G.P., Matthews H.R., et al. Studies of the DNA binding properties of histone H4 amino terminus: thermal denaturation studies reveal that acetylation markedly reduces the binding constant of the H4 "tail" to DNA. J. Biol. Chem. 1993,268:305-314
[105] Steger D.J., Workman J.L. Remodeling chromatin structures for transcription: what happens to the histones? BioEssays. 1996,18:875-884
[106] Fletcher T.M., Hansen J.C. The nucleosomal array: structure/function relationships. Crit. Rev. Eukaryot. Gene Expression. 1996,6:149-188
[107] Luger K., Richmond T.J. The histone tails of the nucleosome. Curr. Opin. Genet. Dev. 1998,8: 140-146
[108] Norton V.G., Imai B.S., Yau P., et al. Histone acetylation reduces nucleosome core particle linking number change. Cell. 1989,57:449-457
[109] Hebbes T.R., Thorne A.W., Crane-Robinson C. A direct link between core histone acetylation and transcriptionally active chromatin. EMBO J. 1988,7:1395-1402
[110] Sterner R., Vidali G., Allfrey V.G. Studies of acetylation and deacetylation in high mobility group proteins: identification of the sites of acetylation in high mobility group proteins 14 and 17. J. Biol. Chem. 1981,256:8892-8895
[111] Arri E., Rhea T.U., Amine N., et al. The yeast NuA4 and Drosophila MSL complexes contain homologous subunits important for transcription regulation. Biol. Chem. 2001,276:3484-3491
[112] Edwin R.S., Christelle C., Rong H., et al. A Human Protein Complex Homologous to the Drosophila MSL Complex Is Responsible for the Majority of Histone H4 Aeetylation at Lysine 16. Mol. Cell. Biol, 2005.25:9175-9188
[113] Yannick D., William S., William S.L., et al. Structural and Functional Conservation of the NuA4 Histone Acetyltransferase Complex from Yeast to Human. Mol. Cell Biol. 2004, 24: 1884-1896
[114] Edwin R.S., Antonio P., Weigang G., et al. The Drosophila MSL Complex Aeetylates Histone H4 at Lysine 16, a Chromatin Modification Linked to Dosage Compensation. Mol. Cell Biol. 2000,20:312-318
[115] Muller H.J. Further studies on the nature and causes of gene mutations. Proc. Ⅵ Int. Congr. Genet. 1932,1: 213-255
[116] Muller H.J. Evidence of the precision of genetic adaptation. Harvey Lect. Ser. 1950, ⅩLⅢ: 165-229
[117] Baker B.S., Gorman M. Marin I. Dosage compensation in Drosophila. Annu. Rev. Genet. 1994, 28:491-521
[118] Hilfiker A., Hilfiker-Kleiner D., Pannuti A. mof, a putative acetyl transferase gene related to the Tip60 and MOZ human genes and to the SAS genes of yeast, is required for dosage compensation in Drosophila. EMBO J. 1997,16:2054-2060
[119] Fukunaga A., Tanaka A. Oishi A. Maleless, a recessive autosomal mutant of Drosophila melanogaster that specifically kills male zygotes. Genetics. 1975,81:135-141
[120] Belote J.M. Lucchesi J.C. Sex determination in Drosophila melanogaster: from the X:A ratio to doublesex. Genetics. 1980a,96:165-186
[121] Uchida S., Uenoyama T., Oishi K. Studies on the sex-specific lethals of Drosophila melanogaster. Ⅲ. A third chromosome malespecific lethal mutant. Jpn. J Genet. 1981,56: 523-527
[122] Lucchesi J.C., Skripsky T. Tax F.E. A new male-specific lethal mutation in Drosophila melanogaster. Genetics. 1982,100:s42
[123] Breen T.R. Lucchesi J.C. Analysis of the dosage compensation of a specific transcript in Drosophila melanogaster. Genetics. 1986,112:483-491
[124] Kelley R.L., Kuroda M.I. Equality for X chromosomes. Science. 1995,270:1607-1610
[125] Bone J.R., Lavender J., Richman R., et al. Acetylated histone H4 on the male X-chromosome is associated with dosage compensation in Drosophila. Genes Dev. 1994,8:96-104
[126] BAKER B.S., GORMAN M., MARIN I. Dosage compensation in Drosophila. Annu. Rev. Genet. 1994,28:491-521
[127] LUCCHESI J.C. Dosage compensation in flies and worms: the ups and downs of X-chromosome regulation. Curr. Opin. Genet. Dev. 1998,8:179-184
[128] FRANKE A., BAKER B.S. The roxl and rox2 RNAs are essential components of the "compensasome", which mediates dosage compensation in Drosophila. Mol. Cell. 1999,4: 117-122
[129] KELLEY R.L., KURODA M.I. The role of chromosomal RNAs in marking the X for dosage compensation. Curr. Opin. Genet. Dev. 2000,10:555-561
