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Research

The Weiler lab studies chromosome and chromatin structure. We use the fruit fly, Drosophila melanogaster, as a model system to investigate factors important for differences in chromatin structure that regulate genome activity i.e. epigenetic regulatory mechanisms.  We are especially interested in  two chromatin proteins of unknown function, encoded by the E(var)3-9 and D1 genes.

E(var)3-9Identification of PEV Modifiers
    The E(var)3-9 gene was originally identified by dominant mutations that resulted in enhancement of position effect variegation (PEV) of In(1)wm4 [Dorn et al. 1993].  PEV is observed as a result of chromosome rearrangements with breakpoints in the heterochromatin and euchromatin.  Genes located near the novel breakpoints exhibit cell-to-cell differences in gene activity due to the altered chromatin environment.  The classic example of this phenomenon is the wm4 inversion.  As illustrated in the figure at right, dominant second-site modifiers of In(1)wm4 variegated expression were isolated in several large-scale genetic screens as suppressors (Su(var) genes) or enhancers (E(var) genes).  The Su(var) genes were proposed to encode proteins that promote heterochromatin formation as structural proteins or enzymes that modify chromatin.  In contrast, the E(var) genes were proposed to antagonize heterochromatin formation and/or promote euchromatin formation.
    As an enhancer of variegation, E(var)3-9 is thought to alter the balance between heterochromatin and euchromatin in favor of euchromatin formation. In support of this hypothesis, our laboratory showed that E(var)3-9 mutations had opposite effects on the variegation of euchromatic and heterochromatic genes [Weiler 2007].  While mutation of E(var)3-9 enhanced wm4 variegation, it suppressed the variegation of the heterochromatic lt gene. We cloned the E(var)3-9 gene.  It encodes a zinc finger protein, consistent with a potential role as a DNA-binding protein [Weiler 2007].
    Mutation of E(var)3-9 causes maternal effect lethality.  E(var)3-9 mutant females lay eggs that fail to hatch. This phenotype reveals that the E(var)3-9 gene product is required for early embryonic development [Weiler 2007].

D1
    The D1 protein is a nonhistone chromosomal protein that binds to the minor groove of AT-DNA tracts in vitro and associates with the AATAT and 359 bp repeat satellite sequences, in vivo [Levinger and Varshavsky, 1982a and 1982b].  Immunostaining of mitotic chromosomes from the drosophila embryo or larval neuroblasts confirms a heterochromatic localization for D1 [Renner et al. 2000; Aulner et al. 2002].  However, the D1 protein was also observed to associate with euchromatin by immunostaining of the larval salivary gland polytene chromosomes [Alfageme et al. 1976; Rodriguez Alfageme et al. 1980; Smith and Weiler 2010].   Although identified over 30 years ago, the cellular functions of the D1 protein remain obscure.
    Our laboratory is taking a genetic approach to investigate the function(s) of D1.  We have isolated deletion alleles of the D1 gene and are analyzing the D1-null phenotype [Weiler and Chatterjee 2009].  Surprisingly, we found that the D1 gene is not essential for viability or fertility.  D1 mutant alleles are also not suppressors of PEV, as might have been expected given the predominantly heterochromatic localization of the D1 protein.  We have also used gain-of-function studies to examine D1 function [Smith and Weiler 2010].  We found that ubiquitous D1 overexpresion is lethal.  By overexpressing D1 in specific tissues, a variety of phenotypes is elicited that reveal a disruption of normal differentiation and development.  Strikingly, overexpression of D1 in the larval salivary glands, which have visible interphase (polytene) chromosomes, leads to abnormal ectopic associations between and within chromosomes.  Another line of experimentation underway to investigate D1 function is the yeast two-hybrid approach, as a means to identify proteins with which the D1 protein interacts.

Publications

Smith, M.B. and K.S. Weiler (2010) Drosophila D1 overexpression induces ectopic pairing of polytene chromosomes and is deleterious to development. Chromosoma 119: 287-309

Weiler, K.S. and S. Chatterjee (2009) The multi-AT hook chromosomal protein of Drosophila melanogaster, D1, is dispensable for viability. Genetics 182: 145-159

Weiler, K.S. (2007) E(var)3-9 of Drosophila melanogaster encodes a zinc finger protein. Genetics 177:167-178.

Weiler, K.S. and B.T. Wakimoto (2002) Suppression of heterochromatic gene variegation can be used to distinguish and characterize E(var) genes potentially important for chromosome structure in Drosophila melanogaster. Molecular Genetics & Genomics 266: 922-932.

Weiler, K.S. and B.T. Wakimoto (1998)  Chromosome rearrangements induce both variegated and reduced, uniform expression of heterochromatic genes in a development-specific manner.  Genetics 149:1451-1464.

Weiler, K.S. and B.T. Wakimoto (1995)  Heterochromatin and gene expression in Drosophila.  Annu. Rev. Genetics 29:577-605.

Weiler, K.S., L. Szeto and J.R. Broach (1995)  Mutations affecting donor preference during mating type interconversion in Saccharomyces cerevisiae.  Genetics 139:1495-1510.

Weiler, K.S. and J.R. Broach (1992)  Donor locus selection during Saccharomyces cerevisiae mating type interconversion responds to distant regulatory signals.  Genetics 132:929-942.


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