Zebrafish Ubc13 is required for Lys63-linked polyubiquitination and DNA damage tolerance.
Ubiquitination is an important post-translational protein modification that functions in diverse cellular processes of all eukaryotic organisms. Conventional Lys48-linked poly-ubiquitination leads to the degradation of specific proteins through 26S proteasomes, while Lys63-linked polyubiquitination appears to regulate protein activities in a non-proteolytic manner. To date, Ubc13 is the only known ubiquitin-conjugating enzyme capable of poly-ubiquitinating target proteins via Lys63-linked chains, and this activity absolutely requires a Ubc variant (Uev or Mms2) as a co-factor. However, Lys63-linked poly-ubiquitination and error-free DNA damage tolerance in zebrafish are yet to be defined. Here, we report molecular cloning and functional characterization of two zebrafish ubc13 genes, ubc13a and ubc13b. Analysis of their genomic structure, nucleotide and protein sequence indicates that the two genes are highly conserved during evolution and derived from whole genome duplication. Zebrafish Ubc13 proteins are able to physically interact with yeast or human Mms2 and both zebrafish ubc13 genes are able to functionally complement the yeast ubc13 null mutant for spontaneous mutagenesis and sensitivity to DNA damaging agents. In addition, upon DNA damage, the expression of zebrafish ubc13a and ubc13b is induced during embryogenesis and zebrafish Ubc13 is associated with nuclear chromatin. These results suggest the involvement of Lys63-linked poly-ubiquitylation in DNA damage response in zebrafish.
Li
Jie
J
Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan, Hubei, People's Republic of China.
Wen
Rui
R
Andersen
Parkeer
P
Liang
Yuping
Y
Li
Qing
Q
Xiao
Wei
W
Cui
Zongbin
Z
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D004249:DNA Damage; D017931:DNA Primers; D018076:DNA, Complementary; D005455:Fluorescent Antibody Technique; D005816:Genetic Complementation Test; D008239:Lysine / Q000378:metabolism*; D008969:Molecular Sequence Data; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D020798:Two-Hybrid System Techniques; D044763:Ubiquitin-Conjugating Enzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D054875:Ubiquitination; D015027:Zebrafish
D017931:DNA Primers; D018076:DNA, Complementary; D044763:Ubiquitin-Conjugating Enzymes; D008239:Lysine