[130] Kuo M.H., Allis C.D. Roles of histone acetyltransferases and deacetylases in gene regulation. Bioessays. 1998,20:615-626
[131] Pazin M., Kadonaga J. What's up and down with histone deacetylation and transcription? Cell. 1997,89:325-328
[132] Astrid S.C., Joanna E.L., Sandra J.J., et al. Esalp is an essential histone acetyltransferase required for cell cycle progression. Mol. Cell. Biol. 1999,19:2515-2526
[133] Mikko T., Stephen R., Karsten R., et al. hMOF histone acetyltmnsferase is required for histone H4 lysine 16 acetylation in mammalian cells, Mol. Cell Biol. 2005,25:6798-810
[134] Fraga M.F., Ballestar E., Villar-Garea A., et al. Loss of acetylation at Lys 16 and trimethylation at Lys20 of histone H4 is a common hallmark of human cancer. Nat. Genet. 2005,37:391-400
[135] Ikura T., Ogryzko V.V., Grigoriev M., et al. Involvement of the TIP60 histone acetylase complex in DNA repair and apoptosis. Cell. 2000102:463-473
[136] Kamine J., Elangovan B., Subramanian T., et al. Identification of a cellular protein that specifically interacts with the essential cysteine region of the HⅣ-1 Tat transactivator. Virology. 1996,216:357-366
[137] Kusch T., Florens L., Macdonald W.H., et al. Acetylation by Tip60 Is Required for Selective Histone Variant Exchange at DNA Lesions. Science. 2004,306:2084-2087
[138] Yochum G.S., Ayer D.E. Role for the Mortality Factors MORF4, MRGX, and MRG15 in Transcriptional Repression via Associations with Pfl, mSin3A, and Transduein-Like Enhancer of Split. Mol. Cell. Biol. 2002,22:7868-7876
[139] Matin I., Baker B.S. Origin and Evolution of the Regulatory Gene male-specific lethal-3. Mol. Biol. Evol. 2000,17:1240-1250
[140] Pardo P.S., Leung J.K., Lucchesi. C., et aL MRG15, a Novel Chromodomain Protein, Is Present in Two Distinct Multiprotein Complexes Involved in Transcriptional Activation. J Biol. Chem. 2002,277:50860-50866
[141] Zhang P., Zhao J.Y., Wang B., et al. The MRG domain of hunman MRG15 uses a shallow hydrophobic pocket to interact with the N-terminal region of PAM14. Protein Science. 2006,15: 2423-2434
[142] Ball L.J., Murzina N.V., Broadhurst R.W., et al. Structure of the chromatin binding (chromo) domain from mouse modifier protein 1. EMBO J. 1997,16:2473-2481
[143] Fields S., Song O. A Novel genetic system to detect protein-protein intersctions. Nature. 1989, 340:245-246
[144] 赵永芳．生物化学技术原理及应用．北京：科学出版社．2002：7-82
[145] Kaelin W.G., Pallas D.C., DeCaprio J.A., et al. Identification of cellular proteins that can interact specifically with the T/E1A-binding region of the retinoblastoma gene product. Cell. 1991,64:521-532
[146] Harlow E. and Lane D. Antibodies: A laboratory manual Cold spring Harbor Laboratory, Cold Spring Harbor, New York.
[147] Harlow E. and Lane D. Using antibodies: A laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York.
[148] Edmondson S.P., Qiu L., Shriver J.W. Solution structure of the DNA-binding protein Sac7d from the hyperthermophile Sulfolobus acidocaldarius. Biochemistry. 1995,34:13289-13304
[149] Riddihough G., Pennisi E. The Evolution of Epigenetics. Science.2001,293:1063
[150] Madireddi M.T., Coyne R.S., Smothers J.F., et al. Pddlp, a novel chromodomain-containing protein, links heterochromatin assembly and DNA elimination in Tetrahymena.Cell.1996,87: 75-84
[151] Xia Q.Y., Zhou Z.Y., Lu C., et al. A Draft Sequence for the Genome of the Domesticated Silkworm (Bombyx mori). Science. 2004, 306:1937-1940
[152] Mita K., Kasahara M., Sasaki S., et al. The Genome Sequence of Silkworm, Bombyx mori DNA Research. 2004,11:27-35
[153] Isobe M., Kai H., Kurahashi T., et al. The Molecular Mechanism of the Termination of Insect Diapause, Part 1: A Timer Protein, TIME-EA4, in the Diapause Eggs of the Silkworm, Bombyx mori is a Metallo-Glycoprotein. Chembiochem. 2006, 7:1590-1598.
[154] MiaoX.X., Xu S.J., Li M.H., et al. Simple sequence repeat-based consensus linkage map of Bombyx mori. PNAS. 2005, 102:16303-16308
[155] Shilatifard A. Chromatin Modifications by Methylation and Ubiquitination: Implications in the Regulation of Gene Expression. Ann. Rev. Biochem. 2006, 75: 243～269.
