Molecular characterization and functional analyses of a diapause hormone receptor-like gene in parthenogenetic Artemia.
In arthropods, mature females under certain conditions produce and release encysted gastrula embryos that enter diapause, a state of obligate dormancy. The process is presumably regulated by diapause hormone (DH) and diapause hormone receptor (DHR) that were identified in the silkworm, Bombyx mori and other insects. However, the molecular structure and function of DHR in crustaceans remains unknown. Here, a DHR-like gene from parthenogenetic Artemia (Ar-DHR) was isolated and sequenced. The cDNA sequence consists of 1410bp with a 1260-bp open reading frame encoding a protein consisting of 420 amino acid residues. The results of real-time PCR (qRT-PCR) and Western blot analysis showed that the mRNA and protein of Ar-DHR were mainly expressed at the diapause stage. Furthermore, we found that Ar-DHR was located on the cell membrane of the pre-diapause cyst but in the cytoplasm of the diapause cyst by analysis of immunofluorescence. In vivo knockdown of Ar-DHR by RNA interference (RNAi) and antiserum neutralization consistently inhibited diapause cysts formation. The results indicated that Ar-DHR plays an important role in the induction and maintenance of embryonic diapause in Artemia. Thus, our findings provide an insight into the regulation of diapause formation in Artemia and the function of Ar-DHR.
College of Life Sciences, Zhejiang University, Hangzhou, Zhejiang 310058, PR China.
Hui-Li
HL
Ye
College of Life Sciences, Zhejiang University, Hangzhou, Zhejiang 310058, PR China.
Dong-Rui
DR
Li
College of Life Sciences, Zhejiang University, Hangzhou, Zhejiang 310058, PR China.
Jin-Shu
JS
Yang
College of Life Sciences, Zhejiang University, Hangzhou, Zhejiang 310058, PR China.
Dian-Fu
DF
Chen
Laboratory of Aquaculture & Artemia Reference Center, Ghent University, Belgium.
Stephanie
S
De Vos
Laboratory of Aquaculture & Artemia Reference Center, Ghent University, Belgium.
Marnik
M
Vuylsteke
Laboratory of Aquaculture & Artemia Reference Center, Ghent University, Belgium.
Patrick
P
Sorgeloos
Laboratory of Aquaculture & Artemia Reference Center, Ghent University, Belgium.
Gilbert
G
Van Stappen
Laboratory of Aquaculture & Artemia Reference Center, Ghent University, Belgium.
Peter
P
Bossier
Department of Applied Biological Chemistry, The University of Tokyo, Yayoi, Bunkyo, Tokyo 113-8657, Japan. Electronic address: anagahi@mail.ecc.u-tokyo.ac.jp.
Hiromichi
H
Nagasawa
College of Life Sciences, Zhejiang University, Hangzhou, Zhejiang 310058, PR China. Electronic address: w_jyang@zju.edu.cn.
Wei-Jun
WJ
Yang
D000595:Amino Acid Sequence / Q000235:genetics; D000818:Animals; D001156:Artemia / Q000235:genetics*; D012831:Bombyx / Q000235:genetics / Q000254:growth & development; D064666:Diapause, Insect / Q000235:genetics*; D005260:Female; D005786:Gene Expression Regulation; D019476:Insect Proteins / Q000235:genetics; D009479:Neuropeptides / Q000235:genetics*; D010312:Parthenogenesis / Q000235:genetics; D011498:Protein Precursors / Q000235:genetics; D034622:RNA Interference; D012333:RNA, Messenger / Q000235:genetics
C092658:DH-PBAN precursor protein, Bombyx mori; D019476:Insect Proteins; D009479:Neuropeptides; D011498:Protein Precursors; D012333:RNA, Messenger; C075027:diapause hormone
Antiserum neutralization; Artemia; Diapause; Diapause hormone (DH); Diapause hormone receptor (DHR); RNAi