The cells' ability to proliferate in response to growth factor stimulation is significantly altered during cancer progression. To investigate the mechanisms underlying these alterations in prostate cancer, the role and expression of β_1A integrin and type 1 insulin-like growth factor receptor (IGF-IR), known to contribute to cell proliferation and transformation, were analyzed. Using small interfering RNA oligonucleotides to down-regulate β_1A, we show that β_1A expression is required for IGF-IR–mediated prostate cancer cell proliferation and anchorage-independent growth. In vivo, using age-matched transgenic adenocarcinoma of mouse prostate (TRAMP) mice at different stages of prostate cancer [prostatic intraepithelial neoplasia, PIN; well-differentiated adenocarcinoma, WD; and poorly differentiated adenocarcinoma, PD], the expression of β_1A and of IGF-IR was studied. β_1A and IGF-IR expression levels were concurrently up-regulated in high PIN and WD, whereas their expression did not correlate in late-stage PD. In contrast to the up-regulated expression of β_1A, the levels of β_1C, a β₁ cytoplasmic variant that inhibits cell proliferation, were down-regulated in all stages of prostate cancer. A similar expression pattern was observed for a β_1C downstream effector, Grb2-associated binder-1 (Gab1) which is known to inhibit IGF-IR phosphorylation. To analyze in vitro the mechanistic implications of β_1A, β_1C, and Gab1 deregulation in prostate cancer, we investigated whether expression of either β₁ variant in β₁-null cells affected IGF-IR localization. We found that IGF-IR and β_1A were colocalized in highly specialized integrin signaling compartments, designated focal contacts. However, in the presence of β_1C, IGF-IR remained diffuse on the cell surface and did not localize to focal contacts. The findings that β₁ integrins and IGF-IR are concurrently deregulated and that expression of β₁ integrins is necessary to achieve appropriate IGF-IR intracellular distribution point to the important role that the cross-talk between these receptors may have during prostate cancer progression and will be helpful in formulating new therapeutic strategies.