[156] Suzuki M.G., Shimada T., Kobayashi M. Bm kettin, homologue of the Drosophila kettin gene, is located on the Z chromosome in Bombyx mori and is not dosage compensated. Heredity. 1999, 82:170-179
[157] Koike Y., Mita K., Suzuki M.G., et al. Genomic sequence of a 320-kb segment of the Z chromosome of Bombyx mori containing a kettin ortholog. Mol Gen Genomics. 2003,269: 137-149
[158] Marin I. Evolution of chromatin-remodeling complexes: comparative genomics reveals the ancient origin of "novel" compensasome genes. J Mol. Evol. 2003,56:527-539
[159] Gorinsek F.B., Gubensek K.D. Phylogenomic analysis of chromoviruses. Cytogenet Genome Res. 2005, 110:543-552
[2] Cowell I.G. and Austin C.A. Self-association of chromo domain peptides. Biochim. Biophys. Acta. 1997,1337:198-206
[3] Strutt H. and Paro R. The polycomb group protein complex of Drosophila melanogaster has different compositions at different target genes. Mol. Cell. Biol. 1997,17:6773-6783
[4] Akhtar A., Zink D., and Becker P.B. Chromodomains are protein-RNA interaction modules. Nature. 2000,407:405-409
[5] Bouazoune K., Mitterweger A., Langst G., et al. The dMi-2 chromodomains are DNA binding modules important for ATP-dependent nucleosome mobilization. EMBO J. 2002,21: 2430-2440
[6] Paro R., Hogness D. The polycomb protein shares a homologous domain with a heterochromatin associated protein of Drosophila. Proc. Natl. Acad. Sci. USA. 1991,88:263-267
[7] Breiling A., Bonte E., Ferrari S., et al. The drosophila polycomb protein interacts with nucleosomal core particles in vitro via its repression domain. Mol. Cell. Biol. 1999,19: 8451-8460
[8] Nielsen A.L., Oulad-Abdelghani M., Ortiz J.A., et al. Heterochromatin formation in mammalian cells: interaction between histories and HP1 proteins. Mol. Cell.. 2001,7:729-739
[9] James T.C., Elgin S.C.R. Identification of a nonhistone chromosomal protein associated with heterochromatin in Drosophila and its gene. Mol. Cell. Biol. 1986,6:3862-3872
[10] Eissenberg J.C., James T.C., Foster-Harmett D.M., et al. Mutation in a heterochromatinspecific chromosomal protein is associated with suppression of position-effect variegation in Drosophila melanogaster. Proc. Natl. Acad. Sci. USA. 1990,87:9923-9927
[11] Paro R. Imprinting a determined state into the chromatin of Drosophila. Trends Genet. 1990,6: 416-421
[12] Paro R. Mechanisms of heritable gene repression during development in Drosophila. Curr. Opin. Cell Biol. 1993,5:999-1005
[13] Singh P.B., Miller J.R., Pearce J., et al. Asequece motif found in a drosophila heterochromatin protein is conserved in animals and plants. Nucleic Acids Res. 1991,19:789-794
[14] Epstein H., James T.C., and Singh P.B. Cloning and expression of Drosophila HP1 homologs from a mealybug, Planococcus citri. J. Cell Sci. 1992,101:463-474
[15] Pearce J.J.H., Singh P.B., and Gaunt S.J. The mouce has a polycomb-like chromobox gene. Development. 1992,114:921-929
[16] Saunders W.S., Chue C., Goebl M., et al. Molecular cloning of a human homologue of Drosophila heterochromatin protein HP1 using anti-centromere autoantibodies with anti-chromo specificity. J. Cell Sci. 1993,104:573-582
[17] Lorentz A., Ostermann K., Fleck O., et al. Switching gene swi6 involved in repression of silent mating-type loci in fission yeast, encodes a homologue of chromatin-assoeiated proteins from Drosophila and mammals. Gene. 1994,143:139-143
[18] Aasland R. and Stewart A.F. The chromo shadow domain, a second chromo domain in heterochromatin-binding protein-1, HP-1. Nucleic Acids. Res. 1995,23: 3168-3173
[19] Woodage T., Basrai M. A., Baxevanis A. D., et al. Characterization of the CHD family of proteins. Proc. Natl Acad. Sci. USA 1997,94:11472-11477
[20] Asifa A., Peter B.B. Activation of Transcription through Histone H4 Acetylation by MOF, an Acetyltransferase Essential for Dosage Compensation in Drosophila. Molecular Cell. 2000,2: 367-375
[21] Alessia B., Thomas K., Jop H.K., et al. MOF-Regulated Acetylation of MSL-3 in the Drosophila Dosage Compensation Complex. Molecular Cell. 2003,11:1265-1277
[22] Qi C., Zhu Y., Pan J., et al. Absence of spontaneous peroxisome proliferation in enoyl-CoA Hydratase/L-3-hydroxyacyl-CoA dehydrogenase-deficient mouse liver. J. Biol. Chem. 1999,274: 15775-15780
[23] Lai A., Marcellus R.C., Corbeil H.B. et al. RBP1 induces growth arrest by repression of E2F-dependent transcription. Oncogene. 1999,18:2091-2100
[24] Kim J.K., Huh S.O., Choi H., et al. Srg3, a mouse homolog of yeast SWI3, is essential for early embryogenesis and involved in brain development. Mol. Cell. Biol. 2001,21:7787-7795
[25] Matsumoto-Taniura N., Pirollet F., Monroe R., et al. Identification of novel M phase phosphoproteins by expression cloning. Mol. Biol. Cell. 1996,7:1455-1469
