Identification and Analysis of the Core Region of Goat NPC1 Gene Promoter
Niemann-pick C1(NPC1)encoded by the NPC1 gene participates in lipid raft formation,endocytosis of pathogenic microorganisms and intracellular transport of endosomes.We aimed to study the transcriptional regulation mechanism of NPC1 gene in Hainan black goats and to provide a theoretical reference for studying the interactions between pathogenic microorganisms and host cells.First,the 1466 bp promoter sequence of the goat NPC1 gene was analyzed and cloned for the study.The 5'-end truncated NPC1 promoters were amplified and constructed into the dual-luciferase reporter vector to screen the core promoter region of the NPC1 gene.Then,the key transcription factors in the core promoter region were further predicted and analyzed and the deletion vectors of transcription factor binding sites were constructed to screen the key transcription factor binding sites of NPC1 gene.The results showed that the core region of goat NPC1 gene promoter located from-195 bp to-60 bp upstream of the transcription initiation site and there were E2F3(-134/-120 bp),SREBP-1(-107/-96 bp)SP1(-68/-62 bp)and other transcription factor binding sites.The results of the dual-luciferase reporter assay showed that deletion of three transcription factor binding sites E2F3,SREBP-1 and SP1 significantly decreased the transcriptional activity of goat NPC1 gene promoter(P＜0.01)and deletion of SREBP-1 binding site significantly decreased the transcriptional activity of NPC1 gene.These results suggested that SREBP-1 transcription factor played an important role in the transcriptional activity of NPC1 gene in Hainan black goats.In this experiment,identification of the core region(-195/-60 bp)of goat NPC1 gene promoter and the key transcription factor binding site SREBP-1 in this region gained better understanding of the molecular mechanism of goat NPC1 gene-mediated interaction between pathogenic microorganisms and hosts.
Hainan black goatNPC1 genetranscriptional regulationcore promoterSREBP-1
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