Integrin heterodimers play crucial roles in epithelial adhesion, proliferation, differentiation, wound healing and tumorigenesis. However, the effects of acute loss of many of the 26 integrin subunits in epidermis have not been determined. This information is critical since many inhibitors for specific integrin heterodimers are entering clinical trials for various cancers. We performed an shRNA-based multiplexed screen to define roles for each integrin subunit in a 3-dimensional in vitro organotypic skin model. In this screen, we transduced individual populations of human keratinocytes with shRNAs against each integrin subunit. We then quantitatively determined the fitness of each cell population in a competition assay in reconstituted human skin. This screen identified the integrin αv class of heterodimers as essential for generation of human skin tissue: the population of keratinocytes lacking αv was selected against 125-fold compared to non-silencing control cells (p<0.001). An identical screen in human melanocytes showed no change in fitness upon loss of integrin αv, indicating cell-intrinsic functions for this subunit. Loss of integrin αv alone in keratinocytes leads to cell cycle arrest with a 4-5 fold increase in the G1/S ratio. Surprisingly, we found that integrin αv is not localized within focal adhesions in human keratinocytes and does not control focal adhesion dynamics in this cell type. Instead, it forms an independent signaling complex at the cellular membrane that controls keratinocyte growth through a FAK c-myc signaling axis. Both FAK and c-myc total protein levels are lost upon αv knock down, and re-expression of c-myc in αv-null keratinocytes is sufficient to completely rescue the cell cycle defects and skin tissue phenotypes. The effects of integrin αv loss are dependent on αv's binding partners β5 and β6, but not β1 or β8. Using a doxycycline-inducible system, we show that αv loss plays a crucial role in tissue generation and tumor invasion, but is not necessary for epidermal tissue maintenance. Furthermore, both β5 and β6 binding partners are required for tumor invasion, thus identifying αvβ5 and αvβ6 as therapeutic targets for metastatic squamous cell carcinoma.
Citation Format: Elizabeth K. Duperret, Ankit Dahal, Todd W. Ridky. Integrin αv is necessary for skin tissue generation and SCC tumor invasion, but dispensable for tissue maintenance. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2380. doi:10.1158/1538-7445.AM2015-2380