Cloning of LHR and expression analysis during the reproductive cycle in female yellow catfish(Pelteobagrus fulvidraco)
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Graphical Abstract
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Abstract
The complete cDNA sequence of luteinizing hormone receptor(LHR)was cloned through degenerate primer PCR amplification and SmartTM Race technology.The length of the cDNA of yellow catfish(Pelteobagrus fulvidraco) LHR gene is 2 524 bp and encodes the protein of 701 amino acids.This gene belongs to glycoprotein hormone receptor(GpHR)family,which contains the conserved seven transmembrane helix domains(TM helix).Meanwhile the amino acids sequence contains nine Leucine-rich repeats(LRRs).Potential N-linked glycosylation sites are 29NFTC, 82NVSR, 203NGSR, 548NLTV in yellow catfish LHR gene and phosphorylation site predictions identified 24 Ser,6 Thr and 5 Tyr potential phosphorylation sites.By Clustal X and MEGE 4.0 alignment revealed the highly conserved in GpHRs’ specific signature sequences: 287CCAF, 471ERW, 584FTD and 657NPFLY.The only one protein kinase C phosphorylation site is 400S.Tissue distribution pattern showed preferential expression of LHR in ovary,and trace amounts were detected in brain,kidney,heart,liver and intestine(brain>kidney>heart>liver>intestine).Plasma 17β-Estradiol(E2)levels in reproductive cycle were detected by radio immunoassay(RIA).The result showed that the highest peak in reproductive cycle is stage Ⅳ.The high transcript level of LHR in ovary was revealed in the reproductive phase of ovarian cycle,which was the relatively high expression from Ⅲ to Ⅴ phase and the highest level in stage Ⅴ.And we also analyzed the brain expression level in reproductive cycle,found the highest level in stage Ⅳ consistent with the level of E2.Results demonstrated the gene of LHR in brain and ovary might also be involved in the processes which preferentially promote vitellogenesis and regulate ovulation maturation and ovulate either indirectly or directly.
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