Transcriptional and post-translational regulation of beta 1 integrin expression during keratinocyte terminal differentiation.

During suspension-induced terminal differentiation of human epidermal keratinocytes, the alpha 5 beta 1 integrin is down-regulated in two stages: first, the ability of the receptor to bind fibronectin is reduced; later, the receptor is lost from the cell surface, and the level of the subunit mRNAs declines. We have begun to examine the mechanisms that regulate these events. Pulse-chase experiments showed that when keratinocytes were placed in suspension to induce terminal differentiation maturation of the beta 1 subunit and its associated alpha subunits was prevented. The inhibition of maturation was at the stage of N-linked glycosylation in the Golgi, because the immature integrin subunits were sensitive to endoglycosidase H digestion and the inhibition could be mimicked in adherent cells by treatment with 1-deoxymannojirimycin. In 1-deoxymannojirimycin-treated adherent keratinocytes, immature integrin subunits reached the cell surface; however, in keratinocytes induced to differentiate in suspension, no beta 1-integrin precursors were detected on the cell surface. Thus commitment to terminal differentiation results in a block both in integrin glycosylation and transport to the cell surface; down-regulation of receptor function must therefore involve modulation of pre-existing receptor on the cell surface. Although fibronectin or rabbit antiserum to alpha 5 beta 1 can inhibit suspension-induced terminal differentiation they did not overcome the inhibition of glycosylation. Nuclear run-on assays showed that transcription of the alpha 5 and beta 1 genes was switched off during suspension-induced terminal differentiation, and treatment of adherent keratinocytes with actinomycin D suggested that the half-lives of the alpha 5 and beta 1 mRNAs were similar in adherent and suspended cells. Thus, loss of alpha 5 beta 1 from the cell surface reflects both inhibition of transcription of the subunit genes and inhibition of maturation and intracellular transport of newly synthesized subunits.