Abstract
BackgroundIntegrins play a critical role in carcinogenesis. Integrin β1 localization is regulated by the guanosine-5′-triphosphate hydrolase Rab25 and integrin β1 levels are elevated in the serum of colon cancer patients; thus, the present study examined the effects of epidermal growth factor (EGF) and Rab25 on integrin β1 localization in colon cancer cells.MethodsHCT116 human colon cancer cells were treated with increasing concentrations of EGF, and cell proliferation and protein expression were monitored by MTT and western blot analyses, respectively. Cell fractionation was performed to determine integrin β1 localization in the membrane and cytosol. Integrin β1 extracellular shedding was monitored by enzyme-linked immunosorbent assays (ELISAs) with culture supernatants from stimulated cells. HCT116 cells were transfected with Rab25-specific siRNA to determine the significance of Rab25 in integrin β1 trafficking in the presence of EGF.ResultsTotal integrin β1 expression increased in response to EGF and subsequently decreased at 24 h post-stimulation. A similar decrease was observed in purified membrane fractions, whereas no changes were observed in cytosolic levels. ELISAs using media from stimulated cell cultures demonstrated increased integrin β1 levels corresponding to the decrease observed in membrane fractions, suggesting that EGF induces integrin receptor shedding. EGF stimulation in Rab25-knockdown cells resulted in integrin β1 accumulation in the membrane, suggesting that Rab25 promotes integrin endocytosis.ConclusionsIntegrin β1 is shed from colon cancer cells in response to EGF stimulation in a Rab25-dependent manner. These results further the present understanding of the role of integrin β1 in colon cancer progression.
Highlights
Integrins play a critical role in carcinogenesis
Integrin β1 has been identified as a drug target in several solid tumors [7, 19,20,21,22]; further analysis of integrin signaling may have implications for cancer therapy [6, 11]. Because both epidermal growth factor (EGF)/epidermal growth factor receptor (EGFR) pathway activation and integrin β1 have been associated with colon cancer progression, the present study aimed to investigate changes in integrin β1 expression and trafficking in response to EGF stimulation
These analyses revealed that HCT116 cell proliferation gradually increased in a concentration- and time-dependent manner (Fig. 1)
Summary
Integrins play a critical role in carcinogenesis. Integrin β1 localization is regulated by the guanosine5′-triphosphate hydrolase Rab and integrin β1 levels are elevated in the serum of colon cancer patients; the present study examined the effects of epidermal growth factor (EGF) and Rab on integrin β1 localization in colon cancer cells. Integrins are a family of heterodimeric transmembrane proteins composed of α and β subunits that function as bidirectional receptors for extracellular matrix proteins to regulate cell adhesion, motility, and proliferation [1]. These activities are dynamically regulated by receptor internalization and recycling back to the plasma membrane, a process known as trafficking [2,3,4,5]. Integrin β1 is detected in the serum of colon cancer patients, where its expression correlates with stage, invasive potential, and the presence of micrometastasis [16].
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