Biology subjects

Mathew, V.

Publications and source records attributed to Mathew, V..

8 recordsLinked to original sources

bioRxiv2025

DNA2 variant analysis supports the nuclease activity as a preferred therapeutic target

DNA2 is a combined nuclease and helicase that processes long 5 flaps and other structures that occur during DNA replication and repair. Current models invoke an essential function of DNA2 in processing stalled replication forks to dampen toxic recombination based restart. As an essential replication stress response factor, DNA2 has been proposed as an anti-cancer drug target with several tool compounds developed. Here we sought to model inhibited DNA2 states using dominant genetics with separation-of-function alleles to identify the optimal mechanisms for DNA2 targeting. We find that expression of DNA2 nuclease-dead alleles exert dominant effects on fitness and DNA damage repair that are dependent on its helicase and RPA binding activities. Furthermore, we find an imperfect link wherein a subset of ALT+ models are hypersensitive to the presence of DNA2-nuclease dead protein. These results were replicated with a nuclease-specific inhibitor of DNA2. Together, our data suggests that DNA2 targeting in cancer should be nuclease focused and will rely on identification of specific biomarker subsets within ALT+ or other tumor cell states.

cell biology↗

bioRxiv2024

Mapping intratumoral myeloid-T cell interactomes at single-cell resolution reveals targets for overcoming checkpoint inhibitor resistance

Effective cancer immunotherapies restore anti-tumor immunity by rewiring cell-cell communication. Treatment-induced changes in communication can be inferred from single-cell RNA-sequencing (scRNA-seq) data, but current methods do not effectively manage heterogeneity within cell types. Here we developed a computational approach to efficiently analyze scRNA-seq-derived, single-cell-resolved cell-cell interactomes, which we applied to determine how agonistic CD40 (CD40ag) alters immune cell crosstalk alone, across tumor models, and in combination with immune checkpoint blockade (ICB). Our analyses suggested that CD40ag improves responses to ICB by targeting both immuno-stimulatory and immunosuppressive macrophage subsets communicating with T cells, and we experimentally validated a spatial basis for these subsets with immunofluorescence and spatial transcriptomics. Moreover, treatment with CD40ag and ICB established coordinated myeloid-T cell interaction hubs that are critical for reestablishing antitumor immunity. Our work advances the biological significance of hypotheses generated from scRNA-seq-derived cell-cell interactomes and supports the clinical translation of myeloid-targeted therapies for ICB-resistant tumors.

systems biology↗

bioRxiv2021

DYNLL1 mis-splicing is associated with replicative genome instability in SF3B1 mutant cells

Genome instability is a hallmark of cancer that arises through a panoply of mechanisms driven by oncogene and tumour-suppressor gene mutations. Oncogenic mutations in the core splicing factor SF3B1 have been linked to genome instability. Since SF3B1 mutations alter the selection of thousands of 3 splice sites affecting genes across biological pathways, it is not entirely clear how they might drive genome instability. Here we confirm that while R-loop formation and associated replication stress may account for some of the SF3B1-mutant genome instability, a mechanism involving changes in gene expression also contributes. An SF3B1-H662Q mutant cell line mis-splices the 5UTR of the DNA repair regulator DYNLL1, leading to higher DYNLL1 protein levels, mis-regulation of DNA repair pathway choice and PARP inhibitor sensitivity. Reduction of DYNLL1 protein in these cells restores genome stability. Together these data highlight how SF3B1 mutations can alter cancer hallmarks through subtle changes to the transcriptome.

cell biology↗

bioRxiv2020

The Effect of Refractive Blur in the Vividness of Mental Imagery

BackgroundVisual mental imagery or "seeing with the minds eye" is an everyday phenomenon. Visual mental imagery and visual perception share common neural-networks. Hence deficits that affect the visual perception may also affect visual mental imagery. AimWe aimed to study the effect of refractive blur on the vividness of mental imagery. MethodsSubjects were recruited from volunteers and divided into two groups; individuals with refractive errors-Ametropes(AM), and individuals without refractive errors - Emmetropes(EM). After filling in the Verbalizer-Visualizer-Questionnaire (VVQ), the subjects were asked to perform a mental imagery task with and without refractive blur. The participants were asked to generate a mental image of a specific object initially with eyes closed, eyes open and then with refractive blur in random order, and then judge the vividness of the mental image on a Likert scale ranging from 1 (low vividness) to 5 (good vividness). The EM participants had to wear a + 2D spectacles to produce refractive blur. ResultsA total of 162 participants were recruited to the study. Of these 73 were EM and 89 were AM. Of the AM, 30 had additional astigmatism. The mean VVQ score was 64.9(11.2). The mean refractive error was 1.8(1.3)D. Following the mental imagery task, at baseline with eyes closed, 138 (85.5%)subjects had vivid mental imagery close to visual perception(Likert scale:5). With the opening of the eyes, the vividness dropped by at least 1 point in the Likert scale in 139(85.8%). With the introduction of refractive blur, 153(94.4%) subjects had a drop in the vividness of the image by at least 1 point and 22(13.6%) subjects by at least 2 points. ConclusionIntroduction of refractory blur results in the reduction of the vividness of mental imagery.

