Abstract: Compared to hormone receptor (HR) positive breast cancer (BC), basal or triple negative BC (TNBC) suffers a poor prognosis and limited treatment options because of the lack of understanding of its driver signaling pathways. Mitochondrial retrograde signaling is a pathway of communication from mitochondria to the nucleus. We have previously published that TNBC has energy dependency to mitochondrial FA β-oxidation (FAO). FAO induces tumor and metastatic potential of TNBC. Src oncopathway is one of the most frequently upregulated pathways in TNBC and a critical player of TNBC metastasis. Like most protein kinases, Src family kinases (SFKs) require phosphorylation within a segment of the kinase for its full catalytic activity. Src is mainly phosphorylated (pSrc) at either Y419 (active state) or Y530 (inactive state). Compared to most BC subgroups, pSrc (Y419) is significantly upregulated in TNBC subgroups. However, how pSrc oncopathway is activated in TNBC was largely unknown. In this context, our lab made a breakthrough discovery that in TNBC, FAO regulates the phosphorylation of Src at Y419 without any major impact on the phosphorylation of Y530. Though we have shown that FAO inhibition decreases the pSrc (Y419), so far, the mechanism of this critical regulation is not fully understood. Only very fewer proteins have been proposed for the removal of phosphorylation from pSrc (Y419). Disabled-2 (DAB2), is known to bind at SH3 domain of Src and directly suppress the Y419 phosphorylation. This dephosphorylation results in the inactivation for Src without affecting the phosphorylation at Y530. Previous studies have suggested that the epigenetic transcriptional downregulation of DAB2 is a major determinant of cancer progression and clinical outcome. FA and FAO metabolites are known to epigenetically modify nuclear genes. Our strong preliminary data using different models suggest that the FAO downregulate the DAB2 mRNA in TNBC. Moreover, HDAC inhibitors activate DAB2 mRNA in TNBC cells. All these suggest that FAO may be epigenetically regulating DAB2. Thus, in this project we will evaluate how FAO-mediated retrograde signaling epigenetically modify the tumor suppressor DAB2. We will also analyze if such regulation of DAB2 is critical in the FAO-mediated activation of Src oncopathway in TNBC. In Aim-1, FAO-mediated transcriptional regulation of DAB2 will be confirmed using TNBC cell lines and already available tumor tissues collected after long-term treatment with FAO inhibitors in mice bearing TNBC patient-derived xenograft (PDX) models. We also have TNBC cells after stably knocking down FAO rate-limiting enzymes. Aim-2 will analyze the epigenetic modifications of DAB2 promoter after metabolic modulation. Overall, this will be one of the first mechanical studies in TNBC to show how the mitochondrial retrograde regulation modulate a tumor suppressor gene to control a major oncopathway via its post-translational regulation. The investigator team i...