Graduate Institute of Life Sciences, National Defense Medical Center, Taipei 11490, Taiwan and Institute of Biomedical Sciences, Academia Sinica, Taipei 11529, Taiwan.
Nucleic Acids Res. 2013 Nov;41(21):9858-69. doi: 10.1093/nar/gkt746. Epub 2013 Aug 21.
Deleted in Azoospermia Associated Protein 1 (DAZAP1) is a ubiquitous heterogeneous nuclear ribonucleoprotein (hnRNP) that is expressed abundantly in the testis. DAZAP1 deficiency in mice results in growth retardation and spermatogenic arrest. Previous reports on DAZAP1's binding to several naturally occurring splicing mutations support a role for DAZAP1 in RNA splicing. To elucidate the biological function(s) of DAZAP1 and to search for its natural RNA substrates, we used microarrays to compare the expression profiles and exon usages of wild-type and Dazap1 mutant testes and identified three genes (Crem, Crisp2 and Pot1a) with aberrant RNA splicing in the mutant testes. We further demonstrated that DAZAP1, but not DAZAP1 mutant proteins, promoted the inclusion of Crem exon 4, Crisp2 exon 9 and Pot1a exon 4 in splicing reporter transcripts in cultured cells. Additional studies on the binding of DAZAP1 to the exons and their flanking intronic sequences and the effects of minigene deletions on exon inclusion identified regulatory regions in Crem intron 3, Crisp2 intron 9 and Pot1a intron 4 where DAZAP1 bound and regulated splicing. Aberrant splicing of the Pot1a gene, which encodes an essential protein that protects telomere integrity, may partially account for the growth retardation phenotype of DAZAP1-deficient mice.
缺失于无精症相关基因 1 蛋白（DAZAP1）是一种普遍存在的异质核核糖核蛋白（hnRNP），在睾丸中大量表达。DAZAP1 基因在小鼠中缺失会导致生长迟缓和精子发生停滞。先前关于 DAZAP1 与几种天然剪接突变体结合的报道支持 DAZAP1 在 RNA 剪接中的作用。为了阐明 DAZAP1 的生物学功能并寻找其天然 RNA 底物，我们使用微阵列比较了野生型和 Dazap1 突变型睾丸的表达谱和外显子使用情况，在突变型睾丸中发现了三个基因（Crem、Crisp2 和 Pot1a）存在异常 RNA 剪接。我们进一步证明，DAZAP1 而不是 DAZAP1 突变蛋白，促进了 Crem 外显子 4、Crisp2 外显子 9 和 Pot1a 外显子 4 在培养细胞中的剪接报告转录本中的包含。对 DAZAP1 与外显子及其侧翼内含子序列的结合以及基因缺失对内含子的影响的进一步研究，确定了 Crem 内含子 3、Crisp2 内含子 9 和 Pot1a 内含子 4 中 DAZAP1 结合和调节剪接的调控区域。Pot1a 基因的异常剪接，该基因编码一种保护端粒完整性的必需蛋白，可能部分解释了 DAZAP1 缺失型小鼠的生长迟缓表型。
