Characterization and biological function analysis of the trim3a gene from zebrafish (Danio rerio).
The biological significance of tripartite motif (TRIM) proteins is increasingly being appreciated due to their roles in a broad range of biological processes that associated with innate immunity. In this study, we have described the structural and functional analysis of TRIM3a from zebrafish. Annotation of domain architectures found that the TRIM3a fulfills the TRIM-NHL rule of domain composition with a Filamin/ABP280 domain and NHL repeats at its C-terminal region. In addition, the mRNA expression level of TRIM3a was the highest in brain, and with a relatively higher level in spleen, liver, and gill. A strong expression starting at 36 h post fertilization (hpf) was observed by real-time PCR and could be detected in brain by in situ hybridization, suggesting that TRIM3a protein might play an important role in brain development in zebrafish. Considering that TRIM3a has a RING finger domain, we expressed and purified the TRIM3a protein and performed ubiquitylation assays, our results showed that TRIM3a underwent self-polyubiquitylation in combination with E1, UbcH5c, biotin-ubiquitin in vitro. Meanwhile, TRIM3a-R without the RING domain was expressed and purified as well, in vitro ubiquitylation assays showed that the self-ubiquitylation of TRIM3a was dependent on its RING domain, suggesting that TRIM3a might function as a RING finger E3 ubiquitin ligase.
Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing, PR China.
Xinshang
X
Zhang
Heng
H
Zhao
Yeyu
Y
Chen
Chao
C
Liu
Kun
K
Meng
Peilong
P
Yang
Yaru
Y
Wang
Guozeng
G
Wang
Bin
B
Yao
D000818:Animals; D001921:Brain / Q000254:growth & development / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D018507:Gene Expression Regulation, Developmental; D009928:Organ Specificity; D010802:Phylogeny; D054829:RING Finger Domains; D012333:RNA, Messenger / Q000032:analysis; D060888:Real-Time Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000032:analysis; D020539:Sequence Analysis, Protein; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D000070584:Tripartite Motif Proteins; D025801:Ubiquitin / Q000032:analysis / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D054875:Ubiquitination; D015027:Zebrafish / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D029961:Zebrafish Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
D012333:RNA, Messenger; D011994:Recombinant Proteins; D000070584:Tripartite Motif Proteins; D025801:Ubiquitin; D029961:Zebrafish Proteins; C573823:TRIM3a protein, zebrafish; D044767:Ubiquitin-Protein Ligases