title: Identification, expression pattern, cellular location and potential role of the caveolin-1 gene from Artemia sinica.
abstract: Caveolins are integral membrane proteins that serve as scaffolds to recruit numerous signaling molecules. Caveolins play an important role in membrane trafficking, signal transduction, substrate transport and endocytosis in differentiated cells. In this study, a caveolin-1 gene from Artemia sinica (As-cav-1) was successfully cloned for the first time. The full-length cDNA of As-cav-1 comprises 974 bp, with a 675 bp open reading frame (ORF) that encodes a polypeptide of 224 amino acids with a caveolin scaffolding domain (CSD) and two transmembrane domains. Multiple sequence alignment revealed that the putative As-CAV-1 protein sequence was relatively conserved across species, especially in the CSD domain. Real-time PCR revealed high levels of the As-cav-1 transcript at 0h of embryo development. Furthermore, As-cav-1 transcripts were highly upregulated under high salinity (200‰) and low temperature stresses (15°C). To further characterize As-cav-1, recombinant pET30a-cav-1 protein was expressed using a prokaryotic expression system. The recombinant protein comprised 290 amino acids with a theoretical molecular weight of 32kDa, and a predicted isoelectric point of 5.6. Western blotting of the expression levels of As-CAV-1 during different embryo development stages revealed that As-CAV-1 levels decreased gradually during development stages from 0 h to 40 h, and increased at 3d. Furthermore, western blotting showed that As-CAV-1 was upregulated to its highest expression level by low temperature stress (15°C) and high salinity. Confocal laser microscopy analysis, using antibodies generated against the recombinant As-CAV-1 protein, showed that As-CAV-1 was mostly located in the cell membrane. Our results suggested that As-cav-1 plays a vital role in protecting embryos from high salt damage and low temperature stress, especially during post-diapause embryonic development.
authors.affiliation: College of Life Sciences, Liaoning Normal University, Dalian 116081, China.
authors.forename: Xuejie
authors.initials: X
authors.lastname: Li
authors.affiliation: College of Life Sciences, Liaoning Normal University, Dalian 116081, China.
authors.forename: Feng
authors.initials: F
authors.lastname: Yao
authors.affiliation: College of Life Sciences, Liaoning Normal University, Dalian 116081, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Zhang
authors.affiliation: College of Life Sciences, Liaoning Normal University, Dalian 116081, China.
authors.forename: Cheng
authors.initials: C
authors.lastname: Cheng
authors.affiliation: College of Life Sciences, Liaoning Normal University, Dalian 116081, China.
authors.forename: Bing
authors.initials: B
authors.lastname: Chu
authors.affiliation: College of Life Sciences, Liaoning Normal University, Dalian 116081, China.
authors.forename: Yan
authors.initials: Y
authors.lastname: Liu
authors.affiliation: College of Life Sciences, Liaoning Normal University, Dalian 116081, China.
authors.forename: Yanli
authors.initials: Y
authors.lastname: Mei
authors.affiliation: College of Life Sciences, Liaoning Normal University, Dalian 116081, China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Wu
authors.affiliation: Department of Biology, Dalian Medical University, Dalian 116044, China. Electronic address: zouxiangyang@126.com.
authors.forename: Xiangyang
authors.initials: X
authors.lastname: Zou
authors.affiliation: College of Life Sciences, Liaoning Normal University, Dalian 116081, China. Electronic address: houlin@lnnu.edu.cn.
authors.forename: Lin
authors.initials: L
authors.lastname: Hou
mesh_terms: D000222:Adaptation, Physiological; D000595:Amino Acid Sequence; D000818:Animals; D001156:Artemia / Q000166:cytology / Q000196:embryology / Q000235:genetics / Q000378:metabolism*; D060829:Arthropod Proteins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D051242:Caveolin 1 / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D058639:Cold-Shock Response; D017124:Conserved Sequence; D008969:Molecular Sequence Data; D010802:Phylogeny; D021381:Protein Transport; D060888:Real-Time Polymerase Chain Reaction; D055049:Salt Tolerance / Q000235:genetics; D059467:Transcriptome
chemical_list: D060829:Arthropod Proteins; D051242:Caveolin 1
keywords: Artemia sinica; Caveolin-1; Cellular location; Development; Prokaryotic expression