title: 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.
authors.affiliation: National Centre for Aquatic Animal Health, Cochin University of Science and Technology, Cochin 682 016, India.
authors.forename: Vrinda
authors.initials: V
authors.lastname: S
authors.affiliation: National Institute of Oceanography, RC Cochin, Dr. Salim Ali Road, Cochin 682 018, India.
authors.forename: Jasmin
authors.initials: J
authors.lastname: C
authors.affiliation: Bioinformatics Facility, Rajiv Gandhi Centre for Biotechnology, Thiruvananthapuram 695 014, India.
authors.forename: Sivakumar
authors.initials: S
authors.lastname: K C
authors.affiliation: National Centre for Aquatic Animal Health, Cochin University of Science and Technology, Cochin 682 016, India.
authors.forename: Seena
authors.initials: S
authors.lastname: Jose
authors.affiliation: National Centre for Aquatic Animal Health, Cochin University of Science and Technology, Cochin 682 016, India.
authors.forename: Blessy
authors.initials: B
authors.lastname: Jose
authors.affiliation: Department of Marine Biology, Microbiology and Biochemistry, Cochin University of Science and Technology, Cochin 682 016, India.
authors.forename: Rosamma
authors.initials: R
authors.lastname: Philip
authors.affiliation: National Centre for Aquatic Animal Health, Cochin University of Science and Technology, Cochin 682 016, India. Electronic address: isbsingh@gmail.com.
authors.forename: Bright Singh
authors.initials: BS
authors.lastname: I S
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D057134:Antibodies, Neutralizing / Q000494:pharmacology; D017756:Aquaculture; D060829:Arthropod Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D001681:Biological Assay; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D017124:Conserved Sequence; D015195:Drug Design; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D005123:Eye; D005260:Female; D007447:Invertebrate Hormones / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D008958:Models, Molecular; D009490:Neurosecretory Systems / Q000166:cytology / Q000187:drug effects / Q000502:physiology; D033561:Penaeidae / Q000166:cytology / Q000187:drug effects* / Q000502:physiology; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D012102:Reproductive Control Agents / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D016415:Sequence Alignment; D040681:Structural Homology, Protein; D013879:Thioredoxins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D047048:Vitellins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D014818:Vitellogenesis / Q000187:drug effects*; D014819:Vitellogenins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism
chemical_list: D057134:Antibodies, Neutralizing; D060829:Arthropod Proteins; D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D007447:Invertebrate Hormones; D011993:Recombinant Fusion Proteins; D012102:Reproductive Control Agents; D047048:Vitellins; D014819:Vitellogenins; C073189:vitellogenesis inhibiting hormone; D013879:Thioredoxins
keywords: CHH family hormone; Fusion/chimeric protein; GIH polyclonal antisera; Gonad-inhibiting hormone (GIH); Penaeus monodon; Thioredoxin-fused protein; XO-SG complex