CDC23(Cell Division Cycle 23) And CDH1 Play Pivotal Roles In The Process Of Early Embryonic Development In Artemia Sinica

Artemia sinica is a crustacean,which is frequently used in developmental biology.It has strong resistant ability.Artemia become special diapause cysts in response to unfavourable environments,and restart the embryonic developmental stages when the situations return to normal.Anaphase promoting complex or cyclosome(APC/C)is a representative ubiquitin ligase;it degrades specific proteins in mitosis.CDH1 is one of the two cofactors of the APC/C,specifically binding and activating the APC/C complex.Cell division cycle 23 plays significant role in the spindle assembly checkpoint associated with CDH1,mediating ubiquitin-mediated degradation of cell-cycle proteins,and ensuring the right separation of chromosomes.Therefore CDC23 and CDH1 participate in cell cycle regulation,but the specific mechanisms of their activities remain unknown.In this study,a 1215 bp full-length cDNA of cdc23 from A.sinica(As-cdc23)was cloned for the first time;it encodes 146 amino acids,17.1kDa protein.Bioinformatics analysis showed As-CDC23 primarily localized in the nucleus,as a non-transmembrane hydrophilic non-secreted protein.Multiple sequence alignment analysis showed that CDC23 was a conserved protein,especially in the APC8 domain.Results of whole mount immunohistochemistry of As-CDH1 and immunofluorescence of As-CDC23 showed no tissue or organ specificity on the expression location.The expression levels of As-CDC23 were analysed by qPCR and Western blotting,results showed that the expression level was up-regulated from 0h to 15 h stage of embryonic development,and gradually reduced with the development of Artemia.But the expression levels showed a down-regulated trend under the temperature and salinity stress.Meanwhile,western blotting showed that the levels of cycle-related proteins,which conformed to their complicated degradation relationships during different embryo stages.Our research revealed that As-CDH1 and As-CDC23 play crucial roles in the process of early embryonic development in A.sinica,participate in the degradation of downstream proteins and regulate the cell cycle.Furthermore,these results have a significance meaning for further studies of embryonic development of A.sinica and CDC23 and CDH1 proteins in vertebrates.