Function and Differential Regulation of the α6 Integrin Isoforms during Parietal Endoderm Differentiation

• Published in: Experimental cell research 1995-04, Vol.217 (2), p.195-204
• Main Authors: Jiang, Rulang, Grabel, Laura B.
• Format: Article
• Language: English
• ISSN: 0014-4827; 1090-2422
• DOI: 10.1006/excr.1995.1079

F9 embryonal carcinoma cells treated with retinoic acid differentiate in monolayer into parietal endoderm (PE) or in suspension into embryoid bodies with an outer layer of visceral endoderm (VE) surrounding a core of largely undifferentiated cells. Previous reports have shown that cell-extracellular matrix interactions mediated by the β1 integrins play a critical role in the differentiation and migration of PE. In the present study we investigated the pattern of expression and function of the integrin α6β1 during the differentiation of F9 cells into VE and PE. F9 cells express integrin subunits α3, α6, α6, and β1. Cell adhesion and migration assays demonstrate that α6β1 is the major laminin receptor in undifferentiated F9 cells as well as F9-derived PE cells. However, the amount of α6 protein decreases significantly upon F9 cell differentiation into either VE or PE, as revealed by immunofluorescent staining and immunoprecipitation analysis. In contrast, the amount of steady-state α6 message stays constant before and after F9 cell differentiation, suggesting that the down-regulation of α6β1 occurs post-transcriptionally. In view of previous reports of two α6 isoforms generated by alternative RNA processing, we carried out reverse transcription-PCR analysis and show that, while α6B is the major mRNA isoform before and after F9 cell differentiation, α6A mRNA is weakly expressed in undifferentiated F9 cells and is substantially increased following F9 differentiation into PE. Immunoprecipitations using the isoform-specific antibodies show an increase in α6A and a dramatic decrease in α6B protein following PE differentiation. Pulse-chase experiments indicate that, whereas the stability of α6B protein is unaltered, synthesis of α6B protein is decreased at least threefold following PE differentiation. Further experiments demonstrate that α6A localizes to focal contacts in PE cells. The switch from α6B to α6A and the localization of α6A at focal contacts correlate with the acquisition of PE cell motility, which suggests distinct functions for the two α6 isoforms.