David Wotton
· Professor of Regulation of Gene Expression, Development and Tumor Progression by TGF beta SignalingVerifiedUniversity of Virginia · Biochemistry and Molecular Genetics
Active 1959–2026
About
David Wotton is a Professor of Biochemistry and Molecular Genetics at the University of Virginia School of Medicine. He holds a BS in Genetics from the University of York, England, and a PhD in Genetics from the Imperial Cancer Research Fund in London. His research disciplines include biochemistry, cancer biology, cell and developmental biology, epigenetics, genetics, metabolism, and molecular biology. His research focuses on the regulation of gene expression, development, and tumor progression by TGF beta signaling. He analyzes the role of TGIF proteins, which are transcriptional repressors regulating TGF beta signaling and retinoid-regulated gene expression. His work involves studying mutations in human TGIF associated with holoprosencephaly, and he has generated and analyzed Tgif1 and Tgif2 knockout mice to understand developmental defects such as holoprosencephaly and the disruption of Sonic Hedgehog signaling. Additionally, Wotton investigates the mechanisms of transcriptional repression by TGIFs, their interactions with nuclear receptors and neural zinc finger transcription factors, and their potential role in neural development, differentiation, and glioblastoma. He also researches the role of TGF beta signaling in prostate cancer progression, developing mouse models based on mutations in tumor suppressor genes such as Pten and Tgfbr2. His work aims to understand how these mutations contribute to tumor invasiveness and metastasis, and to evaluate therapeutic approaches. Overall, his contributions advance understanding of gene regulation during development and cancer, with a focus on TGF beta signaling pathways.
Research topics
- Genetics
- Cell biology
- Biology
- Chemistry
- Cancer research
- Endocrinology
- Biochemistry
Selected publications
bioRxiv (Cold Spring Harbor Laboratory) · 2026-04-14
articleOpen accessSenior authorCorrespondingABSTRACT The TGIF1 transcription factor gene is present on chromosome 18, which is subject to whole chromosome copy number reduction in colon cancer. Despite this, TGIF1 expression is significantly higher in cancer than in normal. In mice complete deletion of Tgif1 reduced tumor burden in an Apc mutant model of intestinal cancer. Here we show that reducing TGIF1 expression in a human colon cancer cell line slows proliferation and reduces growth of orthotopic xenografts. To ask if additional genes with copy number loss are more highly expressed in tumors we identified chromosomal regions subject to copy number reductions from ten TCGA cancer datasets. Within these regions a small proportion of genes, generally less than 10%, are expressed at higher levels in the tumor than in corresponding normal samples. Enrichment analysis using a set of 435 genes that have copy number reduction and increased expression identified mitosis as the most enriched gene set and FOXM1 and E2F family transcription factors as potential regulators. For mitotic genes, the average expression increase in tumor compared to normal is independent of copy number. In contrast, while DepMap common essential genes are generally more highly expressed in cancer than normal tissue, the relative increase in expression tracks well with copy number. Similarly, expression differences for gene sets such as S-phase, rRNA processing and DNA repair show increased expression in cancer versus normal, but changes also track with copy number. Thus, genes with increased expression despite copy number reduction may represent the output of key pro-tumorigenic transcriptional programs and could be potential therapeutic targets.
