Journal: bioRxiv

Article Title: The deacylase SIRT5 supports melanoma viability by regulating chromatin dynamics

doi: 10.1101/2020.09.07.286526

Figure Lengend Snippet: A. Immunoblot analysis demonstrating induction of caspase 3 cleavage 72 and 96 hrs. post-transduction with shRNAs targeting SIRT5 (KD1 or KD2) in A2058 and SK-MEL-2 cell lines. B . Viability of MP-41, A2058 or YUMM5.2 cells infected with control (C) or one of five SIRT5 shRNAs (KD1-KD5) against human SIRT5 (top and middle panels) or mouse Sirt5 (bottom panel). Average results (n=6/timepoint) are graphed. Error bars represent standard deviation. Right panels: immunoblot analysis demonstrating loss of SIRT5 and induction of caspase 3 cleavage following SIRT5 KD. C. Flow cytometric analysis of A2058 cells stained with Annexin V and propidium iodide (PI), as indicated, showing an increased fraction of Annexin V-positive cells 96 hrs. after SIRT5 KD. D. Average of n=3 technical replicates is plotted. Error bars represent (SD). Significance calculated using unpaired Student’s t-test. Increased Annexin V + staining is observed in both the PI-positive and PI-negative populations.

Article Snippet: MP-38, MP-41 and MP-46 cells were cultured in RPMI (Gibco) containing 2 mM L-glutamine, 10 mM HEPES, 110mg/L sodium pyruvate, 4.5g/L glucose, 100units/mL penicillin, 100µg/mL streptomycin and 25% heat-inactivated FBS; 92-1 cells were cultured in RPMI (Gibco) containing 2 mM L-glutamine, 10 mM HEPES, 110mg/L sodium pyruvate, 4.5g/L glucose, 100units/mL penicillin, 100µg/mL streptomycin and 10% heat-inactivated FBS.

Techniques: Western Blot, Transduction, Infection, Standard Deviation, Staining

Journal: bioRxiv

Article Title: The deacylase SIRT5 supports melanoma viability by regulating chromatin dynamics

doi: 10.1101/2020.09.07.286526

Figure Lengend Snippet: A. Immunoblot analysis demonstrating induction of caspase 3 cleavage 72 and 96 hrs. post-transduction with shRNAs targeting SIRT5 (KD1 or KD2) in A2058 and SK-MEL-2 cell lines. B . Viability of MP-41, A2058 or YUMM5.2 cells infected with control (C) or one of five SIRT5 shRNAs (KD1-KD5) against human SIRT5 (top and middle panels) or mouse Sirt5 (bottom panel). Average results (n=6/timepoint) are graphed. Error bars represent standard deviation. Right panels: immunoblot analysis demonstrating loss of SIRT5 and induction of caspase 3 cleavage following SIRT5 KD. C. Flow cytometric analysis of A2058 cells stained with Annexin V and propidium iodide (PI), as indicated, showing an increased fraction of Annexin V-positive cells 96 hrs. after SIRT5 KD. D. Average of n=3 technical replicates is plotted. Error bars represent (SD). Significance calculated using unpaired Student’s t-test. Increased Annexin V + staining is observed in both the PI-positive and PI-negative populations.

Article Snippet: Uveal melanoma cell lines MP-38, MP-41 and MP-46 were purchased from ATCC; cell line 92-1 was purchased from Sigma-Aldrich (Table S2).

Techniques: Western Blot, Transduction, Infection, Standard Deviation, Staining