[26] Henikoff S. and Comai L. A DNA methyltransferase homolog with a chromodomain exists in multiple polymorphic forms in Arabidopsis. Genetics. 1998,149:307-318
[27] Marin I. and Llorens C. Ty3/Gypsy retrotransposom: Description of new Arabidopsis thaliana elements and evolutionary perspectives derived from comparative genomic data. Mol. Biol. Evol. 2000,7:1040-1049
[28] Kozak L., Gopal G., Yoon J.H., et al. Elflp, a member of the ABC class of ATPases, functions as a mRNA export factor in Schizosacchromyces pombe. J. Biol. Chem. 2002,277:33580-33589
[29] Hamvas R.M., Reik W., Gaunt S.J., et al. Mapping of a mouse homolog of a heterochromatin protein gene to the X chromosome. Mamm. Genome. 1992,2:72-75
[30] Horseley D., Hutching A., Butcher G.W., et al. M32, a murine homologue of Drosophila heterochromatin protein 1(HP1), localises to euchromatin within interphase nuclei and is largely excluded from constitutive heterochromatin. Cytogenet. Cell Genet. 1996,73:308-311
[31] Le Douarin B., Bour E.V., Zechel C., et al. Ligand-dependent interaction of nuclear receptors with potential transcriptional intermediary factors(mediators) Philos. Trans. R. Soc. Lond. Ser.B 1996,351: 569-578
[32] Ekwall K., Nimmo E.R., Javerzat J.P., et al. Mutation in the fission yeast silencing factors clr4+ and rikl+disrupt the localization of the chromo domain protein Swi6p and impair centromere function, J. Cell Sci. 1996,109:2637-2648
[33] James T.C., Eissenberg J.C., Craig C., et al. 1989. Distribution patterns of HP1, a heterochromatin-associaged nonhistone chromosomal protein of Drosophila. Eur. J. Cell Biol. 1989,50:170-180
[34] Clark R.F., and Elgin S.C.R. heterochromatin protein 1, a known suppressor of position-effect variegation, is highly conserved in drosophila. Nucleic Acids Res. 1992,20:6067-6074
[35] Singh P.B. and Klar A.J. Actie genes in budding yeast display enhanced in vivo probe for chromatin structure of yeast. Genes Dev. 1992,6:186-196
[36] Ekwall K., Javerzat J.P., Lorentz A., et aL The chromodomam protein Swi6: a key component at fission yeast centromeres. Science. 1955,269: 1429-1431
[37] Eissenberg J.C., Morris G.D., Reuter G., et al. The heterochromatin-associated protein HP1 is an essential protein in Drosophila with a dosage-dependent effect on position-effect variegation. Genetics. 1992,131: 345-352
[38] Pak D.T., Pflumm M., Chesnokov I., et al., Association of the origin recognition complex with heterochromatin and HP1 in higher eukaryotes. Cell. 1997, 91: 311-323.
[39] Fanti L., Giovinazzo G., Berloco M., et al. The heteroehromatin protein 1 prevents telomere fusions in Drosophila. Mol. Cell. 1998, 2:527-538
[40] Kellum R. and Alberts B.M. Heterochromatin protein 1 is required for correct chromosome segregation in Drosophila embryos. J. Cell Sci. 1995,108:1419-1431
[41] Zhao T., Heyduk T., Allis C.D. et al. Heterochromatin protein 1 binds to nucleosomes and DNA in vitro. J. Biol. Chem. 2000, 275:28332-28338
[42] Delattre M., Spierer A., Tonka C.H. et al. The genomic silencing of position-effect variegation in Drosophila melanogaster: interaction between the heterochromatin-associated proteins Su(var)3-7 and HP1. J. Cell Sci.2000,113: 4253-4261
[43] Wallrath L. Unfolding the mysteries of heterochromatin. Curr. Opin. Genet.Dev. 1998,8: 147-153
[44] Eissenberg J.C. and Elgin S.C.R. The HP1 protein family: getting a grip on chromatin. Curr. Opin. Genet. Dev. 2000,10:204-210
[45] Pflumm M.E and Botchan M.R. Orc mutants arrest in metaphase with abnormally condensed chromosomes. Development. 2001,128:1697-1707
[46] Vassalo M.F., Tanese N. Isoform-specific interaction of HP1 with human TAFII130. Proc. Natl. Acad. Sci. USA. 2002,99:5919-5924
[47] Zhao T., Heyduk T., Eissenberg J.C. Phosphorylation site mutations in Heterochromatin Protein 1 (HP1) reduce or eliminate silencing activity. J. Biol. Chem. 2001,276:9512-9518
[48] Fanti L., Dorer D.R., Berloco M., et al. Heterochromatin protein 1 binds transgene arrays. Chromosoma. 1998,107:286-292
[49] Bannister A.J., Zegerman P., Partridge J.F., et al. Selective recognition of methylated lysine 9 on histone H3 by the HP1 chromo domain. Nature. 2001,410:120-124
[50] Lachner M., O' Carrol, D., Rea S., et al. Methylation of histone H3 lysine 9 creates a binding site for HP1 proteins. Nature. 2001,410:116-120
[51] Nakayama J., Rice J.C., Strahl B.D., et al. Role of histone H3 lysine 9 methylation in epigenetic control of heterochromatin assembly. Science. 2001,292:110-113
[52] Jacobs S.A., Taverna S.D., Zhang Y., et al. Specificity of the HP1 chromo domain for the methylated N-terminus of historic H3. EMBO J. 2001,20:5232-5241
[53] Platero J.S., Hartnett T., Eissenberg J.C. Functional analysis of the chromo domain of HP1. EMBO J. 1995,14: 3977-3986.
