Abstract 2658: GATA2 loss promotes Piezo1 expression and stiffness-dependent invasion in uterine serous carcinoma

Abstract Background: Piezo1 is a mechanosensitive ion channel that couples extracellular matrix (ECM) properties to intracellular signaling cascades that can modulate cell phenotypes. Piezo1 was recently shown to regulate metastasis in high-grade ovarian serous carcinoma (HGSOC) by promoting tumor budding in an ECM stiffness-dependent manner. Uterine serous carcinoma (USC) is an aggressive gynecologic malignancy that is increasing in incidence and responsible for 40% of uterine cancer deaths. USC rapidly invades and metastasizes via mechanisms that remain incompletely understood. Whether Piezo1 may have a similar function in USC, which shares many molecular and clinical features of HGSOC, is not known. We recently showed that expression of the transcription factor GATA2 is lost in USCs that locally invade into the wall of the uterus versus non-invasive USCs. However, the transcriptional targets of GATA2 that mediate this activity are not known. Methods: We performed RNAseq in patient-derived USC cells lines after GATA2 depletion to identify differentially expressed genes. We performed anti-GATA2 ChIPseq in patient derived USC cells to identify genome wide sites of GATA2 occupancy. Western blots were performed for GATA2, Piezo1, and Tubulin after GATA2 depletion in patient-derived USC cells modified to express doxycycline-inducible anti-GATA2 shRNAs or shScramble control. In vitro models of ECM with different stiffnesses (3.7 and 14 kPa) were generated using an interpenetrating network of collagen I and methacrylated gelatin (GelMA), crosslinked within transwell inserts. USC cells were seeded on top of the gels and monitored over 24 hours for invasion in response to GATA2 depletion. Results: GATA2 depletion led to an increase in Piezo1 transcripts in patient-derived USC cells (log2fold change = 0.28, FDR<0.005), while western blot revealed a significant increase in Piezo1 protein expression after GATA2 depletion in two unique patient-derived USC cell lines. Anti-GATA2 ChIPseq identified two significant peaks of GATA2 occupancy within intron 1 of the Piezo1 gene, each harboring conserved canonical GATA motifs. Depletion of GATA2 in patient-derived USC cells led to increased tumor cell invasion, but only on the stiffer substrates. Conclusions: GATA2 binds two putative GATA enhancers within intron 1 of the Piezo1 gene in USC. GATA2 depletion leads to increased Piezo1 RNA and protein expression, which is accompanied by increased tumor cell invasion in a stiffness-dependent manner. Our findings support a model where loss of GATA2-mediated Piezo1 suppression facilitates increased Piezo1 expression, and Piezo1-driven ECM stiffness-dependent USC tumor invasion. They justify further studies to evaluate whether a GATA2-Piezo1 axis is a critical driver of USC invasion in vivo. Citation Format: Brittany Baikie, Mayuri Dutta, Pamela K. Kreeger, Daniel R. Matson. GATA2 loss promotes Piezo1 expression and stiffness-dependent invasion in uterine serous carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 2658.