A simple method for analyzing competitive growth of multiple cell types in xenograft tumors
bioRxiv (Cold Spring Harbor Laboratory) · 2026-01-26 · 1 citations
articleOpen accessSenior authorCorrespondingABSTRACT Low take rates and inter-tumor variability in growth rates can limit the effectiveness of mouse xenograft models when comparing between groups. To address this problem we developed a simple method to compare multiple cell types within a single mixed xenograft. Individual cell lines or clones were transduced with a lentiviral vector that includes a unique PCR tag, allowing the use of qPCR to determine the proportion of each tagged cell type within a mixed xenograft tumor. We generated vectors with six distinct PCR tags, and two different selectable markers, and have optimized the approach for determining their relative proportions within a mix. An initial pre-amplification step is used to increase the amount of material for subsequent qPCR reactions. This also removes the bulk of the genomic DNA, increasing the specificity of the qPCR step. Samples are then used for qPCR with specific pairs of primers that distinguish between each of the individual PCR tags, and the relative proportion of each tag is determined relative to that in the starting mix. We have tested this approach for in vitro growth of mixed cell cultures and in an orthotopic cecal xenograft model using a human colon cancer cell line. Since each individual tumor is initiated with a mix of cells, multiple tumors within a single animal can be analyzed separately, and overall tumor size is not important. Similarly, multiple metastatic lesions from the same animal can be analyzed individually. Thus, each tumor provides a direct comparison between individually tagged cell lines or clones. This low throughput “bar-coding” approach is simple and cost effective and has the potential to reduce the number of animals needed for xenograft experiments.
Abstract 1207 Interaction of metabolic and epigenetic control in colon cancer
Journal of Biological Chemistry · 2024-03-01
articleOpen accessColorectal cancer (CRC) is third most common cause of cancer mortality worldwide. More then 150 000 new cases and 50 000 deaths from CRC are expected in the US in 2023. Although CRC can be treated by surgery, chemotherapy or targeted therapy, patients with advanced disease are resistant to current treatments. Acetyl-CoA synthetase (ACSS2) is a nuclear and cytosolic enzyme that converts acetate to Acetyl coenzyme A (Ac-CoA). In CRC, ACSS2 expression is decreased in both primary tumors and metastases, and lower expression correlates with reduced patient survival. Since Ac-CoA is essential for lipid synthesis and histone acetylation, we hypothesized that CRC with low ACSS2 might be sensitized to inhibition of ACSS2 or processes dependent on Ac-CoA. DOT1L (disruptor of telomeric silencing-1-like) is the only enzyme that methylates H3K79, a histone mark that promotes transcription elongation and DNA damage repair. High DOT1L expression in CRC has been linked to shorter survival and DOT1L knockdown has been shown to reduce the growth of subcutaneous CRC xenografts. We have tested the effect of inhibiting both ACSS2 and DOT1L in human CRC cell lines. CRISPR inactivation of ACSS2 increased sensitivity to DOT1L inhibitors in CRC cells. Combining small molecule inhibitors of ACSS2 and DOT1L resulted in a synergistic reduction in proliferation in multiple CRC cell lines, but had less effect in a normal colon epithelial line. RNA-seq data from CRC cells treated with ACS22 or DOT1L inhibitors alone identified relatively few gene expression changes, whereas in cells treated with both inhibitors gene expression changes were much more numerous. Interestingly, the majority of genes involved in the cholesterol synthesis pathway is deregulated by ACSS2 and/or DOT1L inhibition. Using a cell cell-based luciferase assay with an SREBP-responsive promoter, we found effects of DOT1L and ACSS2 inhibitors consistent with those seen by RNA-seq. Based on the effect on the cholesterol pathway, we tested the effects of statins, to inhibit cholesterol synthesis, together with either ACSS2 or DOT1L inhibition. These results show a synergistic effect of ACSS2 inhibition and statins in all CRC cell lines tested, as well as synergy between statins and DOT1L inhibitors in a subset. We are currently testing whether the effect of statins with ACSS2 inhibitors is due to a requirement for cholesterol, or for other biosynthetic intermediates of the cholesterol synthesis pathway. In summary we have found a synergistic interaction between ACSS2 and DOT1L inhibitors, which has profound effects on gene expression and proliferation in CRC cells in vitro, and identified cholesterol synthesis as a downstream target. Future work will test these combinations of inhibitors in pre-clinical CRC models. Research is funded from the NIH grant (CA259571).
