Kindlin-2 regulates the oncogenic activities of integrins and TGF-β in triple-negative breast cancer progression and metastasis

Kindlin-2, an adapter protein, is dysregulated in various human cancers, including triple-negative breast cancer (TNBC), where it drives tumor progression and metastasis by influencing several cancer hallmarks. One well-established role of Kindlin-2 involves the regulation of integrin signaling, achieved by directly binding to the cytoplasmic tail of the integrin beta subunit. In this study, we present novel insights into Kindlin-2's involvement in stabilizing the beta 1-Integrin:TGF-beta type 1 receptor (T beta RI) complexes, acting as a physical bridge that links beta 1-Integrin to T beta RI. Loss of Kindlin-2 results in the degradation of this protein complex, leading to the inhibition of downstream oncogenic pathways. We used a diverse range of in vitro assays, including CRISPR/Cas9 gene editing, cell migration, 3D-tumorsphere formation and invasion, solid binding, co-immunoprecipitation, cell adhesion and spreading assays, as well as western blot and flow cytometry analyses, utilizing MDA-MB-231 and 4T1 TNBC cell lines. Additionally, preclinical in vivo mouse models of TNBC tumor progression and metastasis were employed to substantiate our findings. Our studies established the direct interaction between Kindlin-2 and beta 1-Integrin and between Kindlin-2 and T beta RI. Disruption of these interactions, via CRISPR/Cas9-mediated knockout of Kindlin-2, led to the degradation of beta 1-Integrin and T beta RI, resulting in the inhibition of oncogenic pathways downstream of both proteins, subsequently hindering tumor growth and metastasis. Treatment of Kindlin-2-deficient cells with the proteasome inhibitor MG-132 restored the expression of both beta 1-Integrin and T beta RI. Furthermore, the rescue of Kindlin-2 expression reinstated their oncogenic activities in vitro and in vivo, while Kindlin-2 lacking domains involved in the interaction of Kindlin-2 with beta 1-Integrin or T beta RI did not. This study identifies a novel function of Kindlin-2 in stabilizing the beta 1-Integrin:T beta RI complexes and regulating their downstream oncogenic signaling. The translational implications of these findings are substantial, potentially unveiling new therapeutically targeted pathways crucial for the treatment of TNBC tumors.