A genetic and molecular analysis of histone deacetylase one in Drosophila melanogaster : specific missense mutations suppress position effect variegation

• Main Author: Mottus, Randall C.
• Format: Others
• Language: English
• Published: 2009
• Online Access: http://hdl.handle.net/2429/14807

Essentially all higher organisms are made up of two or more types of tissues. The specific identity of those tissues is dependent on the genes that are expressed within the cells of the particular tissue type. The correct set of genes must be expressed and genes, that are not part of the set specific to that tissue, must be kept silenced. In addition, in most cells, the decision whether a gene will be active or not is made early in development and therefore must be passed on to daughter cells. The focus of this thesis is an investigation into the mechanism or mechanisms employed by eukaryotes to silence genes and to maintain that silenced state throughout development. The model system our laboratory has been using to investigate silencing is position effect variegation (PEV) in D. melanogaster. In PEV a gene is silenced in a certain proportion of the cells of a tissue in which it is normally expressed due to its proximity to an heterochromatic breakpoint. The decision whether a gene will be active or inactive is made early in development and that decision is passed on to daughter cells with reasonable fidelity. Thus PEV mimics normal development in many ways. This has led our lab, and several others, to try to dissect the mechanisms underlying PEV with the hope they will shed some light on the more general silencing mechanisms that occur during normal development. In Chapter 2 of this thesis I describe the cloning and characterization of a gene identified in a screen for dominant suppressors of the variegation associated with PEV [Su(var)s]. The gene encodes HDAC1 , an histone deacetylase homologous to HDAC1 from mammals and Rpd3 from S. cerevisiae. Specific mis-sense mutations in HDAC1 cause strong dominant suppression of PEV while null or hypomorphic mutations have no effect on the variegating phenotype. I present a model proposing that the mis-sense mutations are acting as anti-morphic mutations that "poison" the deacetylase complex. The level of variegation of a gene subject to PEV is very sensitive to a wide variety of factors, some, which may be acting directly and some, which may be acting indirectly. HDAC1 localizes to a large number of sites on the polytene chromosomes of D. melanogaster (Pile and Wasserman, 2000) and therefore appears to regulate a large number of genes. Thus it is a possibility that the Su(var) mutations in HDAC1 are affecting PEV indirectly. In Chapter 3 I present data from chromatin immuno-precipitation experiments (X-ChIP) that provides compelling evidence that HDAC1 is acting directly on the euchromatic region subject to silencing in PEV. I propose a model linking the histone deacetylase activity of HDAC1 to the function of other proteins known to be involved in the silencing associated with PEV. === Science, Faculty of === Zoology, Department of === Graduate