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Development of an Allele-Aware Method to Study the Nuclear Organization of the FSHD Locus

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dc.contributor.advisor Trask, Barbara en_US Siebenthall, Kyle en_US 2012-09-13T17:27:31Z 2013-09-14T11:05:27Z 2012-09-13 2012 en_US
dc.identifier.other Siebenthall_washington_0250E_10581.pdf en_US
dc.description Thesis (Ph.D.)--University of Washington, 2012 en_US
dc.description.abstract The human genome carries out its functions, including the establishment and maintenance of tissue-specific gene expression programs, within the confines of the cell nucleus. The genome is not randomly organized within this space: its three-dimensional arrangement plays a role in executing its functions. One hallmark of nuclear organization is the compartmentalization of active and inactive genomic regions, with little contact between regions of different states. In many different cell types, the 4q subtelomere is located at the nuclear periphery, a repressive region adjacent to the nuclear lamina. Deletions in a tandem repeat array at this locus lead to the muscular dystrophy FSHD through an epigenetic mechanism that results in inappropriate expression of DUX4, a transcription factor encoded in each repeat unit that is normally active in the germline. DUX4 is toxic to muscle cells, yet its high abundance in only a small fraction of muscle cells from patients hints that there might be multiple determinants of its expression. My thesis work addressed the hypothesis that nuclear organization is one of these determinants. Since the FSHD locus is duplicated in the genome, I developed an allele-aware method (4C-seq) that I used to interrogate the "nuclear neighborhood" of specific copies of the locus in primary muscle cells from control individuals and individuals with FSHD. 4C-seq uses a polymorphic "bait" to capture regions of the genome ("prey") that are in close physical proximity to the bait in a population of nuclei. By characterizing prey fragments captured by a bait within the FSHD locus, I found that the locus normally contacts lamin-associated regions with low transcriptional output within its own chromosome territory, and associates with putative insulator sequences (bound by the CTCF protein) and centromeres of other chromosomes. In FSHD cells, I found that the locus carrying a pathogenic deletion contacts regions within its own chromosome that have lower lamin-association and higher transcriptional output than the regions contacted in control cells. My results suggest that an altered nuclear neighborhood might play a role in the mis-expression of DUX4 in FSHD. en_US
dc.format.mimetype application/pdf en_US
dc.language.iso en_US en_US
dc.rights Copyright is held by the individual authors. en_US
dc.subject 4C; FSHD; Nuclear organization en_US
dc.subject.other Biology en_US
dc.subject.other Genetics en_US
dc.subject.other Genetics en_US
dc.title Development of an Allele-Aware Method to Study the Nuclear Organization of the FSHD Locus en_US
dc.type Thesis en_US
dc.embargo.terms Restrict to UW for 1 year -- then make Open Access en_US

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