Principal Investigator Gerald Fink
Many of the cell surface genes that show repeat-size variation also vary epigenetically. Our previous work had shown that FLO11, one of the 22 cell surface genes with repeat-size variation, also varies epigenetically, so that some cells express FLO11 ("on" ) and others did not ("off").
Further studies have made promoter fusions to GFP and URA3 of all 78 genes in the yeast genome that are annotated in SGD as producing a "cell wall localized protein". These are fusions to the initiating ATG in each gene and have only the coding region of the reporter. As controls, we have made promoter fusions to many other housekeeping genes randomly chosen to act as controls. The fusions are then tested to determine whether they variegate by three criteria:
(1) Promoter fusions to URA3 variegate(2) Promoter fusions to GFP--Clonal analysis by fluorescence microscopy and FACS documents: Heritable change in phenotype transmitted from generation to generation despite identical genotype(3) Abolition of variegation by trans-acting chromatin factors e.g. histone deacetylases. Remarkably, unlike the controls (genes encoding non-cell wall proteins) many of the genes with internal repeats that we had shown have genetic variation in repeat length, also show epigenetic switching by these criteria.
The variegation of these cell surface proteins is under the control of chromatin binding proteins. Previous work showed that one of these, FLO11, is under the control of a number of trans-acting chromatin binding factors. Knockouts of these factors prevents epigenetic switching and locks the cells into an "on" or "off" expression state.