Biology subjects

Bowman, B. M.

Publications and source records attributed to Bowman, B. M..

5 recordsLinked to original sources

bioRxiv2025

Pyrimethamine and a potent analogue WCDD115 inhibit NRF2 by suppressing DHFR and one-carbon metabolism

Nuclear factor erythroid 2-related factor 2 (NFE2L2/NRF2) is a critical mediator of the cellular oxidative stress response. Aberrant activation of NRF2 is common in lung and upper aerodigestive cancers, where it promotes tumor initiation and progression and confers resistance to chemotherapy, radiation therapy, and immune checkpoint inhibitors. As such, NRF2 therapeutic inhibitors are actively being sought. We previously reported that the antiparasitic drug Pyrimethamine (PYR) inhibits NRF2 in cell lines and in a NRF2-inducible genetically engineered mouse model. Here we design, synthesize, and define structure-activity relationships across a series of 25 PYR-based derivatives to reveal WCDD115 as a 22-fold more potent inhibitor of NRF2 (57nM versus 1.2{micro}M). PYR is known to inhibit plasmodial and human dihydrofolate reductase (DHFR). We found that WCDD115 inhibits hDHFR with 31-fold greater potency than PYR (144nM versus 4.49{micro}M). Metabolomics showed strong similarities between PYR, WCDD115 and methotrexate. Genetic, pharmacological and metabolic epistasis studies reveal that DHFR inactivation is required for NRF2 suppression by WCDD115 and PYR. Global and targeted proteomics revealed overlapping profiles for WCDD115, PYR and methotrexate, including suppression of NRF2 oxidative stress response and activation of TP53 and the DNA damage response. Therefore, PYR and a novel potent derivative WCDD115 are effective, indirect inhibitors of NRF2 and its antioxidant functions. These data underscore the importance of one- carbon metabolism for the NRF2 signaling pathway and support a new therapeutic strategy to suppress NRF2-driven cancer biology.

cancer biology↗

bioRxiv2023

SETD2 maintains nuclear lamina stability to safeguard the genome

Histone methyltransferases play essential roles in the organization and function of chromatin. They are also frequently mutated in human diseases including cancer1. One such often mutated methyltransferase, SETD2, associates co-transcriptionally with RNA polymerase II and catalyzes histone H3 lysine 36 trimethylation (H3K36me3) - a modification that contributes to gene transcription, splicing, and DNA repair2. While studies on SETD2 have largely focused on the consequences of its catalytic activity, the non-catalytic functions of SETD2 are largely unknown. Here we report a catalysis-independent function of SETD2 in maintaining nuclear lamina stability and genome integrity. We found that SETD2, via its intrinsically disordered N-terminus, associates with nuclear lamina proteins including lamin A/C, lamin B1, and emerin. Depletion of SETD2, or deletion of its N-terminus, resulted in widespread nuclear morphology abnormalities and genome stability defects that were reminiscent of a defective nuclear lamina. Mechanistically, the N-terminus of SETD2 facilitates the association of the mitotic kinase CDK1 with lamins, thereby promoting lamin phosphorylation and depolymerization required for nuclear envelope disassembly during mitosis. Taken together, our findings reveal an unanticipated link between the N-terminus of SETD2 and nuclear lamina organization that may underlie how SETD2 acts as a tumor suppressor.

cell biology↗

bioRxiv2023

Modulation of Retinal Inflammation Delays Degeneration in a Mouse Model of Geographic Atrophy

The advanced form of AMD, geographic atrophy, is associated with increased RPE oxidative stress and chronic inflammation. Here we evaluated the effects of delivering an anti-inflammatory viral gene by an AAV-vector in a mouse model of geographic atrophy. We measured changes in retinal function, structure, and morphology over nine months with electroretinography, optical coherence tomography, and fundoscopy, respectively. In addition, we used retinal tissue to quantify changes in markers of inflammation by multiplex ELISA, RT-qPCR, and immunofluorescence staining. Our AAV significantly delayed the loss of retinal function and structure and decreased retinal inflammation compared to the control AAV treatment. Our results suggest that modulating retinal inflammation could significantly slow the progression of geographic atrophy.

neuroscience↗

bioRxiv2021

Protein proximity networks and functional evaluation of the Casein Kinase 1 γ family reveals unique roles for CK1γ3 in WNT signaling

The WNT/{beta}-catenin signaling pathway is evolutionarily conserved and controls normal embryonic development, adult tissue homeostasis and regeneration. Aberrant activation or suppression of WNT signaling contributes to cancer initiation and progression, developmental disorders, neurodegeneration, and bone disease. Despite great need and more than 40 years of research, targeted therapies for the WNT pathway have yet to be fully realized. Kinases are exceptionally druggable and occupy key nodes within the WNT signaling network, but several pathway-relevant kinases remain understudied and dark. Here we studied the function of the CSNK1{gamma} subfamily of human kinases. miniTurbo-based proximity biotinylation and mass spectrometry analysis of CSNK1{gamma}1, CSNK1{gamma}2, and CSNK1{gamma}3 revealed numerous established components of the {beta}-catenin- dependent and independent WNT signaling pathway, as well as novel interactors. In gain-of- function experiments leveraging a panel of transcriptional reporters, CSNK1{gamma}3 but not CSNK1{gamma}1 or CSNK1{gamma}2 activated {beta}-catenin-dependent WNT signaling and the Notch pathway. Within the family, CSNK1{gamma}3 expression uniquely induced LRP6 phosphorylation. Conversely, siRNA- mediated silencing of CSNK1{gamma}3 alone had no impact on WNT signaling, though co-silencing of all three family members decreased WNT pathway activity. We characterized two moderately selective and potent small molecule inhibitors of the CSNK1{gamma} family. These inhibitors and a CSNK1{gamma}3 kinase dead mutant suppressed but did not eliminate WNT-driven LRP6 phosphorylation and {beta}-catenin stabilization. Our data suggest that while CSNK1{gamma}3 expression uniquely drives pathway activity, potential functional redundancy within the family necessitates loss of all three family members to suppress the WNT signaling pathway.

cell biology↗

bioRxiv2019

Gain-of-function genetic screen of the kinome reveals BRSK2 as an inhibitor of the NRF2 transcription factor

NFE2L2/NRF2 is a transcription factor and master regulator of cellular antioxidant response. Aberrantly high NRF2-dependent transcription is recurrent in human cancer, and conversely NRF2 protein levels as well as activity is diminished with age and in neurodegenerative disorders. Though NRF2 activating drugs are clinically beneficial, NRF2 inhibitors do not yet exist. Here we used a gain-of-function genetic screen of the kinome to identify new druggable regulators of NRF2 signaling. We found that the understudied protein kinase Brain Specific Kinase 2 (BRSK2) and the related BRSK1 kinases suppress NRF2-dependent transcription and NRF2 protein levels in an activity-dependent manner. Integrated phosphoproteomics and RNAseq studies revealed that BRSK2 drives AMPK activation and suppresses mTOR signaling. As a result, BRSK2 kinase activation suppressed ribosome-RNA complexes, global protein synthesis, and NRF2 protein levels. Collectively, our data establish the catalytically active BRSK2 kinase as a negative regulator of NRF2 via the AMPK/mTOR signaling. This signaling axis may prove useful for therapeutically targeting NRF2 in human diseases.\n\nSummary StatementBRSK2 suppresses NRF2 signaling by inhibiting protein synthesis through mTOR downregulation.

cell biology↗