Download functional screening of human genes in zebrafish animal model

FUNCTIONAL SCREENING OF HUMAN GENES IN ZEBRAFISH
ANIMAL MODEL
Cheol-Hee Kim
Department of Biology and GRAST, Chungnam National University, Daejeon,
Korea
Tel: +82-42-821-5494
E-mail: [email protected]
Abstract
The zebrafish (Danio rerio) is a vertebrate model system suitable for fishing
novel genes in post-genome era. Two neural mutants isolated from the ENUbased chemical mutagenesis, were characterized and the point mutations were
identified by the positional cloning method. The first mutant is headless: Wnt
antagonists secreted by the organizer have been identified as head inducers. The
severe head defects in headless are due to a mutation in Tcf-3, a component of
the Wnt pathway. The second is mind bomb: Reduced lateral inhibition in mutant
permits too many neural progenitors to differentiate as neurons. Mind bomb
encodes a novel gene in the Notch signaling pathway. In addition to chemical
mutagenesis, we developed a novel transgene reporter system and tried to apply
its use for the insertional mutagenesis. The first mutant "ondal" from this new
screening approach, was characterized and mapped to a gene involved in axonal
myelination and human schizophrenia. The other mutants, moguri, samdori, and
hayan-4, are being mapped and characterized.
As a separate approach, we are challenging a genome-wide screen against
the human UniGene collections. Currently, more than 15,000 human full-length
cDNAs are available from the Korean UniGene Information (KUGI). After in silico
screening of functionally unknown genes through database search, we performed
overexpression experiments by microinjecting the single human genes into the
zebrafish embryos. So far, we performed microinjections more than 2,600 human
full-length cDNAs and identified 50 functionally active new genes. Zebrafish
embryos display variable phenotypes from developmental defects to neuronal cell
death in injected embryos. Many of these genes were involved in not only
developmental process but also diseases, such as cancer. After these phenotypebased screening, homologous zebrafish genes were cloned and further functional
studies were performed by expression analysis, overexpression and antisense
knockdown approaches. From these functionally active genes, we found that the
human clone #462, now named as ottogi, plays a role in vertebrate head
formation during early development. Overexpression of ottogi caused a big head
phenotype, indicating its possible role in the Wnt signaling. Ottogi physically
interacted with the Wnt receptor Frizzled 8 within the endoplasmic reticulum.
These findings suggest that Ottogi is a novel component negatively regulating
the Wnt signal pathway.
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Jung SH et al. Developmental dynamics 2009 in press.
Choi TY et al. J. Investigative Dermatology. 2009 in press.
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Kim CH et al. Nature 2000;407:913
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