Department of Biochemistry, Indian Institute of Science, Bangalore, India.
Institute of Bioinformatics and Applied Biotechnology, Electronics City, Bangalore, India.
J Immunol. 2024 Oct 15;213(8):1125-1138. doi: 10.4049/jimmunol.2300344.
The process of Ag receptor diversity is initiated by RAGs consisting of RAG1 and RAG2 in developing lymphocytes. Besides its role as a sequence-specific nuclease during V(D)J recombination, RAGs can also act as a structure-specific nuclease leading to genome instability. Thus, regulation of RAG expression is essential to maintaining genome stability. Previously, the role of miR29c in the regulation of RAG1 was identified. In this article, we report the regulation of RAG1 by miR-29a in the lymphocytes of both mice (Mus musculus) and humans (Homo sapiens). The level of RAG1 could be modulated by overexpression of miR-29a and inhibition using anti-miRs. Argonaute2-immunoprecipitation and high-throughput sequencing of RNA isolated by crosslinking immunoprecipitation studies established the association of miR-29a and RAG1 with Argonaute proteins. We observed a negative correlation between miR-29a and RAG1 levels in mouse B and T cells and leukemia patients. Overexpression of pre-miR-29a in the bone marrow cells of mice led to the generation of mature miR-29a transcripts and reduced RAG1 expression, which led to a significant reduction in V(D)J recombination in pro-B cells. Importantly, our studies are consistent with the phenotype reported in miR-29a knockout mice, which showed impaired immunity and survival defects. Finally, we show that although both miR-29c and miR-29a can regulate RAG1 at mRNA and protein levels, miR-29a substantially impacts immunity and survival. Our results reveal that the repression of RAG1 activity by miR-29a in B cells of mice and humans is essential to maintain Ig diversity and prevent hematological malignancies resulting from aberrant RAG1 expression in lymphocytes.
Ag 受体多样性的形成过程始于发育淋巴细胞中的 RAG 蛋白，该蛋白由 RAG1 和 RAG2 组成。除了在 V(D)J 重组过程中作为序列特异性核酸内切酶发挥作用外，RAG 还可以作为结构特异性核酸内切酶，导致基因组不稳定。因此，RAG 表达的调控对于维持基因组稳定性至关重要。先前已经确定了 miR29c 在调控 RAG1 表达中的作用。在本文中，我们报告了 miR-29a 在小鼠（Mus musculus）和人类（Homo sapiens）淋巴细胞中对 RAG1 的调控作用。通过过表达 miR-29a 和使用抗 miR 抑制，RAG1 的水平可以被调节。Argonaute2 免疫沉淀和交联免疫沉淀研究中分离的 RNA 的高通量测序确立了 miR-29a 和 RAG1 与 Argonaute 蛋白的关联。我们观察到在小鼠 B 和 T 细胞和白血病患者中 miR-29a 和 RAG1 水平之间存在负相关。在小鼠骨髓细胞中过表达 pre-miR-29a 会导致成熟 miR-29a 转录本的产生和 RAG1 表达的减少，这导致前 B 细胞中的 V(D)J 重组显著减少。重要的是，我们的研究与 miR-29a 敲除小鼠报告的表型一致，该表型显示免疫受损和生存缺陷。最后，我们表明，尽管 miR-29c 和 miR-29a 都可以在 mRNA 和蛋白质水平上调节 RAG1，但 miR-29a 对免疫和生存的影响要大得多。我们的研究结果表明，miR-29a 在小鼠和人类 B 细胞中对 RAG1 活性的抑制对于维持 Ig 多样性以及预防淋巴细胞中异常 RAG1 表达导致的血液恶性肿瘤至关重要。