2023-04-17
preprintOpen access<div><p>The ADP-ribosyltransferase PARP7 modulates protein function by conjugating ADP-ribose to the side chains of acceptor amino acids. PARP7 has been shown to affect gene expression in prostate cancer cells and certain other cell types by mechanisms that include transcription factor ADP-ribosylation. Here, we use a recently developed catalytic inhibitor to PARP7, RBN2397, to study the effects of PARP7 inhibition in androgen receptor (AR)-positive and AR-negative prostate cancer cells. We find that RBN2397 has nanomolar potency for inhibiting androgen-induced ADP-ribosylation of the AR. RBN2397 inhibits the growth of prostate cancer cells in culture when cells are treated with ligands that activate the AR, or the aryl hydrocarbon receptor, and induce PARP7 expression. We show that the growth-inhibitory effects of RBN2397 are distinct from its enhancement of IFN signaling recently shown to promote tumor immunogenicity. RBN2397 treatment also induces trapping of PARP7 in a detergent-resistant fraction within the nucleus, which is reminiscent of how inhibitors such as talazoparib affect PARP1 compartmentalization. Because PARP7 is expressed in AR-negative metastatic tumors and RBN2397 can affect cancer cells through multiple mechanisms, PARP7 may be an actionable target in advanced prostate cancer.</p>Significance:<p>RBN2397 is a potent and selective inhibitor of PARP7 that reduces the growth of prostate cancer cells, including a model for treatment-emergent neuroendocrine prostate cancer. RBN2397 induces PARP7 trapping on chromatin, suggesting its mechanism of action might be similar to clinically used PARP1 inhibitors.</p></div>
2023-04-17
supplementary-materialsOpen access<p>Supplementary Table 1: RBN2397 growth effects in prostate cencer cell lines</p>
2023-04-04
preprintOpen access<p>Cell cycle distributions of cell lines treated with RBN2397 plus androgen or plus AHR agonist</p>
2023-04-17
preprintOpen access<p>RBN2397 inhibition of prostate cancer cell growth requires PARP7 over-expression</p>
2023-04-04
preprintOpen access<div>Abstract<p>The ADP-ribosyltransferase PARP7 modulates protein function by conjugating ADP-ribose to the side chains of acceptor amino acids. PARP7 has been shown to affect gene expression in prostate cancer cells and certain other cell types by mechanisms that include transcription factor ADP-ribosylation. Here, we use a recently developed catalytic inhibitor to PARP7, RBN2397, to study the effects of PARP7 inhibition in androgen receptor-positive and androgen receptor-negative prostate cancer cells. We find that RBN2397 has nanomolar potency for inhibiting androgen-induced ADP-ribosylation of the androgen receptor. RBN2397 inhibits the growth of prostate cancer cells in culture when cells are treated with ligands that activate the androgen receptor, or the aryl hydrocarbon receptor, and induce PARP7 expression. We show that the growth inhibitory effects of RBN2397 are distinct from its enhancement of interferon signaling recently shown to promote tumor immunogenicity. RBN2397 treatment also induces trapping of PARP7 in a detergent-resistant fraction within the nucleus, which is reminiscent of how inhibitors such as Talazoparib affect PARP1 fractionation. Because PARP7 is expressed in AR negative metastatic tumors and RBN2397 can affect cancer cells through multiple mechanisms, PARP7 may be an actionable target in advanced prostate cancer.</p></div>
2023-04-17
preprintOpen access<p>Cell cycle distributions of cell lines treated with RBN2397 plus androgen or plus AHR agonist</p>
Cancer Research · 2023-06-02