[54] Nielsen P.R., Nietlispach D., Mott H.R., et al. Structure of the HP1 chromodomain bound to histone H3 methylated at lysine 9. Nature. 2002,416:103.
[55] Furuta K., Chart E.K.L., Kiyosawa K., et al. Heterochromatin protein HP1 Hsbeta(p25beta)and its localization with centromeres in mitosis. Chromosoma. 1997,106:11-19
[56] Nicol L. and Jeppesen P. Human autoimmune sera recognize a conserved 26kD protein associated with mammalian heterochromatin that is homologus to heterochromatin protein 1 of Drosophila. Chromosome Res. 1994,2:245-23
[57] Ye Q., and Worman H.J. Interaction between an integral protein of the nuclear envelop inner membrane and human chromodomain proteins homologous to Drosophila HP1. J. Biol. Chem. 1996,271:14653-14656
[58] Kirschmann D.A., Lininger R.A., Gardner L.M., et al. Down-regulation of HP1hs{alpha} expression is associated with the metastatic phenotype in breast cancer. Cancer Res 2000, 60: 3359-63
[59] Tschiersch B.A. Hofmann V. Krauss R. et al. The protein encoded by the Drosophila position-effect variegation suppressor gene Su(var)3-9 combines domains of antagonistic regulators of homeotic gene complexes. EMBO J. 1994, 13: 3822-3831
[60] Ivanova A.V., Bonaduce M.J., Ivanov S.V. et al. The chromo and SET domains of the Clr4 protein are essential for silencing in fission yeast. Nat. Genet. 1998, 9: 192-195.
[61] Aagaard L., Laible G., Selenko P., et al. Functional mammalian homologues of the Drosophila PEV-modifier Su(var)3-9 encode centromere-associated proteins which complex with the heterochromatin component M31.EMBO J. 1999,18:1923-1938
[62] Reuter. Central role of Drosophila SU(VAR)3-9 in histone H3-K9 methylation and heterochromatic gene silencing. EMBO J. 2002,21:1121-1131
[63] Schotta G., Ebert A., Reuter G. SU(VAR)3-9 is a conserved key function in heterochromatic gene silencing. Genetica. 2003,117:149-158
[64] Cleard F., Delattre M., and Spierer P. SU(VAR)3-7, a Drosophila heterochromatin-associated protein and companion of HP1 in the genomic silencing of position-effect variegation. EMBO J. 1997, 16: 5280-5288
[65] Krauss V. and Reuter G. Two Genes Become One: The Genes Encoding Heterochromatin Protein SU(VAR)3-9 and Translation Initiation Factor Subunit eIF-2g Are Joined to a Dicistronic Unit in Holometabolic Insects. Genetics. 2000,156:1157-1167
[66] Melcher M., Schmid M., Aagaard P., et al. Structure-Function Analysis of SUV39H1 Reveals a Dominant Role in Heterochromatin Organization, Chromosome Segregation, and Mitotic Progression Mol. Cell Biol. 2000, 20:3728-3741
[67] Messmer S., Franke A., and Paro R. Analysis of the functional role of the Polycomb chromo domain in Drosophila melanogaster. Genes & Dev. 1992,6:1241-1254
[68] Cao R., Wang L., Wang H., et al. Role of histone H3 lysine 27 methylation in Polycomb-group silencing. Science. 2002,298:1039-1043
[69] Min J., Zhang Y., Xu R.M. Structural basis for specific binding of Polycomb chromodomain to historic H3 methylated at Lys 27. Genes & Dev. 2003,17:1823-1828
[70] Fischle W., Wang Y., Jacobs S.A., et al. Molecular basis for the discrimination of repressive methyl-lysine marks in histone H3 by Polycomb and HP1 chromodomains Genes & Dev. 2003,17:1870-1881
[71] Schwartz, Y.B., Kahn T.G., Nix D.A., et al. Genome-wide analysis of Polycomb targets in Drosophila melanogaster. Nat. Genet. 2006,38:700-705
[72] Rea S., Eisenhaber F., O'Carroll D., et al. Regulation of chromatin structure by site-specific histone H3 methyltransferases. Nature. 2000,406:593-599
[73] Sengupta A.K., Kuhrs A. and Muller J. General transcriptional silencing by a Polycomb response element in Drosophila. Development. 2004,131: 1959-1965
[74] Czermin B., Melfi R., McCabe D., et al. Drosophila enhancer of Zeste/ESC complexes have a histone H3 methyltransferase activity that marks chromosomal Polycomb sites. Cell. 2002,111: 185-196
[75] Kuzmichev A., Nishioka K., Erdjument-Bromage H., et al. Histone methyltransferase activity associated with a human multiprotein complex containing the Enhancer of Zeste protein. Genes & Dev. 2002, 16: 2893-2905
[76] Muller J., Hart C.M., Francis N.J., et al. Histone methyltransferase activity of a Drosophila Polycomb group repressor complex. Cell. 2002, 111: 197-208
[77] Tai H.H., Geisterfer M., Bell J.C., et al. CHD1 associates with NcoR and histone deacetylase as well as with RNA splicing proteins. Biochem Biophy Res. Com. 2003,308:170-176
[78] Hosoi Y., Miyachi H., Matsumoto Y., et al. Induction of interleukin-1 beta and intefleukin-6 mRNA by low doses of ionizing radiation in macrophages. Int J Cancer. 2001,96:270-276
[79] Robson T., Joiner M.C., McCullough W., et al. A novel human stress-related gene with a potential role in induced resisance. Radiat Res. 1999,152:451-461
[80] Tran H.G., Steger D.J., Iyer V.R., et al. The chromo domain protein chdlp from budding yeast is an ATP-dependent chromatin modifying factor. EMBO J. 2000,19:2323-2331
[81] Kelley D.E., Stokes D.G., Perry R.P. CHD1 interacts with SSRP1 and depends on both its chromodamain and its ATPase/helicase-like domain for proper association with chromatin. Chromosoma. 1999,108: 10-25.
