Genome-wide loss-of-function analysis of deubiquitylating enzymes for zebrafish development
1 Laboratory of Developmental Signalling and Patterning, Genes and Development Division, Institute of Molecular and Cell Biology, Agency for Science, Technology and Research (A*STAR), 61 Biopolis Drive, 138673 Singapore
2 Experimental Therapeutics Centre, Agency for Science, Technology and Research (A*STAR), 31 Biopolis Way, 138669 Singapore
3 Bioinformatics Institute, Agency for Science, Technology and Research (A*STAR), 30 Biopolis Street, 138671 Singapore
4 Department of Biological Sciences, National University of Singapore, 14 Science Drive, 117543 Singapore
5 School of Computer Engineering, Nanyang Technological University, 50 Nanyang Drive, 637553 Singapore
6 Department of Biochemistry, National University of Singapore, 8 Medical Drive, 117597 Singapore
7 Current address: Laboratory of Physiology, Ocean Research Institute, The University of Tokyo, 1-15-1 Minamidai, Nakano-ku, Tokyo 164-8639, Japan
8 Current address: Division of Molecular and Genomic Medicine (DMGM), National Health Research Institutes (NHRI), No 35, Keyan Road, Zhunan Town, Miaoli County 35053, Taiwan
BMC Genomics 2009, 10:637 doi:10.1186/1471-2164-10-637Published: 30 December 2009
Deconjugation of ubiquitin and/or ubiquitin-like modified protein substrates is essential to modulate protein-protein interactions and, thus, signaling processes in cells. Although deubiquitylating (deubiquitinating) enzymes (DUBs) play a key role in this process, however, their function and regulation remain insufficiently understood. The "loss-of-function" phenotype studies can provide important information to elucidate the gene function, and zebrafish is an excellent model for this goal.
From an in silico genome-wide search, we found more than 90 putative DUBs encoded in the zebrafish genome belonging to six different subclasses. Out of them, 85 from five classical subclasses have been tested with morpholino (MO) knockdown experiments and 57 of them were found to be important in early development of zebrafish. These DUB morphants resulted in a complex and pleiotropic phenotype that, regardless of gene target, always affected the notochord. Based on the huC neuronal marker expression, we grouped them into five sets (groups I to V). Group I DUBs (otud7b, uchl3 and bap1) appear to be involved in the Notch signaling pathway based on the neuronal hyperplasia, while group IV DUBs (otud4, usp5, usp15 and usp25) play a critical role in dorsoventral patterning through the BMP pathway.
We have identified an exhaustive list of genes in the zebrafish genome belonging to the five established classes of DUBs. Additionally, we performed the corresponding MO knockdown experiments in zebrafish as well as functional studies for a subset of the predicted DUB genes. The screen results in this work will stimulate functional follow-up studies of potential DUB genes using the zebrafish model system.