Spp38 MAPK participates in maintaining the homeostasis of hemolymph microbiota in Scylla paramamosain

The p38 mitogen-activated protein kinases (MAPKs) are evolutionally conserved from yeasts to mammals, and are involved in the regulation of cells response to various extracellular stimuli. In this study, the p38 MAPK gene (designated as Spp38) of mud crab (Scylla paramamosain) was identified and studied. Spp38 contained the conserved Thr-Gly-Tyr (TGY) motif and a Ala-Thr-Arg-Trp (ATRW) substrate-binding site. Spp38 transcript was ubiquitously expressed in all tissues examined, with the highest expression found in muscle and hepatopancras. Quantitative real-time PCR revealed that Spp38 was upregulated in hemocytes and hepatopancras after infection with Vibrio parahemolyticus and Lipopolysaccharides (LPS). Reporter gene assays indicated that Spp38 activated the expression of anti-lipopolysaccharides (SpALF1 - SpALF6) in S. paramamosian. RNA interference (RNAi)-mediated knockdown of Spp38 or inhibition of Spp38 by SB203580 decreased the expression levels of SpALF1-6 and dual oxidase (SpDuox1 and SpDuox2) in S. paramamosian, which consequently reduced reactive oxygen species (ROS) production thereby significantly increasing the bacterial count in the hemolymph of mud crabs. Similarly, there was a significant reduction in bacterial clearance ability of hemolymph after Spp38 knockdown followed by V. parahemolyticus infection. Taken together, the current data indicated that Spp38 could play a vital role in maintaining the homeostasis of hemolymph microbiota in S. paramamosain.