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首页> 外文期刊>Molecular and Cellular Biology >Mammalian DNA ligase III: molecular cloning, chromosomal localization, and expression in spermatocytes undergoing meiotic recombination.
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Mammalian DNA ligase III: molecular cloning, chromosomal localization, and expression in spermatocytes undergoing meiotic recombination.

机译：哺乳动物DNA连接酶III：分子克隆，染色体定位和减数分裂重组在精母细胞中的表达。

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Three biochemically distinct DNA ligase activities have been identified in mammalian cell extracts. We have recently purified DNA ligase II and DNA ligase III to near homogeneity from bovine liver and testis tissue, respectively. Amino acid sequencing studies indicated that these enzymes are encoded by the same gene. In the present study, human and murine cDNA clones encoding DNA ligase III were isolated with probes based on the peptide sequences. The human DNA ligase III cDNA encodes a polypeptide of 862 amino acids, whose sequence is more closely related to those of the DNA ligases encoded by poxviruses than to replicative DNA ligases, such as human DNA ligase I. In vitro transcription and translation of the cDNA produced a catalytically active DNA ligase similar in size and substrate specificity to the purified bovine enzyme. The DNA ligase III gene was localized to human chromosome 17, which eliminated this gene as a candidate for the cancer-prone disease Bloom syndrome that is associated with DNA joining abnormalities. DNA ligase III is ubiquitously expressed at low levels, except in the testes, in which the steady-state levels of DNA ligase III mRNA are at least 10-fold higher than those detected in other tissues and cells. Since DNA ligase I mRNA is also present at high levels in the testes, we examined the expression of the DNA ligase genes during spermatogenesis. DNA ligase I mRNA expression correlated with the contribution of proliferating spermatogonia cells to the testes, in agreement with the previously defined role of this enzyme in DNA replication. In contrast, elevated levels of DNA ligase III mRNA were observed in primary spermatocytes undergoing recombination prior to the first meiotic division. Therefore, we suggest that DNA ligase III seals DNA strand breaks that arise during the process of meiotic recombination in germ cells and as a consequence of DNA damage in somatic cells.
机译：在哺乳动物细胞提取物中已鉴定出三种在生物化学上不同的DNA连接酶活性。我们最近从牛肝和睾丸组织中分别纯化了DNA连接酶II和DNA连接酶III至接近同质。氨基酸测序研究表明，这些酶是由同一基因编码的。在本研究中，使用基于肽序列的探针分离了编码DNA连接酶III的人和鼠cDNA克隆。人DNA连接酶III cDNA编码一个862个氨基酸的多肽，其序列与痘病毒编码的DNA连接酶的序列更紧密，而与诸如人DNA连接酶I的复制DNA连接酶更紧密相关。cDNA的体外转录和翻译产生了具有催化活性的DNA连接酶，其大小和底物特异性与纯化的牛酶相似。 DNA连接酶III基因位于人的17号染色​​体上，从而消除了该基因，使之成为易患癌症的Bloom综合征的候选基因，该综合征与DNA连接异常有关。 DNA连接酶III低水平普遍表达，但在睾丸中除外，在睾丸中，DNA连接酶III mRNA的稳态水平比在其他组织和细胞中检测到的水平至少高10倍。由于DNA连接酶I mRNA在睾丸中的含量也很高，因此我们检查了精子发生过程中DNA连接酶基因的表达。 DNA连接酶I mRNA表达与增殖的精原细胞对睾丸的贡献有关，与该酶在DNA复制中的先前定义作用一致。相反，在第一次减数分裂分裂之前经历重组的原代精母细胞中观察到DNA连接酶III mRNA的水平升高。因此，我们建议DNA连接酶III密封在生殖细胞减数分裂重组过程中以及体细胞中DNA损伤的过程中出现的DNA链断裂。

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