Interplay of chromatin remodeling, transcriptional regulation, and nuclear organization

Abstract

Transcription is regulated on different levels to ensure that genes are expressed at the correct time and in the amounts required. At the chromatin level, DNA is wound onto histone proteins, forming nucleosomes that influence accessibility of DNA elements. Modifications on those histones and interactions with other chromatin proteins can either encourage or inhibit recruitment of the transcription machinery. Genomic regions of similar character form chromatin domains, organizing the genome based on their transcription states. Within the nucleus, both individual loci and entire chromosomes assume non-random positions, based on their transcription levels and interactions with nuclear landmarks. This thesis examines the effects of the Fun30 chromatin remodeling enzymes on transcription regulation and nuclear organization, both on the local chromatin level as well as on a genome-wide scale.

Using the fission yeast Schizosaccharomyces pombe as a model organism, we mapped the interactions between the genome and two inner nuclear membrane proteins, Ima1 and Man1. We observed a preference for lowly expressed genes to associate with the nuclear envelope, similar to what had been observed in mammalian and fruit fly cells. When comparing Ima1 and Man1 binding patterns, we found both common and separate target sites, suggesting a role for inner nuclear membrane proteins in organizing the fission yeast genome.

Following up on these results, we went on to examine subtelomeric chromatin domains, which are regulated through the Fun30 remodeler Fft3. These domains contain repressed genes, whose transcription levels increase in cells carrying an fft3Δ deletion. While the subtelomeres associate with the nuclear envelope through Man1 in wild-type cells, this interaction is lost in fft3Δ cells. In these cells, we also observed changes in nucleosome occupancy at the subtelomeric borders. Interestingly, a strain carrying a catalytically inactive version of the Fft3 remodeler showed the same behavior as the deletion strain, with upregulation of subtelomeric genes and loss of Man1 interactions. Together, these results point to an active role of Fft3 in regulating subtelomeric chromatin, transcription, and nuclear periphery interactions.

In addition to their role at subtelomeres, Fun30 remodelers also control transcription in other parts of the genome. When we examined a strain lacking Fft2, a paralog of Fft3, we found increased transcription of the fission yeast Tf2 retrotransposons. This increase is accompanied by a shift in transcription start site (TSS) further upstream and is especially pronounced when both fft2 and fft3 are deleted. By mapping nucleosome positioning, we were able to establish that Fft2 and Fft3 collaborate in stabilizing a nucleosome over the upstream TSS, resulting in transcription initiation further downstream and production of an mRNA incapable of transposition. Expression of both remodelers is downregulated in stress conditions, allowing for production of the longer transcript under these circumstances. We propose that the shift in TSS choice allows for bursts of transposition in cells under environmental stress. This can enable cells to adapt to changed conditions through favorable insertion events altering expression of nearby genes.