Alexander J Garvin

• Doctor of Philosophy
• Lecturer at University of Leeds

RNF168 is an E3 ubiquitin ligase critical to the mammalian DNA double-strand break repair response. The protein is recruited to and amplifies ubiquitin signals at damaged chromatin and, if not properly regulated, can drive an uncontrolled ubiquitin cascade potentially harmful to repair outcomes. Several indirect mechanisms restrict RNF168 positive...

The amplitudes of small-modifier protein signaling through ubiquitin and the small ubiquitin-like modifiers, SUMO1-3, are critical to the correct phasing of DNA repair protein accumulation, activity, and clearance and for the completion of mammalian DNA double-strand-break (DSB) repair. However, how SUMO-conjugate signaling in the response is delin...

Mammalian DNA replication relies on various DNA helicase and nuclease activities to ensure accurate genetic duplication, but how different helicase and nuclease activities are properly directed remains unclear. Here, we identify the ubiquitin-specific protease, USP50, as a chromatin-associated protein required to promote ongoing replication, fork r...

The preservation of genome integrity requires specialised DNA damage repair (DDR) signalling pathways to respond to each type of DNA damage. A key feature of DDR is the integration of numerous post-translational modification signals with DNA repair factors. These modifications influence DDR factor recruitment to damaged DNA, activity, protein-prote...

Mammalian DNA replication employs several RecQ DNA helicases to orchestrate the faithful duplication of genetic information. Helicase function is often coupled to the activity of specific nucleases, but how helicase and nuclease activities are co-directed is unclear. Here we identify the inactive ubiquitin-specific protease, USP50, as a ubiquitin-b...

A synthetic lethal relationship exists between disruption of polymerase theta (Polθ), and loss of either 53BP1 or homologous recombination (HR) proteins, including BRCA1; however, the mechanistic basis of these observations are unclear. Here we reveal two distinct mechanisms of Polθ synthetic lethality, identifying dual influences of 1) whether Pol...

Monoclonal antibodies (MAb) to members of the Small Ubiquitin-like modifier (SUMO) family are essential tools in the study of cellular SUMOylation. However, many anti-SUMO MAbs are poorly validated, and antibody matching to detection format is without an evidence base. Here we test the specificity and sensitivity of twenty-four anti-SUMO MAbs towar...

The amplitudes of small-modifier protein signalling through ubiquitin and the Small Ubiquitin-like Modifiers, SUMO1-3, are critical to the correct phasing of DNA repair protein accumulation, activity, and clearance, and for the completion of mammalian DNA double-strand break (DSB) repair. However, how SUMO-conjugate signalling in the response is de...

The E3 ubiquitin ligase RNF168 is a rate-limiting component of DNA double-strand break signalling that acts to amplify histone ubiquitylation. The confining of RNF168 chromatin spreading around DNA break sites has been related to constraining its expression levels and to the removal of ubiquitin-conjugates to which it binds. Here we identify a new...

Introductory paragraph Cells lacking several DNA repair proteins, including those promoting homologous recombination (HR), are sensitive to polymerase theta (Polθ) repression 1–4 . Polθ drives theta-mediated end joining (TMEJ) and suppresses HR but what mediates its synthetic lethal relationships is unclear. Here we examine murine Brca1 C61G/C61G 5...

Monoclonal antibodies (MAb) to members of the Small Ubiquitin-like modifier (SUMO) family are essential tools in the study of cellular SUMOylation. However, many reagents are poorly validated, and the choice of which antibody to use for which detection format is without an evidence base. Here we test twenty-four anti-SUMO monoclonal antibodies for...

Mammalian cells possess three conjugatable SUMO variants: SUMO1 and the largely indistinguishable SUMO2 and SUMO3 (designated SUMO2/3). Some SUMOylated substrates are modified by both SUMO1 and SUMO2/3, while others show biased modifications towards SUMO1 or SUMO2/3. How preferential SUMO protein conjugation is coordinated is poorly understood. Her...

The cellular response to genotoxic DNA double strand breaks (DSBs) uses a multitude of post-translational modifications to localise, modulate and ultimately clear DNA repair factors in a timely and accurate manner. Ubiquitination is well established as vital to the DSB response, with a carefully co-ordinated pathway of histone ubiquitination events...

