CELL-SUBSTRATE INTERACTIONS AND LOCOMOTION OF DICTYOSTELIUM WILD-TYPEAND MUTANTS DEFECTIVE IN 3 CYTOSKELETAL PROTEINS - A STUDY USING QUANTITATIVE REFLECTION INTERFERENCE CONTRAST MICROSCOPY

Reflection interference contrast microscopy combined with digital imag e processing was applied to study the motion of Dictyostelium discoide um cells in their pre-aggregative state on substrata of different adhe siveness (glass, albumin-covered glass, and freshly cleaved mica). The temporal variations of the size and shape of the cell/substratum cont act area and the time course of advancement of pseudopods protruding i n contact with the substratum were analyzed. The major goal was to stu dy differences between the locomotion of wild-type cells and strains o f triple mutants deficient in two F-actin crosslinking proteins (a-act inin and the 120-kDa gelation factor) and one F-actin fragmenting prot ein (severin). The size of contact area, A,, of both wild-type and mut ant cells fluctuates between minimum and maximum values on the order o f minutes, pointing toward an intrinsic switching mechanism associated with the mechanochemical control system. The fluctuation amplitudes a re much larger on freshly cleaved mica than on glass. Wild-type and mu tant cells exhibit remarkable differences on mica but not on glass. Th ese differences comprise the population median of A(C) and alterations in pseudopod protrusion. A(C) is smaller by a factor of two or more f or all mutants. Pseudopods protrude slower and shorter in the mutants. It is concluded that cell shape and pseudopods are destabilized by de fects in the actin-skeleton, which can be overcompensated by strongly adhesive substrata. Several features of amoeboid cell locomotion on su bstrata can be understood on the basis of the minimum bending energy c oncept of soft adhering shells and by assuming that adhesion induces l ocal alterations of the composite membrane consisting of the protein/l ipid bilayer on the cell surface and the underlying actin-cortex.