Cesari Eleonora, Loiarro Maria, Naro Chiara, Pieraccioli Marco, Farini Donatella, Pellegrini Livia, Pagliarini Vittoria, Bielli Pamela, Sette Claudio
Department of Neuroscience, Section of Human Anatomy, Catholic University of the Sacred Heart, 00168, Rome, Italy.
Laboratory of Neuroembryology, IRCCS Fondazione Santa Lucia, 00143, Rome, Italy.
Cell Death Dis. 2020 Apr 17;11(4):240. doi: 10.1038/s41419-020-2443-y.
Homologous recombination and chromosome segregation in meiosis rely on the timely expression of two splice variants of the endonuclease SPO11, named α and β, which respectively skip or include exon 2. However, in spite of its physiological importance, the mechanism underlying Spo11 alternative splicing in meiosis is still unknown. By screening the activity of factors that are predicted to bind the alternatively spliced region of Spo11, we identified hnRNPH as a key regulator of SPO11α splicing in mouse spermatocytes. Although hnRNPH was not upregulated in meiosis concomitantly with the switch in splicing, its recruitment to Spo11 pre-mRNA was favored by selective modulation of RNA polymerase II (RNAPII) phosphorylation and processivity in proximity of exon 2. The hnRNPH binding sites were localized near those of splicing factors that promote SPO11β splicing, suggesting that hnRNPH favors exon 2 skipping by competing out positive regulators. Indeed, hnRNPH binds proximal to a consensus motif for Sam68, a positive regulator of SPO11β splicing in vitro and in vivo, and it interferes with Sam68 binding to the Spo11 pre-mRNA. Thus, our work reveals that modulation of RNAPII dynamics in concert with hnRNPH recruitment exerts a combinatorial control of the timely regulated Spo11 splicing during meiosis.
减数分裂中的同源重组和染色体分离依赖于核酸内切酶SPO11的两种剪接变体α和β的适时表达，这两种变体分别跳过或包含外显子2。然而，尽管其具有生理重要性，但减数分裂中Spo11可变剪接的潜在机制仍然未知。通过筛选预测与Spo11可变剪接区域结合的因子的活性，我们确定hnRNPH是小鼠精母细胞中SPO11α剪接的关键调节因子。尽管hnRNPH在减数分裂中并未随着剪接转换而同时上调，但其被募集到Spo11前体mRNA上受到外显子2附近RNA聚合酶II（RNAPII）磷酸化和持续合成能力的选择性调节的促进。hnRNPH结合位点位于促进SPO11β剪接的剪接因子的结合位点附近，这表明hnRNPH通过竞争排除正调节因子来促进外显子2的跳过。事实上，hnRNPH在体外和体内均与SPO11β剪接的正调节因子Sam68的共有基序近端结合，并干扰Sam68与Spo11前体mRNA的结合。因此，我们的研究表明，RNAPII动态变化与hnRNPH募集协同作用，对减数分裂期间Spo11剪接的适时调控发挥组合控制作用。
