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. Author manuscript; available in PMC: 2020 May 15.
Published in final edited form as: Nat Metab. 2019 Nov 15;1(11):1157–1167. doi: 10.1038/s42255-019-0138-4

Extended Data Fig. 1. ABHD5 is required for HDAC4 proteolysis.

Extended Data Fig. 1

(a) Immunoblotting using an antibody directed against endogenous ABHD5 in cardiac and cellular COS as well as HEK293 extracts. GAPDH was used as loading control. (b) Cytosolic and nuclear fractions from adult rat ventricular myocytes were used for immunoblotting using an antibody directed against ABHD5, GAPDH and histone H3. (a-b) Representative results of two independent experiments. (c) Left panel: Immunoblotting analysis of cardiac extracts of control (Ctrl) or isoproterenol (ISO) treated mice using antibodies directed against HDAC4, ABHD5 and GAPDH as loading control. Middle left panel: Densitometric analysis of HDAC4-NT/FL ratio shown as fold change vs Ctrl. Middle-right panel and right panel: Quantification of cardiac ABHD5 protein normalized to GAPDH and abhd5-RNA levels normalized to 18S in unstressed mice and ISO-treated mice. Statistical analysis: Values are presented as Mean±SEM, n=3 (n represents mice per indicated group); by unpaired two tailed t-test, P<0.05 considered as significant. (d) Left panel: Immunoblotting analysis of cardiac extracts of control (Ctrl) or CL 316,243 (CL) treated mice using antibodies directed against HDAC4, ABHD5 and GAPDH as loading control. Middle-left panel: Densitometric analysis of HDAC4-NT/FL ratio shown as fold change vs Ctrl. Middle panel: Quantification of cardiac ABHD5 protein normalized to GAPDH and abhd5-RNA levels normalized to 18S in unstressed mice and CL-treated mice. Right panel: Quantification of ucp1 mRNA normalized to 18S in brown adipose tissue after treatment with CL as a marker of CL effective administration. Statistical analysis: Values are presented as Mean±SEM, n=3 (n represents mice per indicated group); by unpaired two tailed t-test, P<0.05 considered as significant.