Subcellular localization of myosin-V in the B16 melanoma cells, a wild-type cell line for the dilute gene.
The discovery that the dilute gene encodes a class V myosin led to the hypothesis that this molecular motor is involved in melanosome transport and/or dendrite outgrowth in mammalian melanocytes. The present studies were undertaken to gain insight into the subcellular distribution of myosin-V in the melanoma cell line B16-F10, which is wild-type for the dilute gene. Immunofluorescence studies showed some degree of superimposed labeling of myosin-V with melanosomes that predominated at the cell periphery. A subcellular fraction highly enriched in melanosomes was also enriched in myosin-V based on Western blot analysis. Immunoelectron microscopy showed myosin-V labeling associated with melanosomes and other organelles. The stimulation of B16 cells with the alpha-melanocyte-stimulating hormone led to a significant increase in myosin-V expression. This is the first evidence that a cAMP signaling pathway might regulate the dilute gene expression. Immunofluorescence also showed an intense labeling of myosin-V independent of melanosomes that was observed within the dendrites and at the perinuclear region. Although the results presented herein are consistent with the hypothesis that myosin-V might act as a motor for melanosome translocation, they also suggest a broader cytoplasmic function for myosin-V, acting on other types of organelles or in cytoskeletal dynamics.
Nascimento
A A
AA
Department of Morphology, Faculdade de Medicina de Ribeirão Preto-Universidade de São Paulo, Brazil.
Amaral
R G
RG
Bizario
J C
JC
Larson
R E
RE
Espreafico
E M
EM
D000818:Animals; D015153:Blotting, Western; D002458:Cell Fractionation; D052138:Genes, Neoplasm; D007150:Immunohistochemistry; D009074:Melanocyte-Stimulating Hormones / Q000494:pharmacology; D008546:Melanoma, Experimental / Q000235:genetics / Q000473:pathology*; D051379:Mice; D008810:Mice, Inbred C57BL; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D009218:Myosins / Q000032:analysis* / Q000187:drug effects / Q000235:genetics*; D014407:Tumor Cells, Cultured / Q000737:chemistry / Q000187:drug effects / Q000648:ultrastructure
D009074:Melanocyte-Stimulating Hormones; D009218:Myosins