Proteomic analysis to explore potential mechanism underlying pseudomale sperm defect in Cynoglossus semilaevis

Abstract

Females of Chinese tongue sole (Cynoglossus semilaevis) grow 2–4 times faster and bigger than males, but some genetic females can sex reverse into phenotypic males (called pseudomales, ZW karyotype) that presents similar slow growth to normal males. Pseudomales could only produce Z-type sperm, of which the inheritance epigenetic pattern makes their offspring tend to be pseudomales. In this study, to explore the potential mechanism underlying sperm defect (W sperm absence) of pseudomales, we compared the proteomic profiles of pseudomale and male semen samples. Totally, 783 differentially expressed proteins (DEPs) including 495 up-regulated and 288 down-regulated proteins were identified in pseudomale group. Proteins involved with proteasome were enriched by GO and KEGG analyses. A protein-protein interaction (PPI) network was further constructed to understand the relationship of the DEPs. Interestingly, ten molecular chaperones including four heat shock proteins (Hsp90aa1.2, Hsp90ab1, Hsp90b1, and Hsc71/Hspa8) and six chaperonin containing T-complex protein 1 (TCP1) subunits (Cct1, Cct2, Cct3, Cct4, Cct5, and Cct7) were identified, and they were up-regulated in pseudomale. Ubiquitin-related proteins including ubiquitin like 4 A (Ubl4a) and NEDD8 ubiquitin like modifier (NEDD8), proteasome subunits such as proteasome 26S subunit, non-ATPase 14 (Psmd14) and proteasome 26S subunit, ATPase 6 (Psmc6), and redox-related proteins like thioredoxin-disulfide reductase (Txnrd3) and thioredoxin (Trx1) were also found in PPI network. In addition, up-regulation of lysophosphatidylcholine acyltransferase 3 (Lpcat3), acyl-coenzyme A (CoA) synthetase long-chain family members (Acsl1a and Acsl6), glutathione synthetase (GSS), and transferrin receptor 1 (Trf1) but down-regulation of ferritin were observed in pseudomale. Dysfunction of chaperones targeting client polypeptides for degradation via the ubiquitin-proteasome system (UPS) or ferroptosis might be a possible cause of pseudomale sperm abnormalities. The study provides a fundamental basis for explaining W sperm absence, which may have vast application in the future fish breeding with high-female-ratio.