Biology subjects

Sammons, M. A.

Publications and source records attributed to Sammons, M. A..

9 recordsLinked to original sources

bioRxiv2024

Differential transcriptional activity of ΔNp63β is encoded by an isoform-specific C-terminus

p63 is a clinically-relevant transcription factor heavily involved in development and disease. Mutations in the p63 DNA-binding domain lead to severe developmental defects and overexpression of p63 plays a role in the progression of epithelial-associated cancers. Unraveling the specific biochemical mechanisms underlying these phenotypes is made challenging by the presence of multiple p63 isoforms and their shared and unique contributions to development and disease. Here, we explore the function of the p63 isoforms {Delta}Np63[a] and {Delta}Np63{beta} to determine the contribution of C-terminal splice variants on known and unique molecular and biochemical activities. Using RNA-seq and ChIP-seq on isoform-specific cell lines, we show that {Delta}Np63{beta} regulates both canonical {Delta}Np63[a] targets and a unique set of genes with varying biological functions. We demonstrate that the majority of genomic binding sites are shared, however the enhancer-associated histone modification H3K27ac is highly enriched at {Delta}Np63{beta} binding sites relative to {Delta}Np63[a]. An array of {Delta}Np63{beta} C-terminal mutants demonstrates the importance of isoform-specific C-terminal domains in regulating these unique activities. Our results provide novel insight into differential activities of p63 C-terminal isoforms and suggest future directions for dissecting the functional relevance of these and other transcription factor isoforms in development and disease.

molecular biology↗

bioRxiv2024

p53motifDB: integration of genomic information and tumor suppressor p53 binding motifs

The tumor suppressor gene TP53 encodes the DNA binding transcription factor p53 and is one of the most commonly mutated genes in human cancer. Tumor suppressor activity requires binding of p53 to its DNA response elements and subsequent transcriptional activation of a diverse set of target genes. Despite decades of close study, the logic underlying p53 interactions with its numerous potential genomic binding sites and target genes is not yet fully understood. Here, we present a database of DNA and chromatin-based information focused on putative p53 binding sites in the human genome to allow users to generate and test new hypotheses related to p53 activity in the genome. Users can query genomic locations based on experimentally observed p53 binding, regulatory element activity, genetic variation, evolutionary conservation, chromatin modification state, and chromatin structure. We present multiple use cases demonstrating the utility of this database for generating novel biological hypotheses, such as chromatin-based determinants of p53 binding and potential cell type-specific p53 activity. All database information is also available as a precompiled sqlite database for use in local analysis or as a Shiny web application.

bioinformatics↗

bioRxiv2024

Context dependent activity of p63-bound gene regulatory elements

The p53 family of transcription factors regulate numerous organismal processes including the development of skin and limbs, ciliogenesis, and preservation of genetic integrity and tumor suppression. p53 family members control these processes and gene expression networks through engagement with DNA sequences within gene regulatory elements. Whereas p53 binding to its cognate recognition sequence is strongly associated with transcriptional activation, p63 can mediate both activation and repression. How the DNA sequence of p63-bound gene regulatory elements is linked to these varied activities is not yet understood. Here, we use massively parallel reporter assays (MPRA) in a range of cellular and genetic contexts to investigate the influence of DNA sequence on p63-mediated transcription. Most regulatory elements with a p63 response element motif (p63RE) activate transcription, with those sites bound by p63 more frequently or adhering closer to canonical p53 family response element sequences driving higher transcriptional output. The most active regulatory elements are those also capable of binding p53. Elements uniquely bound by p63 have varied activity, with p63RE-mediated repression associated with lower overall GC content in flanking sequences. Comparison of activity across cell lines suggests differential activity of elements may be regulated by a combination of p63 abundance or context-specific cofactors. Finally, changes in p63 isoform expression dramatically alters regulatory element activity, primarily shifting inactive elements towards a strong p63-dependent activity. Our analysis of p63-bound gene regulatory elements provides new insight into how sequence, cellular context, and other transcription factors influence p63-dependent transcription. These studies provide a framework for understanding how p63 genomic binding locally regulates transcription. Additionally, these results can be extended to investigate the influence of sequence content, genomic context, chromatin structure on the interplay between p63 isoforms and p53 family paralogs.

