LOCALIZATION OF MYOSIN-II A-ISOFORMS AND B-ISOFORMS IN CULTURED NEURONS

Tension generated by growth cones regulates both the rate and the dire ction of neurite growth, The most likely effecters of tension generati on are actin and myosins, We are investigating the role of conventiona l myosin in growth cone advance, In this paper we report the localizat ion of the two most prominent isoforms of brain myosin II in growth co nes, neurites and cell bodies of rat superior cervical ganglion neuron s, Affinity purified polyclonal antibodies were prepared against uniqu e peptide sequences from human and rat A and B isoforms of myosin heav y chain. Although each of these antibodies brightly stained nonneurona l cells, antibodies to myosin heavy chain B stained neurons with great er intensity than antibodies to myosin heavy chain A, In growth cones, myosin heavy chain B was most concentrated in the margin bordering th e thickened, organelle-rich central region and the thin, actin-rich pe ripheral region, The staining colocalized with actin bundles proximal and distal to the marginal zone, though the staining was more prominen t proximally. The trailing edge of growth cones and the distal portion of the neurite often had a rimmed appearance, but more proximal regio ns of neurites had cytoplasmic labelling. Localizing MHC-B in growth c ones previously monitored during advance (using differential interfere nce contrast microscopy) revealed a positive correlation with edges at which retraction had just occurred and a negative correlation with la mellipodia that had recently undergone protrusion, Cell bodies were br ightly labelled for myosin heavy chain B, Myosin heavy chain A stainin g was dimmer and its colocalization with filamentous actin bundles in growth cones was less striking than that of myosin heavy chain B, Grow th cones stained for both myosin heavy chain A and B revealed that the two antigens overlapped frequently, but not exclusively, and that myo sin heavy chain A lacked the elevation in the marginal zone that was c haracteristic of myosin heavy chain B, The pattern of staining we obse rved is consistent with a prominent role for myosin heavy chain B in e ither generating tension between widely separated areas of the growth cone, or bundling of actin filaments, which would enable other motors to effect this tension, These data support the notion that conventiona l myosin is important in growth cone advance and turning.