Epithelial-mesenchymal transition downregulates laminin α5 chain and upregulates laminin α4 chain in oral squamous carcinoma cells

Abstract

Basement membranes maintain the epithelial phenotype and prevent invasion and metastasis. We hypothesized that expression of basement membrane laminins might be regulated by epithelial-mesenchymal transition (EMT), hallmark of cancer progression. As EMT is mediated by transcription factor Snail, we used oral squamous carcinoma cells obtained from a primary tumor (43A), from its EMT-experienced recurrence (43B) and Snail-transfected 43A cells (43A-SNA) displaying full EMT, as a model to study laminins and their receptors. Northern blotting, immunofluorescence, and immunoprecipitation showed a gradual loss of laminin-511 and its receptor Lutheran from 43A to 43B and 43A-SNA cells. In contrast, neoexpression of laminin alpha4 mRNA was found congruent with synthesis of laminin-411. Chromatin immunoprecipitation disclosed direct binding of Snail to regions upstream of laminin alpha5 and alpha4 genes. Immunofluorescence and immunoprecipitation showed a switch from hemidesmosomal integrin alpha(6)beta(4) to alpha(6)beta(1) and neoexpression of alpha(1)beta(1) in 43A-SNA cells, and upregulation of integrin-linked kinase in both 43B and 43A-SNA cells. The cells adhered potently to laminin-511 and fibronectin, whereas adhesion to laminin-411 was minimal. In contrast, laminin-411 inhibited cell adhesion to other extracellular matrix proteins. In conclusion, EMT induces a switch from laminin-511 to laminin-411 expression, which may be directly controlled by Snail. Concomitant changes take place in laminin- and collagen-binding receptors. Laminin-411 reduces adhesion to laminin-511 and fibronectin, suggesting that tumor cells could utilize laminin-411 in their invasive behavior.

The role of the transcriptional regulator snail in cell detachment, reattachment and migration

“…In addition to laminin-332, laminin-α5 is also downregulated in snail transfected SCC cells. 13 In contrast, in the same cells, neoexpression of laminin-α4 mRNA and synthesis of laminin-411 (laminin-8) was observed. Laminin-511 is regarded as the most widely expressed laminin and is found in most epithelial BMs.…”

“…15,16 SCC cells potently adhered to laminin-511 whereas adhesion to laminin-411 was minimal. 13 Furthermore, the laminin-α4 chain has been reported to play a role in the detachment of renal carcinoma cells from fibronectin. 17 Laminin-411 decreased adhesion of SCC cells to laminin-511 and to fibronectin by blocking binding sites in the fibronectin molecule.…”

“…17 Laminin-411 decreased adhesion of SCC cells to laminin-511 and to fibronectin by blocking binding sites in the fibronectin molecule. 13 In parental SCC cells integrin α6 is paired with the β4 subunit. When paired with the β4 subunit, integrin α6β4 mediates the formation of hemidesmosomes which link the intermediate filament cytoskeleton to BM laminin-332.…”

“…Integrin α6β4 also mediates cell adhesion to laminin-511. 13 However, in 43A-SNA cells integrin α6 is paired with the β1 subunit. 13 Integrin α6β1 is the main receptor for laminin-411.…”

“…13 However, in 43A-SNA cells integrin α6 is paired with the β1 subunit. 13 Integrin α6β1 is the main receptor for laminin-411. A reduction in integrin α6β4 levels has been shown in snail transfected SCC cells.…”