[82] Dellaire G., Makarov E.M., Cowger J.J., et al, Mammalian PRP4 kinase copurifies and interacts with components of both the U5 snRNP and the N-CoR deacetylase complexes. Mol. Cell Biol. 2002,22:5141-5156
[83] Thompson P.M., Gotoh T., Kok M., et al. CHD5, a new member of the chromodomain gene family, is preferentially expressed in the nervous system. Oncogene. 2003,22:1002-1011
[84] Krogan N.J., Kim M., Ahn S.H., et al. RNA polymerase Ⅱ elongation factors of Saccharomyces cerevisiae: a targeted proteomics approach. Mol. Cell. Biol. 2002,22:6979-6992
[85] Simic R., Lindstrom D.L., Tran H.G., et al. Chromatin remodeling protein Chd1 interacts with transcription elongation factors and localizes to transcribed genes. EMBO J. 2003,22:1846-1856
[86] Pray-Grant M.G., Daniel J.A., Schieltz D., et al. Chd1 chromodomain links histone H3 methylation with SAGA-and SLIK-dependent acetylation. Nature. 2005,433:434-438
[87] Flanagan J.F., Mi L.Z., Chruszcz M., et al. Double chromodomains cooperate to recognize the methylated histone H3 tail. Nature, 2005,438:1181-1185
[88] Okuda M, Horikoshi M, Nishimura Y. 2007. Structural Polymorphism of Chromodomains in Chd1. J Mol Biol. 365:1047-1062
[89] Kehle J., Beuchle D., Treuheit S., et al. dMi-2, a hunchback-interacting protein that functions in polycomb repression. Science. 1998,282:1897-1900
[90] Seelig H.P., Moosbrugger I., Ehrfeld H., et al. The major dermatomyositis-specific Mi-2 autoantigen is a presumed helicase involved in transcriptional activation. Arthritis Rheum. 1995,38:1389-1399
[91] Nilasena D.S., Trieu E.P., Targoff I.N. Analysis of the Mi-2 autoantigen of dermatomyositis. Arthritis Rheum. 1995,38:123-128
[92] Tong J.K., Hassig C.A., Schnitzler G.R., et al. Chromatin deacetylation by an ATP-dependent nucleosome remodelling complex. Nature. 1998,395:917-921
[93] Bowen N.J., Fujita N., Kajita M., et al. Mi-2/NuRD: multiple complexes for many purposes. Biochem Biiphys Acta. 2004,15:52-57
[94] Wade P.A., Gegonne A., Jones P.J., et al. Mi-2 complex couples DNA methylation to chromatin remodelling and histone deacetylation. Nat Genet. 1999,23:62-66
[95] Zhang Y., Ng H.H., Erdjument-Bromage H., et al. Analysis of the NuRD subunits reveals a histone deacetylase core complex and a connection with DNA methylation. Genes Dev. 1999,13: 1924-1935
[96] Guschin D., Wade P.A., Kikyo N., et al. ATP-Dependent histone octamer mobilization and histone deacetylation mediated by the Mi-2 chromatin remodeling complex. Biochemistry. 2000, 39:5238-5245
[97] Shimono Y., Murakami H., Kawai K., et al. Mi-2 beta associates with BRG1 and RET finger protein at the distinct regions with transcriptional activating and repressing abilities. J Biol. Chem. 2003, 278:51638-51645
[98] Khorasanizadeh S. The nucleosome: from genomic organization to genomic regulation. Cell. 2004, 116: 259-272
[99] 孙志增，徐勤枝，隋建丽，等．辐射诱导转录因子RIGb cDNA对Hela细胞增殖的抑制作用．中国生物化学与分子生物学报．2005，21：384-389
[100] 周平坤，隋建丽，耿煜，等．辐射诱导基因LRIGx的细胞周期特异性及编码产物同源性分析表达．中国生物化学与分子生物学报．2002，18：272-276
[101] Loidl P. Historic acetylation: facts and questions. Chromosoma. 1994.103:441-449
[102] Fletcher T.M., Hansen J.C. Core histone tail domains mediate oligonucleosome folding and nucleosomal DNA organization through distinct molecular mechanisms. J. Biol. Chem. 1995,270:25356-25362
[103] Luger K., Mader A.W., Richmond R.K., et al. Crystal structure of the nucleosome core particle at 2.8 A resolution. Nature. 1997,389:251-260
[104] Hong L., Sehroth G.P., Matthews H.R., et al. Studies of the DNA binding properties of histone H4 amino terminus: thermal denaturation studies reveal that acetylation markedly reduces the binding constant of the H4 "tail" to DNA. J. Biol. Chem. 1993,268:305-314
[105] Steger D.J., Workman J.L. Remodeling chromatin structures for transcription: what happens to the histones? BioEssays. 1996,18:875-884
[106] Fletcher T.M., Hansen J.C. The nucleosomal array: structure/function relationships. Crit. Rev. Eukaryot. Gene Expression. 1996,6:149-188
[107] Luger K., Richmond T.J. The histone tails of the nucleosome. Curr. Opin. Genet. Dev. 1998,8: 140-146
