Presentation Authors: Jason Scovell*, Abhishek Seth, Juan Bournat, Joshua Moore, Minerva Solis, Adam Szafran, Carolina Jorgez, Houston, TX
Introduction: We identified PRSS50 as a testis specific serine protease with a possible role in meiosis and male fertility. PRSS50 is thought to signal though NF-ÎºB. One testis specific target of NF-ÎºB is the centrosome protein LRWD1. Our objective was to determine the in vivo effect of PRSS50 loss on fertility and sperm production and identify potential mechanisms driving this phenotype.
Methods: We generated a Prss50 knock-out (KO) mouse using CRISPR/Cas9. Fertility was assessed by 6-month mating (n=10 each group). Testicular histology, sperm analysis, and protein-analysis was performed. Testicular ultrastructural morphology was assessed by transmission electron microscopy (TEM).
Results: PRSS50 expression is localized to the cytoplasm of spermatocytes beginning at post-natal day 14 (beginning of meiosis) and continues throughout adulthood as well as in the sperm midpice. PRSS50 expression patterns were similar between mouse and human sperm. 20% of Prss50-KO males were infertile compared to 0% of WT. Prss50-KO males were severely sub-fertile producing fewer litters and fewer numbers of pups per litter (KO: 63% fewer pups than WT, p < 0.01). All Prss50-KO mice seminiferous tubules show abnormalities with different degrees between mice including: Sertoli cell only (SCO) tubules, a high degree of vacuolation, altered wave of maturation, multinucleated and symplastic germ cells, and increased residual bodies. TEM demonstrated multiple testicular defects including but not limited to multiple spermatids without membranes dividing the nucleus, spermatids lacking or having additional centrosomes, spermatids with multiple axonemes these findings suggested impaired cytokynesis. KO mice had a decreased percentage of normal sperm compared to WT mice (30% vs. 72%, p < 0.01) with defects including sperm heads only (61% vs. 21%, p < 0.01) and multi head/tail sperms. Western blot demonstrated a 2.1-fold reduction in testicular LRWD1 (p < 0.05). LRWD1 plays an important role in centromere formation and histone modification. As a potential consequence of lacking PRSS50, spermatocyte and spermatid H3K9me3 levels were reduced and the pattern of expression was mis-localized.
Conclusions: Prss50-KO unique testicular and sperm morphology defects supports its role in spermatogenesis and meiosis. Lack of PRSS50 signaling through NFkB-LRWD1 may be a causative mechanism in abnormal sperm development in Prss50-KO mice.
Source of Funding: National Institute of General Medical Sciences of the National Institutes of Health under Award Number T32GM088129.