Regulation Of Key Genes In Muskrat Apoptosis Pathway On Scent Gland Development

Muskrat(Ondatra zibethicus)is a kind of economic animal that reproduces seasonally.The morphology of the scent gland in its tail abdomen shows obvious seasonal changes,that is,the development of the muskrat during the breeding period expands rather than shrinks and becomes smaller during the breeding period,and is finally replaced by connective tissue.The muskrat fragrance secreted by adult male muskrat during the breeding period has the same main components and pharmacological effects as natural muskrat,and has the potential to replace natural muskrat.Therefore,it is particularly necessary to explore the mechanism of fragrance gland development and fragrance secretion so as to increase the yield of muskrat fragrance.Apoptosis plays an important role in the development and homeostasis of organisms.Apoptosis can be triggered by mitochondria,endoplasmic reticulum stress and death receptor activation.This study collected in different months of muskrat musk glands,mediated by fluorescence quantitative PCR technique to detect mitochondrial apoptotic pathways related genes of Cyt C,Apaf-1,caspase-9,SMAC,the expression of AIF and PARP,and endoplasmic reticulum stress pathway mediated apoptosis related gene perk,Ire-1,traf-2,the expression of chop and atf-6,understand its mRNA expression rule,the results of inspection by western blot,which discuss apoptotic pathways in the gene regulation function of muskrat musk gland development.The results showed that in the mitochondria-mediated apoptosis pathway,the expression levels of Cyt C,Apaf-1,caspase-9 and SMAC were all up-regulated during the rapid atrophy-phase of aromatic glands,and the expression level of Cyt C gene reached the highest in August(P<0.05,the difference was significant).Apaf-1 gene was gradually upregulated in June,and reached the highest in August(P<0.05,the difference was significant).caspase-9 gene was up-regulated sharply and reached the highest value in September(P<0.01,the difference was extremely significant).The expression level of SMAC gene reached the highest in September(P<0.01,the difference was extremely significant).There was no significant difference in the expression of AIF and PARP during the development of scent gland.In the endoplasmic reticulum stress-mediated apoptosis pathway,the expressions of Ire-1,chop,perk,traf-2 and atf-6 were all upregulated during the peak of fragrance glands.The expression of perk gene was the highest in May(P<0.01,the difference was extremely significant).The transcription level of chop gene was the highest in June(P<0.01,the difference was extremely significant).The expression of atf-6 gene reached the highest level in June(P<0.05,the difference was significant).The transcription level of traf-2 gene was upregulated to the highest in May(P<0.01,the difference was extremely significant).The expression of Ire-1 gene reached the highest level in July(P<0.01,the difference was extremely significant).The trend of Cyt C,caspase,chop and perk immunoblotting was consistent with that of qPCR.The results showed that the transcription levels of atf-6,chop,traf-2 and perk genes in the apoptosis pathway mediated by ER stress were high in the fragrance glands during the fragrance secreting stage,which played a regulatory role in stabilizing the protein synthesis rate and correcting the protein synthesis errors,and promoted cell apoptosis to ensure the stability of fragrance glands during the continuous fragrance secreting stage when the pressure of ER synthesis was too high.In the rapid atrophy of scent glands,mitochondria-mediated apoptosis pathway related genes such as caspase-9 induce cell apoptosis and promote rapid cell apoptosis.The ER stress-mediated apoptosis pathway Ire-1 and traf-2 are also involved in the regulation of scent glands atrophy.However,in the mitochondrial apoptosis pathway,the expression levels of AIF and PARP genes that can induce non-caspase-dependent apoptosis do not change significantly during the development cycle of fragrance glands,and only play a secondary regulatory role in the process of fragrance secretion and atrophy of fragrance glands.The results of this study suggest that the aroma secretion of muskrat can be stable by promoting the expression of atf-6,Ire-1 and perk genes during aroma secretion.Inhibition of the expression of Cyt C,Apaf-1 and caspase-9 genes can delay the atrophy of incense glands,prolong the time of incense secretion,and increase the amount of incense secretion of incense glands.