DiNardo Acadia L, Kurhanewicz Nicole A, Wilson Hannah R, Berg Veronica, Libuda Diana E
Institute of Molecular Biology, Department of Biology, University of Oregon, 1229 Franklin Blvd Eugene, OR 97403, USA.
Department of Biology, University of Oregon, 1229 Franklin Blvd Eugene, OR 97403, USA.
bioRxiv. 2025 Jun 1:2025.05.29.656876. doi: 10.1101/2025.05.29.656876.
Germ cell proliferation and proper genome inheritance are critical for maintaining fertility through generations. To promote proper germ cell development, small RNA pathways employ Argonaute proteins (AGOs) to modulate gene expression and protect against deleterious genomic elements while not silencing against self. Here we identify sexual dimorphisms in localization and function of protein structural features of the Argonaute WAGO-1 that affects sex-specific gene regulation during germ cell development. During meiotic prophase I progression, we find that germ granule structural proteins and the PIWI AGO, PRG-1, display dynamic and distinct localization patterns between egg and sperm development which coincide with differential WAGO-1 localization and biophysical properties. Sexually dimorphic functions of specific WAGO-1 protein structural domains underpin these differences. Disruption or modification to the N-terminus intrinsically disordered region (IDR) of WAGO-1 leads to loss of PGL-1 phase separation only during spermatogenesis. Further, we find that these germ granule disruptions are likely due to prolonged association of the IDR with the RNA-binding pocket of WAGO-1. In addition, deletion of the MID and part of the PIWI domains causes male-specific sterility and disruption to WAGO-1 localization with PGL-1 during oogenesis. Finally, we demonstrate that these disruptions to WAGO-1 protein structure dynamically change the mRNA and sRNA landscape of adult males and hermaphrodites, in which the AGOs ALG-3/4 and VSRA-1 are misregulated. Together, these data suggest that WAGO-1 differentially regulates genes during oogenesis versus spermatogenesis, and that these differences in gene regulation may be due to the sex-specific configuration and biophysical properties of WAGO-1 within the germ granule.
生殖细胞增殖和正确的基因组遗传对于世代维持生育能力至关重要。为促进生殖细胞的正常发育，小RNA通路利用AGO蛋白（AGOs）调节基因表达并抵御有害基因组元件，同时不会对自身进行沉默。在这里，我们鉴定了AGO蛋白WAGO-1的蛋白质结构特征在定位和功能上的性别差异，该差异影响生殖细胞发育过程中的性别特异性基因调控。在减数分裂前期I进程中，我们发现生殖颗粒结构蛋白和PIWI AGO蛋白PRG-1在卵子和精子发育过程中呈现出动态且不同的定位模式，这与WAGO-1的差异定位和生物物理特性相吻合。特定WAGO-1蛋白结构域的性别二态性功能支撑了这些差异。对WAGO-1的N端内在无序区域（IDR）的破坏或修饰仅在精子发生过程中导致PGL-1相分离丧失。此外，我们发现这些生殖颗粒的破坏可能是由于IDR与WAGO-1的RNA结合口袋的长时间结合。另外，MID和部分PIWI结构域的缺失会导致雌性特异性不育，并在卵子发生过程中破坏WAGO-1与PGL-1的定位。最后，我们证明这些对WAGO-1蛋白结构的破坏会动态改变成年雄性和雌雄同体动物的mRNA和sRNA图谱，其中AGO蛋白ALG-3/4和VSRA-1的表达失调。总之，这些数据表明WAGO-1在卵子发生和精子发生过程中对基因的调控存在差异，并且这些基因调控的差异可能是由于生殖颗粒内WAGO-1的性别特异性构象和生物物理特性所致。
