Myosin II antibodies inhibit the resorption activity of isolated rat osteoclasts.
We present microinjection data in support of an indirect approach by which cytoplasmic protein interactions important in the processes of bone resorption can be elucidated. Three polyclonal antibodies (M1, M3, M5) raised against myosin II from perfused rat liver differently affected the actin-activated Mg ATPase of myosin II. These antibodies microinjected into isolated rat osteoclasts affected osteoclast morphology and activity in bone resorption. M1, which completely inhibited myosin ATPase activity at a antibody:myosin ratio of 10:1, initially promoted the extension/retraction motility of lamellipodia but eventually reduced the spread area of osteoclasts along the substrate after 20 hr. M3, which inhibited ATPase activity by 70%, had similar effects; however, M5, which weakly inhibited ATPase activity, neither promoted extension/retraction nor reduced spread area of osteoclasts. Immunofluorescence showed that these antibodies removed myosin II from the majority of actin filaments in injected osteoclasts. Because antibodies that did not bind to a myosin II column had little effect on the extension/retraction of lamellipodia or the osteoclast spread area, these data suggest that myosin II participates in the stabilization of osteoclast lamellipodia along the substrate. M1 injection strongly inhibited injected osteoclasts from excavating resorption lacunae in bone slices, compared to control antibody. M3 and M5 were less effective but also inhibited bone resorption. These data show that myosin II is functionally important in bone resorption and that the osteoclast-differentiated activity of bone resorption is a more sensitive assay for myosin activity than lamellipodia motility or cell morphology.
Sato
M
M
Department of Bone Biology and Osteoporosis Research, Merck Sharp and Dohme Research Laboratories, West Point, Pennsylvania 19486.
Grasser
W
W
D000818:Animals; D000906:Antibodies; D001862:Bone Resorption / Q000378:metabolism*; D008845:Microinjections; D009218:Myosins / Q000502:physiology*; D010010:Osteoclasts / Q000166:cytology / Q000502:physiology*; D051381:Rats; D011919:Rats, Inbred Strains
D000906:Antibodies; D009218:Myosins