Function And Transcriptional Regulation Of Parathyroid Hormone-like Hormone Gene During Follicle Growth And Its Genetic Effects On Egg Production Traits In Hens

Parathyroid hormone-like hormone(PTHLH)was initially identified as the tumor product that is responsible for most instances of humoral hypercalcemia of malignancy.PTHLH regulates cell growth and differentiation,bone development and lactation,embryonic and fetal development and survival,morphogenesis,and placental calcium transport via intracellular,paracrine and endocrine pathways.The parathyroid hormone(PTH)and PTHLH genes are evolutionarily related and share a common receptor,the PTH/PTHLH receptor(PTH1R).In our previous study,by RNA-seq,we found that the relative mRNA expression of PTHLH is similar to that of FSHR in the single small yellow follicle(SY,6-8mm).In this study,the expression and regulation of PTHLH and PTH1 R,and polymorphism in PTHLH promoter region were investigated.We first determined the transcript of chicken ovarian PTHLH transcript by 5?-and 3?-RACE,the product is same to the first transcript in NCBI.The mRNA of PTHLH and PTH1 R is detected at the highest levels within SY follicles,mainly expressed in theca cells except that PTHLH m RNA was mainly expressed in granulosa cells in post-hierarchal follicles,while the protein expression of both were without difference.And PTHLH induces cells proliferation and pre-hierarchal granulosa cells differentiation,PTHLH also induces the mRNA expression of the essential steroidogenic enzymes STAR and CYP11 A.Treatment with FSH increased PTHLH mRNA expression in pre-hierarchal theca cells and hierarchal granulosa cells.Two critical regions regulating chicken PTHLH transcription were revealed,each of which harbored a SNP: C>T(rs72520524)for AP-1 and a SNP: A>G(rs72518862).Compared with other diplotypes,hens with diplotype AC/GT were younger at first laying and laid more eggs at 32 weeks.Based on the luciferase assay,the haplotype(G-1827-T-165)with double mutationshad the greatest promoter activity of chicken PTHLH transcription.The SNP rs72520524,which changed from C to T at-165 site,increased the binding activity of transcription factor AP-1.Collectively,we provide the first evidence of PTHLH expression and regulation in hen follicles and identify polymorphisms in its promoter region.Chicken PTHLH plays an important role in follicle selection by stimulating cell proliferation and steroidogenesis.Polymorphisms in chicken PTHLH promoter region are associated with egg laying traits by affecting the binding of transcription factor AP-1.