Yale Stem Cell Center and Department of Cell Biology, Yale University School of Medicine, New Haven, CT 06509, USA.
Cell Res. 2012 Oct;22(10):1429-39. doi: 10.1038/cr.2012.120. Epub 2012 Aug 21.
piRNAs, a class of small non-coding RNAs associated with PIWI proteins, have broad functions in germline development, transposon silencing, and epigenetic regulation. In diverse organisms, a subset of piRNAs derived from repeat sequences are produced via the interplay between two PIWI proteins. This mechanism, termed "ping-pong" cycle, operates among the PIWI proteins of the primordial mouse testis; however, its involvement in postnatal testes remains elusive. Here we show that adult testicular piRNAs are produced independent of the ping-pong mechanism. We identified and characterized large populations of piRNAs in the adult and postnatal developing testes associated with MILI and MIWI, the only PIWI proteins detectable in these testes. No interaction between MILI and MIWI or sequence feature for the ping-pong mechanism among their piRNAs was detected in the adult testis. The majority of MILI- and MIWI-associated piRNAs originate from the same DNA strands within the same loci. Both populations of piRNAs are biased for 5' Uracil but not for Adenine on the 10th nucleotide position, and display no complementarity. Furthermore, in Miwi mutants, MILI-associated piRNAs are not downregulated, but instead upregulated. These results indicate that the adult testicular piRNAs are predominantly, if not exclusively, produced by a primary processing mechanism instead of the ping-pong mechanism. In this primary pathway, biogenesis of MILI- and MIWI-associated piRNAs may compete for the same precursors; the types of piRNAs produced tend to be non-selectively dictated by the available precursors in the cell; and precursors with introns tend to be spliced before processed into piRNAs.
piRNAs 是一类与 PIWI 蛋白相关的小非编码 RNA，在生殖细胞发育、转座子沉默和表观遗传调控中具有广泛的功能。在不同的生物体中，一小部分源自重复序列的 piRNAs 是通过两种 PIWI 蛋白的相互作用产生的。这种机制被称为“乒乓”循环，在原始小鼠睾丸的 PIWI 蛋白中起作用；然而，其在出生后睾丸中的参与仍然难以捉摸。在这里，我们表明成年睾丸 piRNAs 的产生不依赖于乒乓循环机制。我们鉴定并表征了成年和出生后发育的睾丸中与 MILI 和 MIWI 相关的大量 piRNAs，MILI 和 MIWI 是这些睾丸中唯一可检测到的 PIWI 蛋白。在成年睾丸中，未检测到 MILI 和 MIWI 之间的相互作用，也未检测到其 piRNAs 中乒乓循环机制的序列特征。MILI 和 MIWI 相关的 piRNAs 主要来源于同一基因座内的相同 DNA 链。这两种 piRNA 群体都偏向于 5' 尿嘧啶而不是第 10 个核苷酸位置的腺嘌呤，并且没有互补性。此外，在 Miwi 突变体中，MILI 相关的 piRNAs 没有下调，而是上调。这些结果表明，成年睾丸 piRNAs 主要（如果不是完全的话）由初级加工机制而不是乒乓循环机制产生。在这个主要途径中，MILI 和 MIWI 相关的 piRNA 的生物发生可能竞争相同的前体；产生的 piRNA 类型倾向于由细胞中可用的前体非选择性决定；并且具有内含子的前体倾向于在加工成 piRNA 之前被剪接。
