title: A core component of the CUL4 ubiquitin ligase complexes, DDB1, regulates spermatogenesis in the Chinese mitten crab, Eriocheir sinensis.
abstract: Studies in mammals have shown that damaged DNA-binding protein 1 (DDB1) is a multifunctional protein that recognizes UV-induced DNA lesions and activates nucleotide excision repair process, and could also be a linker protein for Cullin4 in ubiquitination to regulate cell cycle progression. However, there are few studies of DBB1 in crustaceans. In this study, a cDNA representing the DDB1 gene from Eriocheir sinensis (Es-DDB1) was cloned successfully. The full length Es-DDB1 cDNA comprises 4871 nucleotides, and encodes an open-reading frame (ORF) of 1137 amino acid residues. Bioinformatics' analysis showed that the domains and structure of Es-DDB1 have been highly conserved during evolution. Antibodies against Es-DDB1 and Es-Cul4 were raised using a prokaryotic expression system. Moreover, a co-immunoprecipitation assay showed that Es-DDB1 could bind Es-Cul4 in the testis of Eriocheir sinensis. Furthermore, quantitative real-time PCR and Western blotting showed high expression in the testis, particularly during the spermatocyte stage. Immunofluorescence assays showed that Es-DDB1 was mainly distributed in the cytoplasm in the early and middle developmental stages. These results indicated that Es-DDB1 might play a key role in spermatogenesis of E. sinensis.
authors.affiliation: Laboratory of Invertebrate Immunological Defense & Reproductive Biology, School of Life Science, East China Normal University, Shanghai, China.
authors.forename: AnYu
authors.initials: A
authors.lastname: Fang
authors.affiliation: Laboratory of Invertebrate Immunological Defense & Reproductive Biology, School of Life Science, East China Normal University, Shanghai, China.
authors.forename: XueJie
authors.initials: X
authors.lastname: Li
authors.affiliation: Laboratory of Invertebrate Immunological Defense & Reproductive Biology, School of Life Science, East China Normal University, Shanghai, China.
authors.forename: YuanLi
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Laboratory of Invertebrate Immunological Defense & Reproductive Biology, School of Life Science, East China Normal University, Shanghai, China.
authors.forename: DiYue
authors.initials: D
authors.lastname: Pan
authors.affiliation: Laboratory of Invertebrate Immunological Defense & Reproductive Biology, School of Life Science, East China Normal University, Shanghai, China. Electronic address: qwang@bio.ecnu.edu.cn.
authors.forename: Qun
authors.initials: Q
authors.lastname: Wang
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D003386:Brachyura / Q000235:genetics* / Q000502:physiology*; D003001:Cloning, Molecular; D017124:Conserved Sequence; D018076:DNA, Complementary / Q000235:genetics; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D015870:Gene Expression; D008297:Male; D008958:Models, Molecular; D010802:Phylogeny; D011487:Protein Conformation; D017386:Sequence Homology, Amino Acid; D013091:Spermatogenesis / Q000235:genetics* / Q000502:physiology*; D014018:Tissue Distribution; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*
chemical_list: D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D044767:Ubiquitin-Protein Ligases
keywords: Co-immunoprecipitation; DDB1; Eriocheir sinensis; Prokaryotic expression; Spermatogenesis