Research in my laboratory is aimed at deciphering mechanisms of gene regulation as governed by chromatin structure. Chromatin boundary elements function to delimit the domains of active and inactive chromatin. A major question in eukaryotic gene regulation is how heterochromatin and euchromatin are physically and functionally separated by chromatin boundaries.
Boundary elements are divided into two classes: insulators are DNA sequences that block enhancer activation of a promoter, preventing inappropriate gene activation. Barriers are sequences that block the propagation of heterochromatin, thereby separating domains of active and inactive chromatin. In both cases, the function is to prevent regulatory regions of a given chromatin domain from inappropriately affecting another region.
Using the yeast Saccharomyces cerevisiae as a model system, we have
identified several DNA elements that function as heterochromatin barriers. One
is a unique tRNA gene (tDNA) located downstream from the silenced HMR
locus. Current genetic and biochemical studies center on analyzing mechanisms
of barrier function, and we have found that bromodomain proteins, which bind
to acetyllysine moieties on histones and other proteins, are important for barrier
integrity. We are currently focusing on the molecular characterization of the
yeast YTA7 gene, which codes for one such bromodomain containing protein.
We are also assessing the potential global role of RNA polymerase III promoters as chromatin boundaries. Recent evidence from our lab and others has shown that DNA bound TFIIIC, an RNA Polymerase III transcription factor, can be sufficient to create a chromatin boundary .We have found that other tDNAs and TFIIIC binding sites in the yeast genome can act as boundary elements, perhaps in the absence of the rest of the polymerase complex. This suggests a more general extra-transcriptional role for TFIIIC in sculpting genome architecture and function.
Selected Publications
RNA Polymerase III and Polymerase II Promoter Complexes are Heterochromatin Barriers in Saccharomyces cerevisiae.
David Donze and Rohinton T. Kamakaka
EMBO Journal, Vol. 20 No. 3, 520-531 (2001)
Breaking the Histone Code of Silence: The Propagation and Blocking of Heterochromatin.
David Donze
Current Organic Chemistry, Vol. 8, 211-221 (2004)
The S. cerevisiae TRT2 tRNAThr gene upstream of STE6 is a barrier to repression in MATa cells and exerts a potential tRNA position effect in MATa cells.
Tiffany A. Simms, Elsy C. Miller, Nicolas P. Buisson, Nithya Jambunathan, and David Donze
Nucleic Acids Research, Vol. 32, 5206-5213 (2004)
Modulation of yeast genome expression in response to defective RNA polymerase III-dependent transcription.
Christine Conesa, Roberta Ruotolo, Pascal Soularue, Tiffany A. Simms, David Donze, André Sentenac, and Giorgio Dieci
Mol. Cell. Biol. Vol. 25, 8631-8642 (2005)
Multiple bromodomain genes are involved in restricting the spread of heterochromatic silencing at the S. cerevisiae HMR-tRNA boundary.
Nithya Jambunathan, Adam W. Martinez, Elizabeth C. Robert, Nneamaka B. Agochukwu, Megan E. Ibos, Sandra L. Dugas, and David Donze.
Genetics, Vol. 171, 913-922 (2005)
Requirement of Nhp6 proteins for transcription of a subset of tDNAs and
heterochromatin barrier function in Saccharomyces cerevisiae.
Priscilla Braglia, Sandra L. Dugas, David Donze and Giorgio Dieci
Mol. Cell. Biol. Vol. 27, 1545-1557 (2007)