Abstract 7245: Mechanistic dissection of ABI1 as DNA-binding transcriptional regulator in cancer cells

Abstract Background: ABI1 (Abelson interactor-1) is classically recognized as a multifunctional adaptor protein with homeostatic roles in cancer biology. It functions as a tumor suppressor in some cancer such as prostate cancer, yet exhibits oncogenic activity in other cancers such as for example breast cancer. Historically, ABI1 has been studied for its actin-cytoskeleton-associated functions—including cell-cell adhesion, cell motility, and lamellipodia formation—as well as its role in regulating major signaling hubs such as c-Abl, PI3K, and Src. Our recent findings reveal an unanticipated function of ABI1: direct DNA binding mediated by a conserved homeodomain homology region (HHR). This discovery led us to hypothesize that ABI1 may act as a previously unrecognized transcriptional regulator. Here, we sought to define the molecular mechanisms through which ABI1 contributes to transcriptional control. Methods: To determine sequence specificity and genomic occupancy, we performed ChIP using HHR-intact and HHR-mutant ABI1 constructs, complemented by in vitro DNA binding assays using purified proteins. Subcellular fractionation and chromatin enrichment assays assessed ABI1 nuclear localization and association with chromatin. ABI1-interacting transcriptional machinery was identified through co-immunoprecipitation (co-IP). RNA-seq comparing cells expressing wild-type ABI1 versus an HHR-defective DNA-binding mutant defined ABI1-dependent transcriptional outputs. Results: ABI1 binds DNA both in vitro and in vivo and displays reproducible sequence motifs from integrated ChIP and in vitro binding analyses. ABI1 variants containing an intact HHR domain localize preferentially to the nucleus and chromatin fractions. Co-IP studies identify ABI1 as a component of a defined transcriptional complex. RNA-seq analyses reveal that HHR-mediated DNA binding is required for a discrete ABI1-dependent transcriptional program. Conclusions: We identify ABI1 as a novel DNA-binding protein with sequence preference and transcriptional regulatory capacity mediated through its HHR domain. These findings expand the functional repertoire of ABI1 beyond actin regulation and kinase signaling, providing the first mechanistic framework for ABI1-driven transcriptional control. Citation Format: Kate Livingston, XIANG Li, Kevin M. Lin, Leszek Kotula. Mechanistic dissection of ABI1 as DNA-binding transcriptional regulator in cancer cells abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 7245.