Figure 7.
Loss of Lysmd1/2 causes abnormal enlargement of melanosomes and reduces melanin production in mouse B16F10 cells. (A) Western blotting of LYSMD1, LYSMD2, Rab32, and Tyrosinase in B16F10 cells with shRNA knockdown of Lysmd1 (shD1), Lysmd1 (shD2), or both (shD1/D2). (B) Representative images (left) and quantification (right) of melanosomes stained with tyrosinase antibody in B16F10 cells with the indicated shRNA treatment. The boxed region is magnified (3×) at the bottom right in each image. Scale bars, 5 μm. Diameters of 80 melanosomes were measured in each group (mean ± SEM, n = 80 melanosomes, unpaired two-tailed t test, ***P < 0.001, ns: not significant). (C) Representative images (left) and quantification (right) of melanin production in shRNA-treated B16F10 cells with or without UV irradiation. Melanin production was quantified by calculating A405/A562 (mean ± SEM, n = 4 independent experiments, unpaired two-tailed t test, **P < 0.01, ***P < 0.001, ns: not significant). (D) Reinforced expression of LYSMD1, LYSMD2, or both rescues melanosome enlargement in B16F10 cells treated with shD1/D2. Representative images are shown on the left and quantification of melanosome diameters is shown on the right. The boxed region is magnified (3×) at the bottom right in each image. Scale bars, 5 μm. Diameters of 80 melanosomes were measured in each group (mean ± SEM, n = 80 melanosomes, unpaired two-tailed t test, ***P < 0.001, ns: not significant). (E) Reinforced expression of LYSMD1, LYSMD2, or both rescues defective melanin production in B16F10 cells treated with shD1/D2. Representative images (left) and quantification of melanin production (right) are shown. Melanin production was quantified by calculating A405/A562 (mean ± SEM, n = 4 independent experiments, unpaired two-tailed t test, **P < 0.01, ***P < 0.001). OE, overexpressing. Source data are available for this figure: SourceData F7.

Loss of Lysmd1/2 causes abnormal enlargement of melanosomes and reduces melanin production in mouse B16F10 cells. (A) Western blotting of LYSMD1, LYSMD2, Rab32, and Tyrosinase in B16F10 cells with shRNA knockdown of Lysmd1 (shD1), Lysmd1 (shD2), or both (shD1/D2). (B) Representative images (left) and quantification (right) of melanosomes stained with tyrosinase antibody in B16F10 cells with the indicated shRNA treatment. The boxed region is magnified (3×) at the bottom right in each image. Scale bars, 5 μm. Diameters of 80 melanosomes were measured in each group (mean ± SEM, n = 80 melanosomes, unpaired two-tailed t test, ***P < 0.001, ns: not significant). (C) Representative images (left) and quantification (right) of melanin production in shRNA-treated B16F10 cells with or without UV irradiation. Melanin production was quantified by calculating A405/A562 (mean ± SEM, n = 4 independent experiments, unpaired two-tailed t test, **P < 0.01, ***P < 0.001, ns: not significant). (D) Reinforced expression of LYSMD1, LYSMD2, or both rescues melanosome enlargement in B16F10 cells treated with shD1/D2. Representative images are shown on the left and quantification of melanosome diameters is shown on the right. The boxed region is magnified (3×) at the bottom right in each image. Scale bars, 5 μm. Diameters of 80 melanosomes were measured in each group (mean ± SEM, n = 80 melanosomes, unpaired two-tailed t test, ***P < 0.001, ns: not significant). (E) Reinforced expression of LYSMD1, LYSMD2, or both rescues defective melanin production in B16F10 cells treated with shD1/D2. Representative images (left) and quantification of melanin production (right) are shown. Melanin production was quantified by calculating A405/A562 (mean ± SEM, n = 4 independent experiments, unpaired two-tailed t test, **P < 0.01, ***P < 0.001). OE, overexpressing. Source data are available for this figure: SourceData F7.

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