Related information:
|
Contact information Recent publications Curriculum vitae Laboratory of Molecular Biology |
Biography: Dr. Trun also holds an Adjunct Assistant Professorship at the University of Maryland at College Park. She received her Ph.D. from Princeton University and was a Jane Coffin Childs Postdoctoral Fellow at NCI.Research: Many cellular processes require spatial and temporal coordination of a complex series of events. One of the most intricate of these is maintenance of daughter chromosomes. Using E. coli and Schizzosaccharomyces pombe as a model systems we have been focusing on condensation of the DNA into a functional state. Both projects were initiated by isolating cells that are resistant to the diploidizing agent, camphor. By isolating high copy number plasmids that confer camphor resistance, we have identified a three gene system in E. coli that functions to condense the chromosome and several potential plasmids that affect chromatin structure in S. pombe.To begin to understand the mechanism of action of camphor and identify its target(s), we isolated high copy number plasmids that confer resistance in E. coli. One fragment of chromosomal DNA (from 14.2 min.), when present on pBR322, gives this phenotype. Unlike chromosomal camphor resistant mutants, the plasmid does not result in diploidy. Deletion analysis and sequencing indicated that three genes are responsible for the phenotype, cspE, a cold shock gene homolog and two other ORF's (crcA and crcB). crcA is most likely the regulator of the system and crcB is predicted to be an integral inner membrane protein. From their natural promoters, all three genes are required in the correct ratios to confer camphor resistance. However, if cspE is greatly overexpressed from a heterologous promoter then it can confer all of the known phenotypes of the three genes.All of our data on crcA, cspE and crcB led us to suspect that the genes are affecting chromosome condensation. In a direct test of this, we have demonstrated that, both in vivo and in vitro, camphor does indeed cause unfolding of the chromosome and overproduction of the three genes results in more tightly condensed nucleoids. The original work of Burgi and Worcell on the nucleoid demonstrated that mRNA is one of the major components holding the nucleoid together. The recent discovery that CspE binds to the 5' end of mRNA in an active transcription complex may suggest that this condensing mechanism senses actively transcribed genes and may condense using the mRNA. Alternatively, because CspE also bind dsDNA, a bending mechanism similar to that described below for Sac7d may explain how CspE functions.E. coli contains seven homologs of cspE, three are cold-shock induced, while the other that have been tested, including cspE, are not. The unifying theme of these proteins are that they bind some form of nucleic acids (ssDNA, dsDNA, RNA). A homolog from Sulfolobus, Sac7d, has recently been crystallized with DNA. This protein binds the minor groove of dsDNA and introduces a large bend. These small (8 kDa) proteins may represent a novel chromosome condensation mechanism that relies on randomly bending the DNA.In a new project that we have recently undertaken, we are extending our findings with camphor to the fission yeast, Schizzosaccharomyces pombe. In S. pombe, the two unexpressed mating type cassettes are kept silenced by their chromatin structure. We have isolated a series of plasmids that confer camphor resistance on S. pombe and checked their affects on the silent mating type loci (mat2 and mat3). Approximately 20% of our camphor resistant plasmids activate the silent loci, not only increasing their gene expression but also affecting their stability. These plasmids and the general chromosome disruption phenotype of camphor may allow us to uncover novel aspects of chromatin structure and genome instability in this genetically tractable eukaryote.Recent Publications: |
Contact Information: Laboratory of Molecular Biology,NCI, NIHBuilding 37, Room 2D2137 CONVENT DR MSC 4255BETHESDA MD 20892-4255Phone: 301-496-3874Fax:301-496-2212Email: trun@helix.nih.gov
|
