Cell signalling during male-female interactions by 'Schistosoma mansoni'

The genome of the human parasitic worm Schistosoma mansoni encodes over 260 protein kinases, important regulatory proteins that are well conserved through the evolution of eukaryotes. To understand protein kinase signal transduction and its downstream function during male-female interactions of S. mansoni, we investigated the temporal effects of excretory-secretory products (ESPs) produced by adult male worms over 24 h culture on protein kinase activities in female worms, and vice versa. Western blotting with anti-phospho Tyr/Ser/Thr antibodies revealed that the phosphorylation status of multiple proteins changed over 60 min in response to opposite sex ESPs. Adult male/female worms exposed to opposite sex ESPs displayed a rapid activation of p38 mitogen-activated protein kinase (p38 MAPK) and extracellular signal-regulated kinase (ERK), where activation was evident as early as 5 min, including in the tegument; such responses also occurred with adolescent worms. Phosphorylation (activation) of p38 MAPK and ERK was attenuated significantly by SB203580 and U0126, p38 MAPK and ERK inhibitors, respectively. Immunofluorescence and confocal laser scanning microscopy revealed that the activation occurred in different tissues including the parasite tegument matrix and sub-layers, the testes and ovaries. In addition, the motility of the parasites increased in response to the opposite sex ESPs, whereas incubation with SB203580 or U0126 for 1 h prior to exposure to ESPs profoundly reduced worm motility. Using biotinylated-ESPs and fluorescence confocal laser scanning microscopy, the adult male ESPs were observed bound to the surface of female worms and vice versa. Moreover, cell proliferation was investigated using Click-it EdU incorporation assays. Single sex adult worm cultures displayed significantly reduced cell proliferation after 5 days, however, there was a striking increase in cell proliferation in the testes, ovaries and tegument of worms exposed to opposite sex ESPs on day 5; this ESP-mediated recovery of cell proliferation was attenuated by both SB203580 and U0126. The ESP proteome, with and without extracellular vesicles (EVs), was also investigated. LC-MS/MS analysis of total and EV-depleted ESP fractions of male and female S. mansoni identified a total of 705 and 362 proteins, respectively, the importance of which are further discussed. To the best of our knowledge, this study is the first to report on male and female ESPs activating signal transduction processes in S. mansoni worms of the opposite sex and their importance in driving cell proliferation in both reproductive and somatic tissues. These findings provide valuable insight into male-female schistosome interactions and highlight future possibilities for the development of novel strategies for schistosome control to reduce human schistosomiasis, a disease which affects over 240 million people.