Senegalese sole (Solea senegalensis) is a species with a high commercial value that exhibits a reproductive dysfunction in males born and raised in captivity (F1) that hinders their sustainable culture. The present study evaluates the sperm quality and dopaminergic pathway of males born in the wild environment and of F1 males. Traditional sperm analyses were performed, finding only significant differences in curvilinear velocity (VCL) and no significant differences in viability and total motility. No differences in global sperm methylation were observed either in spermatozoa or brain between the two groups (F1 and wild-born males). However, our results point to a different sperm molecular signature between wild fish and fish born in captivity, specifically the differential expression in miR-let7-d and miR-200a-5p between these two groups. miR-let7-d has been correlated with spermatogenesis and sex preferences, whereas the miR-200 family is implied in target innervation of dopaminergic neurons in zebrafish. When we analysed the dopaminergic pathway, no differences were found in terms of different mRNA expression of dopaminergic markers. However, some differences were detected in terms of tyrosine hydroxylase protein expression by western blot analysis, thus suggesting an altered post-transcriptional regulation in F1 males. The results of this study suggest that an altered sperm miRNA signature in F1 males could be one possible mode of transmission of reproductive dysfunction to the progeny.

Ubiquitination of the TrkA neurotrophin receptor in response to NGF is critical in the regulation of TrkA activation and functions. TrkA is ubiquitinated, among other E3 ubiquitin ligases, by Nedd4-2. To understand mechanistically how TrkA ubiquitination is regulated, we performed a siRNA screening to identify deubiquitinating enzymes and found that USP36 acts as an important regulator of TrkA activation kinetics and ubiquitination. However, USP36 action on TrkA was indirect because it does not deubiquitinate TrkA. Instead, USP36 binds to Nedd4-2 and regulates the association of TrkA and Nedd4-2. In addition, depletion of USP36 increases TrkA⅐Nedd4-2 complex formation, whereas USP36 expression disrupts the complex, resulting in an enhancement or impairment of Nedd4-2-dependent TrkA ubiquitination, respectively. Moreover, USP36 depletion leads to enhanced total and surface TrkA expression that results in increased NGF-mediated TrkA activation and signaling that augments PC12 cell differentiation. USP36 actions extend beyond TrkA because the presence of USP36 interferes with Nedd4-2-dependent Kv7.2/3 channel regulation. Our results demonstrate that USP36 binds to and regulates the actions of Nedd4-2 over different substrates affecting their expression and functions.