genomics↗

bioRxiv2023

Shared gene regulatory strategies for the p53 and ATF4-dependent transcriptional networks

The master tumor suppressor p53 regulates multiple cell fate decisions, like cell cycle arrest and apoptosis, via transcriptional control of a broad gene network. Dysfunction in the p53 network is common in cancer, often through mutations that inactivate p53 or other members of the pathway. Induction of tumor-specific cell death by restoration of p53 activity without off-target effects has gained significant interest in the field. In this study, we explore the gene regulatory mechanisms underlying a putative anti-cancer strategy involving stimulation of the p53-independent Integrated Stress Response (ISR). Our data demonstrate the p53 and ISR pathways converge to independently regulate common metabolic and pro-apoptotic genes. We investigated the architecture of multiple gene regulatory elements bound by p53 and the ISR effector ATF4 controlling this shared regulation. We identified additional key transcription factors that control basal and stress-induced regulation of these shared p53 and ATF4 target genes. Thus, our results provide significant new molecular and genetic insight into gene regulatory networks and transcription factors that are the target of numerous antitumor therapies.

molecular biology↗

bioRxiv2021

A feedback loop between heterochromatin and the nucleopore complex controls germ-cell to oocyte transition during Drosophila oogenesis

Germ cells differentiate into oocytes that become totipotent upon fertilization. How the highly specialized oocyte acquires this distinct cell fate is poorly understood. During Drosophila oogenesis, H3K9me3 histone methyltransferase SETDB1 translocates from the cytoplasm to the nucleus of germ cells concurrent with oocyte specification. Here, we discovered that nuclear SETDB1 is required to silence a cohort of differentiation-promoting genes by mediating their heterochromatinization. Intriguingly, SETDB1 is also required for the upregulation of 18 of the ~30 nucleoporins (Nups) that comprise the nucleopore complex (NPC). NPCs in turn anchor SETDB1-dependent heterochromatin at the nuclear periphery to maintain H3K9me3 and gene silencing in the egg chambers. Aberrant gene expression due to loss of SETDB1 or Nups results in loss of oocyte identity, cell death and sterility. Thus, a feedback loop between heterochromatin and NPCs promotes transcriptional reprogramming at the onset of oocyte specification that is critical to establish oocyte identity.

developmental biology↗

bioRxiv2019

Locally acting transcription factors are required for p53-dependent cis-regulatory element activity

The master tumor suppressor p53 controls transcription of a wide-ranging gene network involved in apoptosis, cell cycle arrest, DNA damage repair, and senescence. Recent studies revealed pervasive binding of p53 to cis-regulatory elements (CRE), which are non-coding segments of DNA that spatially and temporally control transcription through the combinatorial binding of local transcription factors (TFs). Although the role of p53 as a strong trans-activator of gene expression is well known, the co-regulatory factors and local sequences acting at p53-bound CREs are comparatively understudied. We designed and executed a massively parallel reporter assay (MPRA) to investigate the effect of transcription factor binding motifs and local sequence context on p53-bound CRE activity. Our data indicate that p53-bound CREs are both positively and negatively affected by alterations in local sequence context and changes to co-regulatory TF motifs. We identified a SP1/KLF family motif located in an intronic p53 CRE that is required for the endogenous expression of the p53-dependent gene CCNG1. We also identified ATF3 as a factor that co-regulates the expression of the p53-dependent gene GDF15 through binding with p53 in an upstream CRE. Loss of either p53 or ATF3 severely reduces CRE activity and alters endogenous GDF15 mRNA levels in the cell. Our data suggests that p53 has the flexibility to cooperate with a variety of transcription factors in order to regulate CRE activity. By utilizing different sets of co-factors across CREs, we hypothesize that p53 activity is guarded against loss of any one regulatory partner allowing for dynamic and redundant control of p53-mediated transcription.

molecular biology↗

bioRxiv2018

Comparison of genotoxic vs. non-genotoxic stabilization of p53 provides insight into parallel stress-responsive transcriptional networks