[108] Norton V.G., Imai B.S., Yau P., et al. Histone acetylation reduces nucleosome core particle linking number change. Cell. 1989,57:449-457
[109] Hebbes T.R., Thorne A.W., Crane-Robinson C. A direct link between core histone acetylation and transcriptionally active chromatin. EMBO J. 1988,7:1395-1402
[110] Sterner R., Vidali G., Allfrey V.G. Studies of acetylation and deacetylation in high mobility group proteins: identification of the sites of acetylation in high mobility group proteins 14 and 17. J. Biol. Chem. 1981,256:8892-8895
[111] Arri E., Rhea T.U., Amine N., et al. The yeast NuA4 and Drosophila MSL complexes contain homologous subunits important for transcription regulation. Biol. Chem. 2001,276:3484-3491
[112] Edwin R.S., Christelle C., Rong H., et al. A Human Protein Complex Homologous to the Drosophila MSL Complex Is Responsible for the Majority of Histone H4 Aeetylation at Lysine 16. Mol. Cell. Biol, 2005.25:9175-9188
[113] Yannick D., William S., William S.L., et al. Structural and Functional Conservation of the NuA4 Histone Acetyltransferase Complex from Yeast to Human. Mol. Cell Biol. 2004, 24: 1884-1896
[114] Edwin R.S., Antonio P., Weigang G., et al. The Drosophila MSL Complex Aeetylates Histone H4 at Lysine 16, a Chromatin Modification Linked to Dosage Compensation. Mol. Cell Biol. 2000,20:312-318
[115] Muller H.J. Further studies on the nature and causes of gene mutations. Proc. Ⅵ Int. Congr. Genet. 1932,1: 213-255
[116] Muller H.J. Evidence of the precision of genetic adaptation. Harvey Lect. Ser. 1950, ⅩLⅢ: 165-229
[117] Baker B.S., Gorman M. Marin I. Dosage compensation in Drosophila. Annu. Rev. Genet. 1994, 28:491-521
[118] Hilfiker A., Hilfiker-Kleiner D., Pannuti A. mof, a putative acetyl transferase gene related to the Tip60 and MOZ human genes and to the SAS genes of yeast, is required for dosage compensation in Drosophila. EMBO J. 1997,16:2054-2060
[119] Fukunaga A., Tanaka A. Oishi A. Maleless, a recessive autosomal mutant of Drosophila melanogaster that specifically kills male zygotes. Genetics. 1975,81:135-141
[120] Belote J.M. Lucchesi J.C. Sex determination in Drosophila melanogaster: from the X:A ratio to doublesex. Genetics. 1980a,96:165-186
[121] Uchida S., Uenoyama T., Oishi K. Studies on the sex-specific lethals of Drosophila melanogaster. Ⅲ. A third chromosome malespecific lethal mutant. Jpn. J Genet. 1981,56: 523-527
[122] Lucchesi J.C., Skripsky T. Tax F.E. A new male-specific lethal mutation in Drosophila melanogaster. Genetics. 1982,100:s42
[123] Breen T.R. Lucchesi J.C. Analysis of the dosage compensation of a specific transcript in Drosophila melanogaster. Genetics. 1986,112:483-491
[124] Kelley R.L., Kuroda M.I. Equality for X chromosomes. Science. 1995,270:1607-1610
[125] Bone J.R., Lavender J., Richman R., et al. Acetylated histone H4 on the male X-chromosome is associated with dosage compensation in Drosophila. Genes Dev. 1994,8:96-104
[126] BAKER B.S., GORMAN M., MARIN I. Dosage compensation in Drosophila. Annu. Rev. Genet. 1994,28:491-521
[127] LUCCHESI J.C. Dosage compensation in flies and worms: the ups and downs of X-chromosome regulation. Curr. Opin. Genet. Dev. 1998,8:179-184
[128] FRANKE A., BAKER B.S. The roxl and rox2 RNAs are essential components of the "compensasome", which mediates dosage compensation in Drosophila. Mol. Cell. 1999,4: 117-122
[129] KELLEY R.L., KURODA M.I. The role of chromosomal RNAs in marking the X for dosage compensation. Curr. Opin. Genet. Dev. 2000,10:555-561
[130] Kuo M.H., Allis C.D. Roles of histone acetyltransferases and deacetylases in gene regulation. Bioessays. 1998,20:615-626
[131] Pazin M., Kadonaga J. What's up and down with histone deacetylation and transcription? Cell. 1997,89:325-328
[132] Astrid S.C., Joanna E.L., Sandra J.J., et al. Esalp is an essential histone acetyltransferase required for cell cycle progression. Mol. Cell. Biol. 1999,19:2515-2526
[133] Mikko T., Stephen R., Karsten R., et al. hMOF histone acetyltmnsferase is required for histone H4 lysine 16 acetylation in mammalian cells, Mol. Cell Biol. 2005,25:6798-810
