DAZ 家族包含三個成員 : BOULE、DAZL (DAZ-like) 和 DAZ，皆會專一性表現在生殖細胞中，具有結合 RNA 與蛋白質之功能。在男性不孕症病人常見其 Y 染色體長臂上的 AZF 區段之缺失，此區段包含 DAZ 基因。DAZ 的同源基因 DAZL 位於第三號染色體，在 DAZL 基因剃除的動物模式研究實驗中也發現生殖細胞缺失的現象，目前已知 DAZL 的缺失與不孕症有相關性，但受其調控之下游基因尚待探討。文獻指出 DAZL 會結合不同基因之 mRNA 來進行後轉錄作用的調控，藉由促進不同基因之蛋白表現，在精子形成過程中執行不同功能，所以 DAZL 在生殖細胞之發育與分化扮演重要角色。實驗室之前在患有無精子症與少精症的台灣男性不孕症病人中篩選到數個 DAZL 核苷酸突變與單一核苷酸多型性的變異，特別是在核苷酸 386 (T54A) 位置由 adenine 變成 guanine，此單一核苷酸多型性與不孕症病人有高度的關聯性。為了闡明單一核苷酸多型性對 DAZL 功能的影響，首先我們先利用 RNA 免疫沉澱分析並結合 deep-sequencing 技術篩選 DAZL 可能作用的 mRNA，我們得到 NUDT21、NUP50、KLHL12、KDELR1、VAPA、ABHD2、SUPT5H、MLL3、ATP6AP2 等可能是 DAZL 調控的 transcripts。其中 NUDT21 為 CFIm complex 的次單元，在文獻中指出會高量表現在老鼠生殖細胞且參與在精子形成過程中的 alternative polyadenylation 的機制，因此選定 NUDT21 作為探討 DAZL 調控機制的標的。結果顯示 DAZL 會結合 NUDT21 ，並透過後轉錄層面促進轉譯效率，增加 NUDT21 蛋白表現。然而帶有單一核苷酸多型性 DAZLT54A 會降低 NUDT21 的轉譯效率，造成 NUDT21 蛋白量減少，同時發現 SREBF2、NR6A1 及 TIMP-2 的 poly(A) 選擇位改變，結果顯示較短片段的 truncated 3’UTR 表現量增加，較長片段的 3’UTR 表現量相對減少，顯示 DAZL 可能參與 alternative polyadenylation。綜合以上結果顯示 NUDT21 為 DAZL 調控的下游基因，且帶有單一核苷酸多型性 DAZL 透過 NUDT21 改變基因 poly(A) 選擇位，此現象可能影響精子正常發育。

The DAZ family encodes RNA-binding proteins that are essential for gametogenesis in metazoans. In humans, loss of the Y chromosomal DAZ genes is associated with oligozoospermia or azoospermia. A DAZ homolog, DAZL (DAZ-Like), is found in diverse organisms, including humans, and is required for germ cell development. Although the loss of DAZL is associated with infertility, the mechanism of how DAZL regulates its substrates is largely unknown. It has been proposed that DAZL controls spermatogenesis through regulating the translation efficiency of targeted mRNAs. Previously, we have identified a SNP of DAZL, the A to G transition at nucleotide 386 (T54A), which confers susceptibility to spermatogenic failure. A number of target transcripts, including NUDT21, NUP50, KLHL12, KDELR1, VAPA, ABHD2, SUPT5H, MLL3 and ATP6AP2, were also identified as targeted mRNA of DAZL. Among these genes, the NUDT21, encoding one of the CFIm subunits, was chose for further study. NUDT21 gene expresses three isoforms, including 1.1, 2.0 and 4.5 kb mRNA. While the 2.0 kb transcript is expressed in multiple tissues, the 1.1 kb transcript is dominantly expressed in male germ lines. NUDT21 participates in the alternative polyadenylation mechanism directed by noncanonical poly(A) signals during spermatogenesis. To study how NUDT21 is regulated by DAZL, RNA-IP and S6-IP were used to examine the binding activity and translational efficiency of NUDT21. We found DAZL increased both the binding activity and translational efficiency of the 1.1-kb transcript. However, the SNP DAZLT54A decreased translational efficiency of the 1.1-kb transcript and resulted in decreased NUDT21 protein level. Moreover, the alternative poly(A) site selection of SREBF2, NR6A1 and TIMP-2 was analyzed. We observed changes in the poly(A) site selection in DAZLT54A overexpressing cells. It suggests a role for DAZL in alternative poly(A) site selection. Together, these data support NUDT21 as a candidate target of DAZL, and also suggest SNP DAZLT54A may cause spermatogenic failure by altering the polyadenylation sites of genes via modulating the function of NUDT21.

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