Abstract 6605: Understanding the role of sumoylated Etv1 in mammary oncogenesis

Abstract

Abstract The Small Ubiquitin-like Modifier (SUMO) pathway is a major post-translational regulatory mechanism involved in a variety of cellular functions including the DNA damage response, transcriptional regulation, and carcinogenesis. Recent discoveries by our group demonstrated that inhibiting the SUMO pathway can repress the growth of breast cancer xenografts and Neu-driven mammary oncogenesis in a genetically engineered mouse model with conditional knockout (CKO) of the SUMOylation E2 conjugating enzyme, Ube2i. However, despite the consistency of the findings, there remains considerable debate concerning the mechanisms through which SUMO inhibition represses tumorigenesis. Our downstream analysis suggested ETS Translocation Variant 1, encoded by ETV1 gene, acts as a SUMO-sensitive transcription factor that drives mammary oncogenesis and cancer growth. To examine oncogenic properties of SUMO-conjugated ETV1 in breast cancer, we cloned HA-tagged WT Etv1, SUMO-resistant Etv15xKR, or SUMO mimetic SUMO-1-Etv15xKR into a lentiviral vector with a GFP cassette. Proliferation, colony formation in soft agar, and in vivo xenograft assays were evaluated in the Ube2iF/F cell line derived from a tumor in a MMTV-c-neu/Ube2ifl/fl animal. We did ChIP-seq in Ube2iF/F cells using SUMO-1 and Etv1 antibodies to examine the effects of SUMO conjugation on Etv1 genomic occupancy. RNA-seq using siEtv1 or AdCre was also performed to compare changes in gene expression with SUMO-conjugated Etv1 genomic occupancy. Inhibition of SUMOylation via AdCre caused a significant reduction in proliferation and colony formation in vitro and completely block the formation of flank xenografts in vivo. Interestingly, expressing a SUMO-resistant ETV15xKR rescued in vitro growth effects, whereas a SUMO mimetic SUMO-1-ETV15xKR had no effect on restoring proliferation. On the contrary, in vivo xenograft assays demonstrated that the Etv15xKR-SUMO1 efficiently induced tumor formation despite the loss of Ube2i while the sumo-resistant Etv15xKR was unable to rescue tumorigenesis. Bulk RNA-seq and ChIP seq performed demonstrated different patterns of gene regulation between sumoylated and unsumoylated Etv1. Gene set enrichment analysis (GSEA) showed genes associated with cancer stemness were significantly enriched in cells expressing Etv15xKR-SUMO1. Collectively, these observations support Etv1 acting as a SUMO-sensitive transcription factor that drives a stem cell phenotype necessary for mammary oncogenesis and progression. Citation Format: Zhijie Li, Kelsey E. Koch, Dakota T. Thompson, Dana M. Van der Heide, Jeremy Chang, Christopher M. Franke, Mohammed O. Suraju, Anna C. Beck, Allison W. Lorenzen, Jeffrey R. White, Mikhail V. Kulak, David K. Meyerholz, Colin Kenny, Ronald J. Weigel. Understanding the role of sumoylated Etv1 in mammary oncogenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6605.