articleAbstract The ADP-ribosyltransferase PARP7 modulates protein function by conjugating ADP-ribose to the side chains of acceptor amino acids. PARP7 has been shown to affect gene expression in prostate cancer cells and certain other cell types by mechanisms that include transcription factor ADP-ribosylation. Here, we use a recently developed catalytic inhibitor to PARP7, RBN2397 (NCT04053673), to study the effects of PARP7 inhibition in androgen receptor-positive (AR+) and androgen receptor-negative (AR-) prostate cancer cells. Ribon Therapeutics developed RBN2397 as a first-in-class mono-ADP-ribosyltransferase inhibitor, and showed that it blocks PARP7 negative regulation of TBK1 [1]. We find that RBN2397 has nanomolar potency for inhibiting androgen-induced ADP-ribosylation of the androgen receptor. RBN2397 inhibits the growth of prostate cancer cells in culture when cells are treated with ligands that activate the androgen receptor (PC3-AR, VCaP, CWR22Rv1), or the aryl hydrocarbon receptor (PC3, DU145, NCI-H660), and induce PARP7 expression. We show that the growth inhibitory effects of RBN2397 are distinct from its enhancement of interferon signaling recently shown to promote tumor immunogenicity in lung cancer models [1]. Chemical inhibitors to PARP1 exert effects on cells by blocking enzyme function, but also via cytotoxic effects attributed to stabilizing PARP1-chromatin interactions in a process termed trapping [2]. Drug-induced trapping of PARP1 can be detected biochemically by immunoblotting the detergent-resistant chromatin fraction. We found that RBN2397 treatment of AR+ and AR- prostate cancer cells induces biochemical trapping of PARP7 within the nucleus, which was also detected by confocal microscopy. Potential therapeutic benefits of RBN2397 are likely to depend on the level of PARP7 expression, given its induction is necessary for growth inhibitory effects of RBN2397 in cell culture. As a first step towards evaluating whether PARP7 levels in human prostate cancer may be actionable with RBN2397, we used computational methods to analyze PARP7 gene expression data from primary prostate tumors and metastatic AR+ and AR- prostate tumors. To assess PARP7 mRNA levels, we used data from the online resource recount3, which uniformly reprocesses publicly available RNA-seq datasets using a Monorail analysis pipeline. Using the level of PARP7 expression in VCaP cells that confers sensitivity to RBN2397 as a threshold, 50% of primary tumors, 41% of metastatic AR- and 11% of AR+ tumors are predicted to have PARP7 expression levels that are sufficient for a response to RBN2397. Because RBN2397 can inhibit the growth of castration-resistant and neuroendocrine prostate cancer cells, PARP7 may be an actionable target in advanced prostate cancer. 1. Gozgit, J.M., et al., PARP7 negatively regulates the Type I interferon response in cancer cells and its inhibition triggers antitumor immunity. Cancer Cell, 2021. 39(9): p. 1214-1226 e10 2. Murai, J., et al., Trapping of PARP1 and PARP2 by Clinical PARP Inhibitors. Cancer Res, 2012. 72(21): p. 5588-99. Citation Format: Chunsong Yang, Krzysztof Wierbilowicz, Natalia M. Dworak, Song Yi Bae, Sachi B. Tengse, Nicki Abianeh, Justin M. Drake, Tarek Abbas, Aakrosh Ratan, David Wotton, Bryce M. Paschal. Induction of PARP7 creates a vulnerability for growth inhibition by RBN2397 in prostate cancer cells [abstract]. In: Proceedings of the AACR Special Conference: Advances in Prostate Cancer Research; 2023 Mar 15-18; Denver, Colorado. Philadelphia (PA): AACR; Cancer Res 2023;83(11 Suppl):Abstract nr B072.
Recent grants
NIH · $26.2M · 2005–2017
NIH · $2.5M · 2012
Regulation of neural development by TGF beta family signaling
NIH · $1.7M · 2012–2019
TGF beta regulation of cilium-dependent signaling
NIH · $1.2M · 2012–2017
NIH · $1.5M · 2013
Frequent coauthors
- 36 shared
Bryce M. Paschal
University of Virginia
- 36 shared
Joan Massagué
Memorial Sloan Kettering Cancer Center
- 35 shared
Chunsong Yang
Sheng Jing Hospital
- 34 shared
Tiffany A. Melhuish
University of Virginia
- 31 shared
Natalia M. Dworak
University of Virginia
- 30 shared
Justin M. Drake
- 30 shared
Krzysztof Wierbiłowicz
- 30 shared
Nicki Abianeh
University of Virginia
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