[134] Fraga M.F., Ballestar E., Villar-Garea A., et al. Loss of acetylation at Lys 16 and trimethylation at Lys20 of histone H4 is a common hallmark of human cancer. Nat. Genet. 2005,37:391-400
[135] Ikura T., Ogryzko V.V., Grigoriev M., et al. Involvement of the TIP60 histone acetylase complex in DNA repair and apoptosis. Cell. 2000102:463-473
[136] Kamine J., Elangovan B., Subramanian T., et al. Identification of a cellular protein that specifically interacts with the essential cysteine region of the HⅣ-1 Tat transactivator. Virology. 1996,216:357-366
[137] Kusch T., Florens L., Macdonald W.H., et al. Acetylation by Tip60 Is Required for Selective Histone Variant Exchange at DNA Lesions. Science. 2004,306:2084-2087
[138] Yochum G.S., Ayer D.E. Role for the Mortality Factors MORF4, MRGX, and MRG15 in Transcriptional Repression via Associations with Pfl, mSin3A, and Transduein-Like Enhancer of Split. Mol. Cell. Biol. 2002,22:7868-7876
[139] Matin I., Baker B.S. Origin and Evolution of the Regulatory Gene male-specific lethal-3. Mol. Biol. Evol. 2000,17:1240-1250
[140] Pardo P.S., Leung J.K., Lucchesi. C., et aL MRG15, a Novel Chromodomain Protein, Is Present in Two Distinct Multiprotein Complexes Involved in Transcriptional Activation. J Biol. Chem. 2002,277:50860-50866
[141] Zhang P., Zhao J.Y., Wang B., et al. The MRG domain of hunman MRG15 uses a shallow hydrophobic pocket to interact with the N-terminal region of PAM14. Protein Science. 2006,15: 2423-2434
[142] Ball L.J., Murzina N.V., Broadhurst R.W., et al. Structure of the chromatin binding (chromo) domain from mouse modifier protein 1. EMBO J. 1997,16:2473-2481
[143] Fields S., Song O. A Novel genetic system to detect protein-protein intersctions. Nature. 1989, 340:245-246
[144] 赵永芳．生物化学技术原理及应用．北京：科学出版社．2002：7-82
[145] Kaelin W.G., Pallas D.C., DeCaprio J.A., et al. Identification of cellular proteins that can interact specifically with the T/E1A-binding region of the retinoblastoma gene product. Cell. 1991,64:521-532
[146] Harlow E. and Lane D. Antibodies: A laboratory manual Cold spring Harbor Laboratory, Cold Spring Harbor, New York.
[147] Harlow E. and Lane D. Using antibodies: A laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York.
[148] Edmondson S.P., Qiu L., Shriver J.W. Solution structure of the DNA-binding protein Sac7d from the hyperthermophile Sulfolobus acidocaldarius. Biochemistry. 1995,34:13289-13304
[149] Riddihough G., Pennisi E. The Evolution of Epigenetics. Science.2001,293:1063
[150] Madireddi M.T., Coyne R.S., Smothers J.F., et al. Pddlp, a novel chromodomain-containing protein, links heterochromatin assembly and DNA elimination in Tetrahymena.Cell.1996,87: 75-84
[151] Xia Q.Y., Zhou Z.Y., Lu C., et al. A Draft Sequence for the Genome of the Domesticated Silkworm (Bombyx mori). Science. 2004, 306:1937-1940
[152] Mita K., Kasahara M., Sasaki S., et al. The Genome Sequence of Silkworm, Bombyx mori DNA Research. 2004,11:27-35
[153] Isobe M., Kai H., Kurahashi T., et al. The Molecular Mechanism of the Termination of Insect Diapause, Part 1: A Timer Protein, TIME-EA4, in the Diapause Eggs of the Silkworm, Bombyx mori is a Metallo-Glycoprotein. Chembiochem. 2006, 7:1590-1598.
[154] MiaoX.X., Xu S.J., Li M.H., et al. Simple sequence repeat-based consensus linkage map of Bombyx mori. PNAS. 2005, 102:16303-16308
[155] Shilatifard A. Chromatin Modifications by Methylation and Ubiquitination: Implications in the Regulation of Gene Expression. Ann. Rev. Biochem. 2006, 75: 243～269.
[156] Suzuki M.G., Shimada T., Kobayashi M. Bm kettin, homologue of the Drosophila kettin gene, is located on the Z chromosome in Bombyx mori and is not dosage compensated. Heredity. 1999, 82:170-179
[157] Koike Y., Mita K., Suzuki M.G., et al. Genomic sequence of a 320-kb segment of the Z chromosome of Bombyx mori containing a kettin ortholog. Mol Gen Genomics. 2003,269: 137-149
[158] Marin I. Evolution of chromatin-remodeling complexes: comparative genomics reveals the ancient origin of "novel" compensasome genes. J Mol. Evol. 2003,56:527-539
[159] Gorinsek F.B., Gubensek K.D. Phylogenomic analysis of chromoviruses. Cytogenet Genome Res. 2005, 110:543-552