蛋白乙酰化在基因组稳定性维护中发挥重要作用

组蛋白的乙酰化水平直接影响着染色质结构,使它表现出最适物理化学特性,从而保证着遗传物质顺利地被复制。遗传物质只有顺利完整地被复制,生物同一种族间遗传物质才会在数量上高度稳定,生物种族才得以延续。然而目前大量研究已经表明蛋白乙酰化还包括大量非组蛋白乙酰化,还有许多更为复杂的作用。它们不仅仅影响着遗传物质的传递和表达,还在DNA损伤修复中发挥重要作用。该文综述列举了大量组蛋白和非组蛋白乙酰化的例子,并通过这些例子详细阐述了蛋白乙酰化在保持基因组稳定中发挥作用的机制。     

CHD1 motor protein is required for deposition of histone variant H3.3 into chromatin in vivo

The organization of chromatin affects all aspects of nuclear DNA metabolism in eukaryotes. H3.3 is an evolutionarily conserved histone variant and a key substrate for replication-independent chromatin assembly. Elimination of chromatin remodeling factor CHD1 in Drosophila embryos abolishes incorporation of H3.3 into the male pronucleus, renders the paternal genome unable to participate in zygotic mitoses, and leads to the development of haploid embryos. Furthermore, CHD1, but not ISWI, interacts with HIRA in cytoplasmic extracts. Our findings establish CHD1 as a major factor in replacement histone metabolism in the nucleus and reveal a critical role for CHD1 in the earliest developmental instances of genome-scale, replication-independent nucleosome assembly. Furthermore, our results point to the general requirement of adenosine triphosphate (ATP)-utilizing motor proteins for histone deposition in vivo.     

Dependence of heterochromatic histone H3 methylation patterns on the Arabidopsis gene DDM1

The Arabidopsis gene DDM1 is required to maintain DNA methylation levels and is responsible for transposon and transgene silencing. However, rather than encoding a DNA methyltransferase, DDM1 has similarity to the SWI/SNF family of adenosine triphosphate-dependent chromatin remodeling genes, suggesting an indirect role in DNA methylation. Here we show that DDM1 is also required to maintain histone H3 methylation patterns. In wild-type heterochromatin, transposons and silent genes are associated with histone H3 methylated at lysine 9, whereas known genes are preferentially associated with methylated lysine 4. In ddm1 heterochromatin, DNA methylation is lost, and methylation of lysine 9 is largely replaced by methylation of lysine 4. Because DNA methylation has recently been shown to depend on histone H3 lysine 9 methylation, our results suggest that transposon methylation may be guided by histone H3 methylation in plant genomes. This would account for the epigenetic inheritance of hypomethylated DNA once histone H3 methylation patterns are altered.     

Phosphorylation of nuclear protein early in the course of gene activation in lymphocytes

Human lymphocytes treated with phytohemagglutinin undergo extensive gene activation, as evidenced by augmented synthesis of ribonucleic acids. This activation is preceded by an early stimulation in the rate of phosphorylation and dephosphorylation of nuclear proteins. This finding is consistent with a hypothesized role of phosphoproteins in the modification of chromatin structure and in modulation of the template activity of DNA in vivo.     

Adenosine 3',5'-monophosphate-dependent protein kinase of cultured mammalian cells

Protein kinase was partially purified from Chang's liver cells, 3T6 mouse embryo fibroblasts, and HeLa cells. The rate of histone phosphorylation catalyzed by the kinase from each of these cell lines was stimulated two- to three-fold by 1 x 10(-6) molar adenosine 3',5'-monophosphate. The same concentration of guanosine 3',5'-monophosphate failed to stimulate these kinases.     

Activation of the DNA replication checkpoint through RNA synthesis by primase

When DNA replication is inhibited during the synthesis (S) phase of the cell cycle, a signaling pathway (checkpoint) is activated that serves to prevent mitosis from initiating before completion of replication. This replication checkpoint acts by down-regulating the activity of the mitotic inducer cdc2-cyclin B. Here, we report the relation between chromatin structure and induction of the replication checkpoint. Chromatin was competent to initiate a checkpoint response only after the DNA was unwound and DNA polymerase alpha had been loaded. Checkpoint induction did not require new DNA synthesis on the unwound template strand but did require RNA primer synthesis by primase. These findings identify the RNA portion of the primer as an important component of the signal that activates the replication checkpoint.     