The tumor suppressor protein p53 is activated in response to diverse intrinsic and extrinsic cellular stresses and controls a broad cell-protective gene network. Whether p53:DNA binding and subsequent transcriptional activation differs downstream of these diverse intrinsic and extrinsic activators within the same cell type is controversial. Using primary human fibroblasts, we assessed the genome-wide profile of p53 binding, chromatin structure, and transcriptional dynamics after either genotoxic or non-genotoxic activation of p53. Activation of p53 by treatment with either etoposide or the small molecule MDM2 inhibitor nutlin 3A yields strikingly similar genome-wide binding of p53 and concomitant changes to local chromatin modifications and structure. DNA damage, but not p53 activation per se, leads to increased expression of genes in an inflammatory cytokine pathway. Etoposide-mediated activation of this inflammation signature is inhibited by treatment with the NF-kB pathway inhibitor Bay 11-7082, but does not affect expression of canonical p53 target genes. Our data demonstrate that differential activation of p53 within the same cell type leads to highly similar genome-wide binding, chromatin dynamics, and gene expression dynamics, and that DNA damage-mediated signaling through NF-{kappa}B likely controls the observed pro-inflammatory cytokine gene expression pattern.

genomics↗

bioRxiv2018

Cell type-dependent control of p53 transcription and enhancer activity by p63

Transcriptional activation by p53 provides powerful, organism-wide tumor suppression. In this work, we demonstrate that the p53-induced transcriptome varies based on cell type, reflects cell type-specific activities, and is considerably more broad than previously anticipated. This behavior is strongly influenced by p53 engagement with differentially active cell type-specific enhancers and promoters. In epithelial cell types, p53 activity is dependent on the p53 family member p63, which displays widespread enhancer binding. Notably, we demonstrate that p63 is required for epithelial enhancer identity including enhancers used by p53 during stress-dependent signaling. Loss of p63, but not p53, leads to site-specific depletion of enhancer-associated chromatin modifications, suggesting that p63 functions as an enhancer maintenance factor in epithelial cells. Additionally, a subset of epithelial-specific enhancers is dependent on the activity of p63 providing a direct link between lineage determination and enhancer structure. These data suggest a broad, cell-intrinsic mechanism for regulating the p53-dependent cellular response to stress through differential regulation of cis-regulatory elements.

genomics↗

bioRxiv2018

The transcription factor Tfap2e/AP-2ε plays a pivotal role in maintaining the identity of basal vomeronasal sensory neurons

The identity of individual neuronal cell types is defined by the expression of specific combinations of transcriptional regulators that control cell type-specific genetic programs. The epithelium of the vomeronasal organ of mice contains two major types of vomeronasal sensory neurons (VSNs): 1) the apical VSNs which express vomeronasal 1 receptors (V1r) and the G-protein subunit Gi2 and; 2) the basal VSNs which express vomeronasal 2 receptors (V2r) and the G-protein subunit Gao. Both cell types originate from a common pool of progenitors and eventually acquire apical or basal identity through largely unknown mechanisms.\n\nThe transcription factor AP-2{varepsilon}, encoded by the Tfap2e gene, plays a role in controlling the development of GABAergic interneurons in the main and accessory olfactory bulb (AOB), moreover AP-2{varepsilon} has been previously described to be expressed in the VSNs. Here we show that AP-2{varepsilon} is expressed in postmitotic VSNs after they commit to the basal differentiation program. Loss of AP-2{varepsilon} function resulted in reduced number of basal VSNs and in an increased number of neurons expressing markers of the apical lineage. Our work suggests that AP-2{varepsilon}, which is expressed in late phases of differentiation, is not needed to initiate the apical-basal differentiation dichotomy but for maintaining the basal VSNs identity by preventing the expression of apical genes. Moreover, our data suggest that differentiated VSNs of mice retain a notable level of plasticity.\n\nHighlightsO_LIThe VNO contains two major cell types that are segregated in apical and basal regions of the VNO\nC_LIO_LIAP-2{varepsilon} is expressed in postmitotic basal vomeronasal sensory neurons.\nC_LIO_LIAP-2{varepsilon} is essential to maintain the identity of basal vomeronasal sensory neurons.\nC_LI

developmental biology↗