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Browsing by Subject "ATRX"

Now showing 1 - 2 of 2
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    Novel roles for the Fanconi Anemia pathway protein FANCD2 in the recovery of stalled replication forks
    (2017-06) Raghunandan, Maya
    Fanconi Anemia (FA) is an inherited cancer predisposition syndrome that is characterized by a cellular hypersensitivity to DNA interstrand crosslinks (ICLs). To repair these DNA lesions, the 21 known FA proteins are thought to act in a linear hierarchy: Following ICL detection, an upstream FA core complex activates two central FA pathway members, FANCD2 and FANCI, via monoubiquitination. Both activated proteins then bind the ICL and recruit downstream FA proteins that repair the ICLs. Importantly, we previously found that FANCD2 has an additional independent role during the cellular replication stress response: it promotes the homologous recombination (HR) dependent restart of hydroxyurea (HU) stalled replication forks in concert with other HR DNA repair proteins such as the BLM helicase. In this work, we show that FANCD2 promotes replication fork restart in concert with downstream FA pathway proteins but independently of the upstream FA core complex and thus, independently of FANCD2 monoubiquitination. To further our understanding of how FANCD2 promotes replication fork recovery, we performed a search for S-phase specific FANCD2 interactors and we identified a novel FANCD2 interacting protein, Alpha Thalassemia Retardation X-linked factor (ATRX). ATRX is a subunit of the ATRX/DAXX histone H3 chaperone complex that plays several key roles in regulating chromatin structure and was recently identified as a replication fork recovery factor. Our new findings demonstrate that ATRX forms a constitutive complex with FANCD2 and promotes FANCD2 protein stability. Moreover, while ATRX is dispensable for DNA ICL repair, it works in concert with FANCD2 to promote HU resistance and the restart of HU-stalled replication forks. Remarkably, the HR-dependent replication fork restart requires the histone H3 chaperone activity of both the ATRX/DAXX complex and FANCD2 indicating that histone exchange at stalled replication forks is a crucial step in fork restart. Altogether, our results support a novel non-linear FA pathway model where individual protein members fulfill distinct cellular roles to support genomic stability. We propose that FANCD2- and possibly other FA pathway proteins- is involved in the deposition of histone H3 variants in the vicinity of HU- stalled replication forks to mediate fork recovery.
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    The role of DNA repair & regulatory proteins in the maintenance of human telomeres and their control of cellular immortalization
    (2017-04) Harvey, Adam
    Telomeres are the nucleoprotein structures that protect the ends of linear chromosomes from recognition as a double-stranded DNA break (DSB). In the absence of proper telomere function, the ends of a chromosome fuse together, creating di-centromeric chromosomes, which can no longer properly segregate at mitosis. Thus, proper telomere maintenance is absolutely essential for all eukaryotic life. Unfortunately, maintaining telomeres at a size that is protective is problematic. For example, as a consequence of “the end-replication problem,” telomeres shorten incrementally during every cell cycle. These short telomeres can, in turn, function to regulate the lifespan of any given cell. Perhaps not surprisingly, therefore, humans have evolved a vast array of genes to enable telomere stability, in order to counteract any premature ageing or cell death. In order to ensure that offspring may begin their life with a default telomere length that is sufficient for stability during the organism’s lifespan, stem cells must not be subjected to overall telomere shortening. Thus, all telomere shortening that a stem cell occurs during its eternal proliferation must be correspondingly compensated for by a lengthening event. This telomere elongation mechanism in essence confers cellular immortality. The most well-characterized of these cellular immortality pathways is controlled by the enzyme telomerase, which precisely elongates telomeres in a stochastic way to maintain a telomere length equilibrium. Unfortunately, this functional, essential pathway can also be conscripted to perform pathological reactions. In human cancer, all malignant growths must enable cellular immortalization to allow for their characteristic uncontrolled proliferation. In most cases this is achieved simply by the reactivation of telomerase. Interestingly, 5 to 15% of all human cancers are telomerase negative. These cancers can be described as ALT cancers, as the Alternative Lengthening of Telomeres pathway enables their immortality. ALT, which is specific to cancer, achieves telomere elongation by aberrant recombination between telomeres. My research has found that DNA repair proteins, such as PARP1, (poly ADP ribose polymerase 1) are critical for both the maintenance of the genome and specifically for proper telomere maintenance. Furthermore, my research has demonstrated that the mutation of a single gene, ATRX, (alpha thalassemia mental retardation on the X chromosome) is an active repressor of ALT immortalization. In summary, I have contributed to the understanding of human telomere length maintenance and these studies have implications for human aging and the genesis of cancer.