The role of extracellular materials in cell movement. I. Inhibition of mucopolysaccharide synthesis does not stop ruffling membrane activity or cell movement.

Abstract

The involvement of mucopolysaccharide synthesis in cell locomotion was investigated by determining the effects of inhibition of synthesis on ruffling membrane activity and cell movement by embryonic heart fibroblasts. Mucopolysaccharide synthesis was inhibited directly by treatment with a glutamine analog, 6-diazo-5-oxo-L-norleucine (DON), and indirectly with cycloheximide. DON treatment reduced synthesis to 20% of control values, and cycloheximide reduced synthesis to less than 10% of control values, as measured by incorporation of [86S]sulfate into mucopolysaccharides. Nevertheless, ruffling membrane activity and cell locomotion continued under both conditions. Cytochalasin B did not inhibit mucopolysaccharide synthesis, although it did stop ruffling and locomotion. These results suggest that if mucopolysaccharides are required for cell movement, they must have long half-lives or represent only a minute fraction of the normal synthetic load.In recent years, use of refined techniques has provided insight into the mechanism of vertebrate cell locomotion in vitro. Time-lapse cinematography (lngram, 1969; Abercrombie et ai., 1970;Harris, 1973), as well as light and electron microscopy (Buckley and Porter, 1967;Spooner et al., 1971;Goldman, 1972), has revealed several behavioral characteristics of migrating cells and implicated specific subcellular structures in generating the motive forces for movement. Understanding cell movement has been facilitated by recognizing that effective translocation results from three successive events. First, the cell extends its leading edge forward into space above the substratum. Second, some portion of the extended leading edge is lowered and adheres to the substratum with a net adhesive force greater than that of more posterior points of adhesion. Third, a cytoplasmic contraction (or an elastic recoil) takes place between the anterior area of adhesion and more posterior ones, resulting in the cell moving forward as posterior adhesions are broken. This locomotory cycle is repeated as the cell continues to translocate.A number of investigators are beginning to gain insight into the extension step of locomotion (Ingram, 1969; Abercromhie et al., 1970;Harris, 1973;, and rapidly accumulating evidence implicates actin and myosin in the contractile phase of the locomotory cycle (Bray, 1973;

Relationship of transformation, cell density, and growth control to the cellular distribution of newly synthesized glycosaminoglycan.

“…The behavioral differences evident at low density probably do not involve differences in cell surface GAG because, although there is a small difference in amount, the types and relative proportions of labeled surface GAG on low density 3T3 and 3T3SV cells are very similar. This conclusion is consistent with the report that cell locomotion is not modified by inhibition of GAG synthesis (44).…”

A requirement for trypsin-sensitive cell-surface components for cell-cell interactions of embryonic neural retina cells

“…A similar pattern of inhibition was found with DON (6-diazo-5-oxo-L-norleucine), a glutamine analog that is thought to inhibit mucopolysaccharide synthesis (37,39). Recovery was partially affected by DON (Table I), and collection of recovered cells was unaffected by DON at a concentration that inhibited incorporation of [asS]sulfate into mucopolysaccharides by about 70% (37). The inhibition pattern of DON was very similar to that of cycloheximide: both inhibitors affected the recovery of cells from trypsin treatment but did not completely inhibit recovery.…”

“…If ceils were given a chance to recover before cycloheximide treatment (Table II), the cycloheximide had little or no effect on the rate of collection for 2 h. We have previously shown that long treatments with cycloheximide eventually inhibit collection of recovered cells (18). A similar pattern of inhibition was found with DON (6-diazo-5-oxo-L-norleucine), a glutamine analog that is thought to inhibit mucopolysaccharide synthesis (37,39). Recovery was partially affected by DON (Table I), and collection of recovered cells was unaffected by DON at a concentration that inhibited incorporation of [asS]sulfate into mucopolysaccharides by about 70% (37).…”

Post-translational alterations in newly synthesized cartilage proteoglycans induced by the glutamine analogue 6-diazo-5-oxo-l-norleucine. Time course of inhibition and recovery

“…A similar time-dependent inhibition of [35S]sulphate incorporation by DON has been reported for cultures of chick heart fibroblasts (Spooner & Conrad, 1975).…”