Histone H4-K16 acetylation controls chromatin structure and protein interactions

Acetylation of histone H4 on lysine 16 (H4-K16Ac) is a prevalent and reversible posttranslational chromatin modification in eukaryotes. To characterize the structural and functional role of this mark, we used a native chemical ligation strategy to generate histone H4 that was homogeneously acetylated at K16. The incorporation of this modified histone into nucleosomal arrays inhibits the formation of compact 30-nanometer-like fibers and impedes the ability of chromatin to form cross-fiber interactions. H4-K16Ac also inhibits the ability of the adenosine triphosphate-utilizing chromatin assembly and remodeling enzyme ACF to mobilize a mononucleosome, indicating that this single histone modification modulates both higher order chromatin structure and functional interactions between a nonhistone protein and the chromatin fiber.     

草莓斑驳病毒的分子杂交检测

为探索分子杂交技术在草莓斑驳病毒(Strawberry mottle virus,SMoV)检测中的运用,利用CTAB法从草莓叶片中提取总核酸,并以此为模板,利用特异引物扩增SMoV非编码区序列,经克隆、测序获得SMoV非编码区序列。系统进化分析结果显示:本研究中的分离物与GenBank中已报道的分离物的亲缘关系较远,独立形成1个小的进化支。以带有病毒特异片段的质粒为模板,运用PCR技术制备了地高辛标记的cDNA探针。以草莓感染SMoV叶片为试材,利用地高辛标记探针可有效检测草莓叶片中的SMoV,草莓总核酸样品稀释10倍后仍可有效检测。     

Chromosomal proteins in the dinoflagellate alga Gyrodinium cohnii

Chromatin has been prepared from nuclei isolated from the dinoflagellate alga Gyrodinium cohnii. This chromatin contains RNA, acid-insoluble proteins, and acid-soluble proteins; the respective ratios to amount of DNA are about 0.09, 0.48, 0.08 (by weight). Not only is the amount of acid-soluble protein associated with the DNA much less than it is in the typical eukaryote, but polyacrylamide gel electrophoresis in urea at pH 3.2 produces a banding pattern different from that of typical histones. There is one predominant band that migrates about as fast as does histone IV from corn. These findings are of interest, because the nuclear organization in the dinoflagellates appears to be intermediate between the prokaryotes and the eukaryotes.     

Regulation of replication fork progression through histone supply and demand

DNA replication in eukaryotes requires nucleosome disruption ahead of the replication fork and reassembly behind. An unresolved issue concerns how histone dynamics are coordinated with fork progression to maintain chromosomal stability. Here, we characterize a complex in which the human histone chaperone Asf1 and MCM2-7, the putative replicative helicase, are connected through a histone H3-H4 bridge. Depletion of Asf1 by RNA interference impedes DNA unwinding at replication sites, and similar defects arise from overproduction of new histone H3-H4 that compromises Asf1 function. These data link Asf1 chaperone function, histone supply, and replicative unwinding of DNA in chromatin. We propose that Asf1, as a histone acceptor and donor, handles parental and new histones at the replication fork via an Asf1-(H3-H4)-MCM2-7 intermediate and thus provides a means to fine-tune replication fork progression and histone supply and demand.     

山梨SRAP反应体系的建立

以山梨为试材,提取其基因组DNA,探讨了SRAP反应体系中各组分浓度对扩增的影响。结果表明,采用该方法提取的基因组DNA纯度好、片段完整、无明显降解;在20μL的反应体系中,各成分适宜浓度分别为TaqDNA聚合酶1.0 U、模板DNA 70 ng、dNTPs 250μmol/L、每个特异性引物0.3μmol/L、10×PCR Buffer 2μL,余下的体积用已灭菌的去离子水补充至20μL。     

A histone acetyltransferase regulates active DNA demethylation in Arabidopsis

Active DNA demethylation is an important part of epigenetic regulation in plants and animals. How active DNA demethylation is regulated and its relationship with histone modification patterns are unclear. Here, we report the discovery of IDM1, a regulator of DNA demethylation in Arabidopsis. IDM1 is required for preventing DNA hypermethylation of highly homologous multicopy genes and other repetitive sequences that are normally targeted for active DNA demethylation by Repressor of Silencing 1 and related 5-methylcytosine DNA glycosylases. IDM1 binds methylated DNA at chromatin sites lacking histone H3K4 di- or trimethylation and acetylates H3 to create a chromatin environment permissible for 5-methylcytosine DNA glycosylases to function. Our study reveals how some genes are indicated by multiple epigenetic marks for active DNA demethylation and protection from silencing.     

Size of the Chloroplast Genome in Codium fragile

Chloroplasts isolated from the siphonous green alga Codium fragile yield circular DNA molecules averaging 27.3 micrometers in length and 56 x 10(6) daltons in molecular size. This chloroplast genome is 25 to 30 percent smaller than any reported. The small size of the Codium chloroplast genome may represent a primitive evolutionary condition in green plants.