Females have two X chromosomes and males have one, but they all have two copies of all other chromosomes. Gene dosage, the number of copies of each gene, needs to be balanced with all other genes in the genome. Recent studies suggested that gene dosage balance is achieved in two steps. First the X chromosome is upregulated two-fold, so as to balance the single X in males to the other chromosomes which are present in two copies. However, upregulation of the two X chromosomes in females leads to excessive X chromosome expression, and one of these X chromosomes is then silenced. This process is called dosage compensation, because it compensates for imbalances caused by different numbers of X chromosomes. This project will investigate the mechanism of dosage compensation in a genetic model organism, the roundworm Caenorhabditis elegans, where animals with one X chromosome are males, and animals with two X chromosomes are hermaphrodites. The genes which function in worm dosage compensation are known, but their molecular mechanism of action needs to be elucidated. The DNA in our chromosomes is packaged into a tight structure around small proteins called histones. Turning genes on or off frequently involves addition or removal of small molecules, such as methyl or acetyl groups, from histone proteins. This project will investigate the role of the addition of an acetyl group to a specific residue, lysine 16 of histone H4. Preliminary evidence indicate that the addition of this acetyl group to histone H4 plays a role in the upregulation of the X chromosome in male worms, and removal of this acetyl group plays a role in repression of the two X chromosomes in hermaphrodite worms. The intellectual merit of this project is that it is the first detailed analysis of the role of histone modifications in worm dosage compensation and the first study of any kind addressing the mechanism of X-upregulation in worms. The results will also shed light on the general mechanism of how histone modifications can affect chromosome architecture and gene expression.
Broader Impacts: The project will have a broad impact on education on several levels. First, undergraduate students will be active participants in performing the experiments and will be exposed to real life research not in a course setting. Second, a larger group of undergraduates will be exposed to the research in an upper level seminar course focusing on chromosome structure and function. To bring research into the classroom, raw data generated during the project will be analyzed by students enrolled in the course, so students can witness how data turns into publication quality images. Finally, training a postdoctoral fellow for a career which combines research and education is a central part of the project. The postdoctoral researcher will be involved in the research project during the first two years, and will be participating in discussion groups and journal clubs focused on teaching. In the final year of the project, under the mentorship of the principle investigator, the postdoctoral fellow will explore part-time teaching opportunities at the university.
Our genetic material, DNA, together with histones and other proteins, is packaged into chromatin. Looser packaging of chromatin is associated with active genomic regions, and tighter packaging with silenced regions. This project investigated how a particular modification of histones (acetylation) influences chromatin packaging and gene activity on the X chromosomes in the nematode Caenorhabditis elegans. Acetylation levels were found to be low the X chromosomes of XX animals (hermaphrodites) and high in XO animals (males). These differences in acetylation contribute to overall higher levels of gene expression on the single male X, and lower level of gene expression on each of the two hermaphrodite Xs, thus equalizing X-linked gene expression in the process called dosage compensation. Results showed that this modification of the chromatin fiber also affects higher order packaging of the genome, leading to compaction of the chromosome when depleted of acetylation (in XX animals) and enlargement of the volume of the X chromosome when enriched for acetylation (in XO animals). The intellectual merit of this project is that it was the first detailed analysis of the role of histone modifications in nematode dosage compensation and their affect on higher order packaging of the genome. The results mirror findings in other organisms, such as flies and humans, implying conservation of mechanisms. The project had a broader impact on education on multiple levels. Two postdoctoral fellows received training for a career that combines research and teaching. As well as being active participants in research, under the supervision of the principal investigator they also taught a mini-module of an upper level undergraduate course on chromosome structure and function. The topic of this project was included in the course. Although enrolled students did not participate in performing experiments, they witnessed the process from experimental design through raw data to publication. A small number of undergraduates were able to become active participants by joining the research group and receiving hands on training in research. Undergraduate students, graduate students and postdoctoral fellows all had the opportunity to attend international and national conferences as part of their professional development.
