Fig. 9. MARCKS is essential for muscle cell spreading. 5-expressing cells were transfected with a MARCKS antisense vector or a vector control, and individual clones are selected for analysis of MARCKS expression and cell spreading. (A) MARCKS expression in transfected clones. Three clones (02, 042 and 011) transfected with the MARCKS-antisense cDNA showed reduced levels of MARCKS protein. A representative clone (`cont') transfected with control vector showed normal levels of MARCKS protein expression. Individual clones were photographed to illustrate the relationship between MARCKS protein expression and cell spreading. The clone transfected with the control vector (empty vector) showed normal cell spreading on FN. By contrast, each of the clones expressing the MARCKS antisense vector showed reduced cell spreading, and the inhibition of cell spreading correlated directly with the extent of reduction of MARCKS protein expression (A). MARCKS protein was undetectable by western blot analysis in clone 02, and this clone showed the most dramatic inhibition of cell spreading. Clone 02 was plated on FN with or without pretreatment with PMA (100 nM). Even activation of PKC by PMA was unable to promote spreading of this clone in which MARCKS protein was undetectable (A).