Regulating gonad inhibition and vitellogenin/vitellin induction in Penaeus monodon using mature GIH fusion protein and polyclonal antisera

Abstract

Gonad inhibiting hormone (GIH), type II class of the CHH family neuropeptides, is released by the neurohaemal XO-SG complex of the eyestalk. The inhibitory function of GIH has a pivotal role in gonad development and reproduction. In this study, we report the expression and production of a thioredoxin-fused mature GIH protein (mf-PmGIH) of Penaeus monodon in a bacterial system and its use as antigen to raise polyclonal antiserum (anti-mf-PmGIH). The mature GIH gene of 237bp that codes for 79 amino acids, was cloned into the Escherichia coli thioredoxin gene fusion expression system. The expression vector construct (mf-PmGIH+pEt32a+) upon induction produced 32.16kDa mature GIH fusion protein (mf-PmGIH)·The purified fusion protein was used as exogenous GIH and as antigen to raise polyclonal antisera. The fusion protein when injected into juvenile shrimp significantly reduced vitellogenin/vitellin levels by 31.55% within 72h in comparison to the controls showing the gonad inhibiting property. Vitellogenin/vitellin levels were significantly induced by 74.10% within 6h when polyclonal antiserum (anti-mf-PmGIH - 1:500) was injected in P. monodon. Anti-mf-PmGIH immunolocalized GIH producing neurosecretory cells in the eyestalk of P. monodon. The present manuscript reports an innovative means of gonad inhibition and vitellogenin/vitellin induction with thioredoxin fused GIH and antisera developed.