Membrane-associated actin from the microvillar membranes of ascites tumor cells.
A membrane fraction (MF2) has been purified from isolated microvilli of the MAT-C1 subline of the 13762 rat mammary ascites adenocarcinoma under conditions which cause F-actin depolymerization. This membrane preparation contains actin as a major component, although no filamentous structures are observed by transmission electron microscopy. Membranes were extracted with a Triton X-100-containing actin-stabilizing buffer (S buffer) or actin-destabilizing buffer (D buffer). In D buffer greater than 90% of metabolically labeled protein and glycoprotein was extracted, and 80-90% of these labeled species was extracted in S buffer. When S buffer extracts of MF2 were fractionated by either gel filtration on Sepharose 6 B or rate-zonal sucrose density gradient centrifugation, most of the actin was found to be intermediate in size between G- and F-actin. In D buffer most of the MF2 actin behaved as G-actin. Extraction and gel filtration of intact microvilli in S buffer also showed the presence of the intermediate form of actin, indicating that it did not arise during membrane preparation. When [35S]methionine-labeled G-actin from ascites cells was added to S buffer extracts of MF2 and chromatographed, all of the radioactivity chromatographed as G-actin, indicating that the intermediate form of actin did not result from an association of G-actin molecules during extraction or chromatography. The results of this study suggest that the microvillar membrane fraction is enriched in an intermediate form of actin smaller than F-actin and larger than G-actin.
D000199:Actins / Q000032:analysis*; D000818:Animals; D001201:Ascites; D002352:Carrier Proteins / Q000032:analysis; D002462:Cell Membrane / Q000648:ultrastructure*; D003599:Cytoskeleton / Q000648:ultrastructure*; D018260:Gelsolin; D008840:Microfilament Proteins; D008871:Microvilli / Q000648:ultrastructure*; D008970:Molecular Weight; D009218:Myosins / Q000378:metabolism; D009374:Neoplasms, Experimental / Q000648:ultrastructure; D011485:Protein Binding; D051381:Rats
D000199:Actins; D002352:Carrier Proteins; D018260:Gelsolin; D008840:Microfilament Proteins; C016953:brevin; D009218:Myosins