The integrity of genomes is constantly threatened by problems encountered by the replication fork. BRCA1, BRCA2 and a subset of Fanconi anaemia proteins protect stalled replication forks from degradation by nucleases, through pathways that involve RAD51. The contribution and regulation of BRCA1 in replication fork protection, and how this role rela...

A. Alignment of the IRF1 GSK3 consensus site with a subset of other known GSK3 substrates, all sequences are human and the location indicated in parenthesis. Grey highlights indicate phosphorylated residues B. Schematic representation of IRF1 functional domains, indicating secondary structure within the DNA binding domain. The TAD (Transcriptional...

IRF1 (Interferon Regulatory Factor-1) is the prototype of the IRF family of DNA binding transcription factors. IRF1 protein expression is regulated by transient up-regulation in response to external stimuli followed by rapid degradation via the ubiquitin-proteasome system. Here we report that DNA bound IRF1 turnover is promoted by GSK3 (Glycogen Sy...

SUMOylation (small ubiquitin-like modifier) in the DNA double-strand break (DSB) response regulates recruitment, activity, and clearance of repair factors. However, our understanding of a role for deSUMOylation in this process is limited. Here we identify different mechanistic roles for deSUMOylation in homologous recombination (HR) and nonhomologo...

IRF1 (Interferon Regulatory Factor-1) is the prototype of the IRF family of DNA binding transcription factors. IRF1 protein expression is regulated by transient up-regulation in response to external stimuli followed by rapid degradation via the ubiquitin-proteasome system. Here we report that DNA bound IRF1 turnover is promoted by GSK3β (Glycogen S...

SUMOylation in the DNA double-strand break (DSB) response regulates recruitment, activity and clearance of repair factors. However, our understanding of a role for deSUMOylation in this process is limited. Here we identify different mechanistic roles for deSUMOylation in homologous recombination (HR) and non-homologous enjoining (NHEJ) through the...

The response to a DNA double-stranded break in mammalian cells is a process of sensing and signalling the lesion. It results in halting the cell cycle and local transcription and in the mediation of the DNA repair process itself. The response is launched through a series of post-translational modification signalling events coordinated by phosphoryl...

The opposing activities of 53BP1 and BRCA1 influence pathway choice in DNA double-strand-break repair. How BRCA1 counteracts the inhibitory effect of 53BP1 on DNA resection and homologous recombination is unknown. Here we identify the site of BRCA1-BARD1 required for priming ubiquitin transfer from E2∼ubiquitin and demonstrate that BRCA1-BARD1's ub...

SUMO conjugation is known to occur in response to double-stranded DNA breaks in mammalian cells, but whether SUMO deconjugation has a role remains unclear. Here, we show that the SUMO/Sentrin/Smt3-specific peptidase, SENP7, interacts with the chromatin repressive KRAB-associated protein 1 (KAP1) through heterochromatin protein 1 alpha (HP1α). SENP7...

The regulation of Ubiquitin (Ub) conjugates generated by the complex network of proteins that promote the mammalian DNA double-strand break (DSB) response is not fully understood. We show here that the Ub protease POH1/rpn11/PSMD14 resident in the 19S proteasome regulatory particle is required for processing poly-Ub formed in the DSB response. Prot...

IRF-1 (Interferon Regulatory Factor 1) is a transcription factor first identified as a regulator of Interferon expression. Two decades after its discovery, IRF-1 has been shown to be involved in numerous other pathways including apoptosis, cell cycle regulation, DNA damage/repair, immune cell development and inflammation. Transcriptional regulation...

IRF1 is a transcription factor that regulates key processes in the immune system and in tumour suppression. To gain further insight into IRF1's role in these processes, we searched for new target genes by performing chromatin immunoprecipitation coupled to a CpG island microarray (ChIP–chip). Using this approach we identified 202 new IRF1-binding s...

Photooxidized tryptophan (TRP) in tissue culture medium elicits a transient cytochrome P450 (CYP1) induction response in cultured cells. We show here that exposure of TRP to window sunlight (aTRP) greatly increased the potency, efficacy, and duration of CYP1A induction by TRP in primary chick embryo hepatocytes and in vivo. Aqueous TRP exposed to s...