Cell substrate adhesion-induced redistribution of proteins among the apical, basal, and internal domains of the plasma membrane of HeLa cells spreading on gelatin.

M.L Lu,B.S Jacobson,P.W Mason

doi:10.1016/s0021-9258(18)61418-8

Abstract

The establishment of the asymmetric distribution of proteins among the apical, basal, and internal plasma membrane (PM) domains was studied in HeLa cells. Comparisons were made of the amount of membrane and the redistribution of individual PM proteins in the three PM domains for cells on substrates that either induced cell attachment and spreading (gelatin), or induced only attachment (bovine serum albumin (BSA]. Many PM proteins were asymmetrically segregated among the apical, basal, and internal PM domains when cells were attached to gelatin. However, most of the proteins were not totally excluded from any of the domains. In contrast, there was no segregation of membrane components in cells attached to BSA. The segregation of most proteins was well established within 7.5 min of cell attachment when the cells were only partially spread. Cell adhesion induced a change in the movement of total membrane between the internal PM and the external PM domains. When cells attached to either gelatin or BSA there was a transient decrease in the internal PM pool that lasted less than 20 min. For cells attached to BSA the transient decrease was followed by the re-establishment of the internal PM pool which was equivalent to that found in cells in suspension, whereas in cells spreading on gelatin there was only a partial re-establishment of the internal PM pool. Taken together, these observations suggest that the internal PM rapidly moved to the external PM domain during cell adhesion, and that particular PM proteins moved from the internal PM pool into the newly established basal and apical PM domains.

Highlights

RESULTSD on geloiin iubstraie efflux of internal plasma membrane (PM) (Fig. 5) might represent membrane movement to thebasal PM followed byan influx of the apical PM to the internaPl M. i Fluorescence microscopy of the basal PM isolated from HeLa cells spreading on gelatin was used to estimate the size of the basal domain during spreading and identify membrane i
Comparisons were made of the amount of membrane peptide composition very similar to the external domain and and the redistributioonf individual plasma membrane (PM) proteins in theis in equilibrium with it (Fishman and Cook, 1982; Hubbard three PM domains for cells on substrates that either induced cell attachmentandspreading(gelatin), or induceodnly attachment (bovine serumalbumin (BSA))
For cells attached to BSA the tran- 1980; Oppenheimer-Marks and Grinnell, 1982; Brandts and sient decrease wafsollowed bythe re-establishment of Jacobson, 1983; Jacobson, 1983; Fairman and Jacobson1, 983; ottttfihhhonoeeeunnigsinednetxsteluteairengrtnrnigannacelalsePlttlPhMlPsteMhMraienptpowdtooshooluae.mlssTpiawnoeaitnnhnkeasilerciypdnnohaunatrrl,wotinigcPawageesMlhtlhrleeeerrq-aareeu,dpasihisttvdhaealbeisylnslieioensmnchtoe,tmobolvlasesteenhndrdtasvtpotaortf-ehaadttwhnIianaonhkgvtigebeccshelaerlaeopersctosclauhtahlror.s,awdni1in9sp18totrh6hliab)et.ruoaAittbiyclostohenimnosicupneeglsehftotuaeflbilpty(lcirisfoss.hptBekeriednunaodrdkwsueyrcnineenltttglhhsaealisc.tn,iesP1lclM9au8snlpt3dpur;reurMoeastd,uaeiaisnitlonlgynis,s particular PM proteins moved from the internal PM and Jacobson, 1985; Rodriguez-Boulan, 1983)

Summary

RESULTS

D on geloiin iubstraie efflux of internal PM (Fig. 5) might represent membrane movement to thebasal PM followed byan influx of the apical PM to the internaPl M. i Fluorescence microscopy of the basal PM isolated from HeLa cells spreading on gelatin was used to estimate the size of the basal domain during spreading and identify membrane i. Plasma membrane proteins (Geiger et al, 1982)and amethod for visualizing the basal PM which remains attached to the gelatin substrate after the apical and internal domains have beenremoved by the silica microbead membrane isolation procedure (Mason and Jacobson, 1985).Fig. 6 shows the LRBSC-labeled basal membranes isolated from cells at different stages of spreading.

HH B A I

HB A I

HBA I

Findings

DISCUSSION