Diversification of the avian primary immunoglobulin (Ig) repertoire is achieved in developing B cells by somatic hypermutation (SHM) and gene conversion (GCV). GCV is a type of homologous recombination that unidirectionally transfers segments of Ig pseudogenes to Ig variable domains. It is regulated by epigenetic mechanisms like histone modifications, but the role of DNA methylation remains unclear. Here, we demonstrate that the chicken B‐cell line DT40 lacking TET3, a member of the TET (Ten‐eleven translocation) family dioxygenases that facilitate DNA demethylation, exhibited a marked reduction in GCV activity in Ig variable regions. This was accompanied by a drop in the bulk levels of 5‐hydroxymethylcytosine, an oxidized derivative of 5‐methylcytosine, whereas TET1‐deficient or TET2‐deficient DT40 strains did not exhibit such effects. Deletion of TET3 caused little effects on the expression of proteins required for SHM and GCV, but induced hypermethylation in some Ig pseudogene templates. Notably, the enhanced methylation occurred preferably on non‐CpG cytosines. Disruption of both TET1 and TET3 significantly inhibited the expression of activation‐induced cytidine deaminase (AID), an essential player in Ig diversification. These results uncover unique roles of TET proteins in avian Ig diversification, highlighting the potential importance of TET3 in maintaining hypomethylation In Ig pseudogenes.

中文翻译：

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TET3双加氧酶通过伪基因模板中非CpG位点的去甲基化来调节禽类免疫球蛋白可变区的基因转化

通过体细胞超突变（SHM）和基因转化（GCV），在发育中的B细胞中实现了禽原代免疫球蛋白（Ig）种类的多样化。GCV是一种同源重组，可将Ig假基因的片段单向转移至Ig可变域。它受组蛋白修饰等表观遗传机制调控，但DNA甲基化的作用仍不清楚。在这里，我们证明了缺乏TET3（促进DNA去甲基化的TET（十一位易位）家族双加氧酶的成员）的鸡B细胞株DT40在Ig可变区中GCV活性显着降低。这伴随着5-羟甲基胞嘧啶（5-甲基胞嘧啶的氧化衍生物）的总体含量下降，而TET1缺乏或TET2缺乏的DT40菌株则没有这种作用。TET3的删除对SHM和GCV所需蛋白质的表达影响不大，但在某些Ig假基因模板中诱导了超甲基化。值得注意的是，增强甲基化最好发生在非CpG胞嘧啶上。TET1和TET3的破坏均显着抑制了活化诱导的胞苷脱氨酶（AID）的表达，这是Ig多样化的重要因素。这些结果揭示了TET蛋白在禽类Ig多样化中的独特作用，突显了TET3在维持Ig假基因的低甲基化方面的潜在重要性。TET1和TET3的破坏均显着抑制了活化诱导的胞苷脱氨酶（AID）的表达，这是Ig多样化的重要因素。这些结果揭示了TET蛋白在禽类Ig多样化中的独特作用，突显了TET3在维持Ig假基因的低甲基化方面的潜在重要性。TET1和TET3的破坏均显着抑制了活化诱导的胞苷脱氨酶（AID）的表达，这是Ig多样化的重要因素。这些结果揭示了TET蛋白在禽类Ig多样化中的独特作用，突显了TET3在维持Ig假基因的低甲基化方面的潜在重要性。