RAC1 activation drives pathologic interactions between the epidermis and immune cells.

Interactions between the epidermis and the immune system govern epidermal tissue homeostasis. These epidermis-immune interactions are altered in the inflammatory disease psoriasis; however, the pathways that underlie this aberrant immune response are not well understood. Here, we determined that Ras-related C3 botulinum toxin substrate 1 (RAC1) is a key mediator of epidermal dysfunction. RAC1 activation was consistently elevated in psoriatic epidermis and primary psoriatic human keratinocytes (PHKCs) exposed to psoriasis-related stimuli, but not in skin from patients with basal or squamous cell carcinoma. Expression of a constitutively active form of RAC1 (RACV12) in mice resulted in the development of lesions similar to those of human psoriasis that required the presence of an intact immune system. RAC1V12-expressing mice and human psoriatic skin showed similar RAC1-dependent signaling as well as transcriptional overlap of differentially expressed epidermal and immune pathways. Coculture of PHKCs with immunocytes resulted in the upregulation of RAC1-dependent proinflammatory cytokines, an effect that was reproduced by overexpressing RAC1 in normal human keratinocytes. In keratinocytes, modulating RAC1 activity altered differentiation, proliferation, and inflammatory pathways, including STAT3, NFκB, and zinc finger protein 750 (ZNF750). Finally, RAC1 inhibition in xenografts composed of human PHKCs and immunocytes abolished psoriasiform hyperplasia and inflammation in vivo. These studies implicate RAC1 as a potential therapeutic target for psoriasis and as a key orchestrator of pathologic epidermis-immune interactions.

A laminin-collagen complex drives human epidermal carcinogenesis through phosphoinositol-3-kinase activation.

Laminin-332 (formerly laminin-5) and collagen VII are basement membrane proteins expressed at the invasive front of human squamous cell carcinoma (SCC) tumors. These proteins have protumorigenic properties, but whether laminin-332 and collagen VII promote SCC tumors by providing adhesion or other nonadhesive extracellular cues, or whether laminin-332 and collagen VII interact together in this process remains unknown. In this study, we examined the role of these molecules by a structural approach using an in vivo model of human SCC tumorigenesis. Here, we show that individual domains (VI and V-III) on the laminin-332 beta3 chain provide distinct and highly divergent cell adhesion and tumor-promoting functions. We found that laminin beta3 domain VI provided a critical role in the assembly of stable adhesion complexes, but this domain was not required in SCC tumors. Instead, we found that laminin beta3 domain V-III played an essential role in SCC carcinogenesis/invasion through binding to collagen VII, which in turn, led to phosphoinositol-3-kinase activation and protection from apoptosis. Overexpression of constitutively active p110 phosphoinositol-3-kinase subunit was sufficient to restore invasion and tumorigenesis in transformed cells lacking laminin-332/collagen VII interaction in a manner independent of cellular adhesion. These studies show distinctive adhesive and signaling functions in individual domains of laminin-332, one which is required for normal epithelial adhesion and one which is required for SCC tumorigenesis. This uncoupling of stable adhesion from tumor progression in our studies suggests that laminin-332/collagen VII interaction promotes epidermal carcinogenesis through signaling rather than adhesion.

Alpha 6 beta 4 integrin regulates keratinocyte chemotaxis through differential GTPase activation and antagonism of alpha 3 beta 1 integrin.

Growth factor-induced cell migration and proliferation are essential for epithelial wound repair. Cell migration during wound repair also depends upon expression of laminin-5, a ligand for alpha 6 beta 4 integrin. We investigated the role of alpha 6 beta 4 integrin in laminin-5-dependent keratinocyte migration by re-expressing normal or attachment-defective beta 4 integrin in beta 4 integrin null keratinocytes. We found that expression of beta 4 integrin in either a ligand bound or ligand unbound state was necessary and sufficient for EGF-induced cell migration. In a ligand bound state, beta 4 integrin supported EGF-induced cell migration though sustained activation of Rac1. In the absence of alpha 6 beta 4 integrin ligation, Rac1 activation became tempered and EGF chemotaxis proceeded through an alternate mechanism that depended upon alpha 3 beta 1 integrin and was characterized by cell scattering. alpha 3 beta 1 integrin also relocalated from cell-cell contacts to sites of basal clustering where it displayed increased conformational activation. The aberrant distribution and activation of alpha 3 beta 1 integrin in attachment-defective beta 4 cells could be reversed by the activation of Rac1. Conversely, in WT beta 4 cells the normal cell-cell localization of alpha 3 beta 1 integrin became aberrant after the inhibition of Rac1. These studies indicate that the extracellular domain of beta 4 integrin, through its ability to bind ligand, functions to integrate the divergent effects of growth factors on the cytoskeleton and adhesion receptors so that coordinated keratinocyte migration can be achieved.