neuroscience↗

bioRxiv2020

Cdc48 regulates intranuclear quality control sequestration of the Hsh155 splicing factor

Cdc48/VCP is a highly conserved ATPase chaperone that plays an essential role in the assembly or disassembly of protein-DNA complexes and in degradation of misfolded proteins. We find that Cdc48 accumulates during cellular stress at intranuclear protein quality control (INQ) sites. Cdc48 function is required to suppress INQ formation under non-stress conditions and to promote recovery following genotoxic stress. Cdc48 physically associates with the INQ substrate and splicing factor Hsh155 and regulates its assembly with partner proteins. Accordingly, cdc48 mutants have defects in splicing and show spontaneous distribution of Hsh155 to INQ aggregates where it is stabilized. Overall, this study shows that Cdc48 regulates deposition of proteins at INQ and suggests a previously unknown role for Cdc48 in the regulation or stability of splicing subcomplexes.

molecular biology↗

bioRxiv2018

The yeast core spliceosome maintains genome integrity through R-loop prevention and alpha-tubulin expression

To achieve genome stability cells must coordinate the action of various DNA transactions including DNA replication, repair, transcription and chromosome segregation. How transcription and RNA processing enable genome stability is only partly understood. Two predominant models have emerged: one involving changes in gene expression that perturb other genome maintenance factors, and another in which genotoxic DNA:RNA hybrids, called R-loops, impair DNA replication. Here we characterize genome instability phenotypes in a panel yeast splicing factor mutants and find that mitotic defects, and in some cases R-loop accumulation, are causes of genome instability. Genome instability in splicing mutants is exacerbated by loss of the spindle-assembly checkpoint protein Mad1. Moreover, removal of the intron from the -tubulin gene TUB1 restores genome integrity. Thus, while R-loops contribute in some settings, defects in yeast splicing predominantly lead to genome instability through effects on gene expression.

cell biology↗

bioRxiv2017

Transient telomerase inhibition alters cell cycle kinetics

Telomerase is the ribonucleoprotein reverse transcriptase that catalyzes the synthesis of telomeres at the ends of linear human chromosomes and contributes to proper telomere-loop (T-loop) formation. Formation of the T-loop, an obligate step before cell division can proceed, requires the generation of a 3-overhang on the G-rich strand of telomeric DNA via telomerase or C-strand specific nucleases. Here, we discover telomerase activity is critical for efficient cell cycle progression using transient chemical inhibition by the telomerase inhibitor imetelstat. Telomerase inhibition caused changes in cell cycle kinetics and increased the proportion of cells in G2 phase, suggesting delayed clearance through this checkpoint. Investigating the possible contribution of unstructured telomere ends to these cell cycle distribution changes, we observed that imetelstat treatment induced {gamma}H2AX DNA damage foci in a subset of telomerase-positive cells but not telomerase-negative primary human fibroblasts. Chromatin-immunoprecipitation with {gamma}H2AX antibodies demonstrated imetelstat treatment-dependent enrichment of this DNA damage marker at telomeres. Notably, the effects of telomerase inhibition on cell cycle profile alterations were abrogated by pharmacological inhibition of the DNA-damage-repair transducer ATM.\n\nAdditionally, imetelstat potentiation of etoposide, a DNA-damaging drug that acts preferentially during S/G2 phases of the cell cycle, also depended on functional ATM signaling. Our results suggest that telomerase inhibition delays the removal of ATM-dependent DNA damage signals from telomeres in telomerase-positive cancer cells. This demonstrates for the first time that telomerase activity directly facilitates the progression of the cell cycle through modulation of transient telomere dysfunction signals.

cancer biology↗

bioRxiv2017

Genotoxin-induced transcriptional repression regulates selective protein aggregation

Upon genotoxic stress, dynamic relocalization events control DNA repair, and alterations of the transcriptome and proteome enabling stress recovery. How these events may influence one another is only partly known. Beginning with a cytological screen for genome maintenance proteins that move under stress, we find that, upon alkylation stress, the splicing factor Hsh155 localizes to both intranuclear and cytoplasmic protein quality control aggregates. Under stress, an ordered sequestration of Hsh155 occurs at nuclear and then cytoplasmic aggregates in a manner that is regulated by molecular chaperones. This dynamic behavior is preceded by a decrease in splicing efficiency. While DNA replication stress signaling is not required for Hsh155 sequestration, Hsh155 aggregation is cell cycle and TOR pathway dependent. Indeed, loss of a TORC1 regulated ribosomal protein gene transcription factor Sfp1 allows general aggregate formation but prevents Hsh155 recruitment. Together, our analyses suggest a model in which some proteins evicted from chromatin undergoing transcriptional remodeling during stress are targeted to protein quality control sites.

cell biology↗