A gene-trap strategy identifies quiescence-induced genes in synchronized myoblasts
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Cellular quiescence is characterized not only by reduced mitotic and metabolic activity but also by altered gene expression. Growing evidence suggests that quiescence is not merely a basal state but is regulated by active mechanisms. To understand the molecular programme that governs reversible cell cycle exit, we focused on quiescence-related gene expression in a culture model of myogenic cell arrest and activation. Here we report the identification of quiescence-induced genes using a gene-trap strategy. Using a retroviral vector, we generated a library of gene traps in C2C12 myoblasts that were screened for arrest-induced insertions by live cell sorting (FACS-gal). Several independent genetrap lines revealed arrest-dependent induction of βgal activity, confirming the efficacy of the FACS screen. The locus of integration was identified in 15 lines. In three lines, insertion occurred in genes previously implicated in the control of quiescence, i.e. EMSY — a BRCA2-interactin proteinm, p8/com1 — a p300HAT-binding protein and MLL5 — a SET domain protein. Our results demonstrate that expression of chromatin modulatory genes is induced in G0, providing support to the notion that this reversibly arrested state is actively regulated.
KeywordsGene-trap MLL5 myoblast p8 quiescence
colony forming units
fluorescent activated cell sorting
long terminal repeat
mixed lineage leukaemia gene 5
multiplicity of infection
myogenic regulatory factor
phenyl ethyl thiogalactoside
plant homoeo domain
polymerase chain reaction
conserved in Su(var), Enhancer of Zeste, Trithorax
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