PTBP2-dependent alternative splicing regulates protein transport and mitochondria morphology in post-meiotic germ cells.

Abstract

The RNA binding protein PTBP2 (polypyrimidine tract binding protein 2) is a key regulator of tissue-specific alternative RNA splicing. In the testis, PTBP2 is expressed in meiotic and post-meiotic germ cells (spermatocytes and spermatids, respectively). In these cells, PTBP2 is required for proper alternative mRNA splicing for over 200 genes, disproportionately affecting genes encoding proteins involved in protein trafficking via transport vesicles. In this study, we used electron microscopy to test the hypothesis that protein trafficking is impaired in the absence of PTBP2, and to further investigate why spermatogenesis abruptly ceases in PTBP2-deficienct spermatids. Ultrastructural analysis shows that protein trafficking in spermatids is aberrant in the absence of PTBP2. Unexpectedly, we also found that mitochondria morphology and number are significantly altered in PTBP2-deficient spermatids, consistent with increased mitochondria fission. Furthermore, we show that genes with key roles in mitochondria dynamics and function are post-transcriptionally regulated by PTBP2 and in different stages of spermatogenesis. Collectively, the data provide ultrastructural evidence that alternative splicing regulation by PTBP2 during spermatogenesis is critical for proper regulation of protein trafficking and mitochondria morphology.

Widespread association of the Argonaute protein AGO2 with meiotic chromatin suggests a distinct nuclear function in mammalian male reproduction

“…5 C). The set of head morphology defects observed in Ago2 cKO sperm parallels the defects expected following depletion of several of the proteins down-regulated in Ago2 cKO postmeiotic germ cells, including HMGB2, YBX2, and PTBP2 ( Catena et al 2006 ; Snyder et al 2015 ; Hannigan et al 2018 ; Lorès et al 2018 ). Further, at least two of the transcription factors with binding motifs enriched among the set of dual AGO2 ChIP and eCLIP targets (NF1 and STAT4) ( Fig.…”

A nuclear role for the Argonaute protein AGO2 in mammalian gametogenesis

“…Defects in formation of the acrosome, the organelle containing enzymes that break down the zona pellucida at fertilization, are the major cause of sperm head defects (Perrin et al, 2013), and evaluation of acrosome formation by lectin-peanut agglutinin staining showed that the acrosome was small and improperly formed in Ago2 cKO sperm compared to controls (Figure 6B). The set of head morphology defects observed in Ago2 cKO sperm parallels the defects expected following depletion of several of the proteins downregulated in Ago2 cKO post-meiotic germ cells, including HMGB2, YBX2, and PTBP2 (Catena et al, 2006;Hannigan et al, 2018;Lorès et al, 2018;Snyder et al, 2015). Further, at least two of the transcription factors with binding motifs enriched among the set of dual AGO2 ChIP and CLIP targets (NF1 and STAT4; Figure 5J) are known to regulate sperm head formation (Chohan et al, 2018;Herrada and Wolgemuth, 1997).…”