Abstract
The most frequently mutated protein in human cancer is p53, a transcription factor (TF) that regulates myriad genes instrumental in diverse cellular outcomes including growth arrest and cell death. Cell context-dependent p53 modulation is critical for this life-or-death balance, yet remains incompletely understood. Here we identify sequence signatures enriched in genomic p53-binding sites modulated by the transcription cofactor iASPP. Moreover, our p53-iASPP crystal structure reveals that iASPP displaces the p53 L1 loop-which mediates sequence-specific interactions with the signature-corresponding base-without perturbing other DNA-recognizing modules of the p53 DNA-binding domain. A TF commonly uses multiple structural modules to recognize its cognate DNA, and thus this mechanism of a cofactor fine-tuning TF-DNA interactions through targeting a particular module is likely widespread. Previously, all tumor suppressors and oncoproteins that associate with the p53 DNA-binding domain-except the oncogenic E6 from human papillomaviruses (HPVs)-structurally cluster at the DNA-binding site of p53, complicating drug design. By contrast, iASPP inhibits p53 through a distinct surface overlapping the E6 footprint, opening prospects for p53-targeting precision medicine to improve cancer therapy.
Highlights
The most frequently mutated protein in human cancer is p53, a transcription factor (TF) that regulates myriad genes instrumental in diverse cellular outcomes including growth arrest and cell death
We focus on the inhibitory iASPP and used RNA sequencing (RNA-seq) combined with chromatin immunoprecipitation followed by sequencing (ChIP-seq) to investigate genome-wide p53 binding and transcriptional activities regulated by iASPP in the HCT 116 colorectal carcinoma cell line, which harbors wild-type p53
Our structure of the p53 DNA-binding domains (DBDs) in complex with iASPP exemplifies a mechanism through which direct interactions with partner proteins impart DNA-binding refinement and target gene selectivity to transcription factors
Summary
Sequence Signatures Enriched in iASPP-Regulated p53 REs. To expand knowledge of iASPP-regulated p53 target gene selectivity from a handful of tested promoters to the breadth of the human log2[fold change]. Apart from CDKN1A, iASPP has not been studied in relation to the transcriptional regulation of these iPTGs. Following iASPP depletion, our RNA-seq and ChIP-seq data showed strong upregulation of transcription and enhanced p53 binding for TIGAR (Fig. 2B), which modulates metabolism and lowers intracellular reactive oxygen species (ROS) levels in response to mild metabolic stress signals in favor of cell survival [52]. Following iASPP depletion, our RNA-seq and ChIP-seq data showed strong upregulation of transcription and enhanced p53 binding for TIGAR (Fig. 2B), which modulates metabolism and lowers intracellular reactive oxygen species (ROS) levels in response to mild metabolic stress signals in favor of cell survival [52] We confirmed this result in a luciferase reporter assay using p53-null cell lines H1299 and Saos-2, in which iASPP inhibited p53-mediated transactivation of a TIGAR response element (Fig. 2D). The results from this binding assay, in conjunction with the structural
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