CaM kinase II selectively signals to histone deacetylase 4 during cardiomyocyte hypertrophy
J. Clin. Invest. Johannes Backs, et al. 116:1853 doi:10.1172/JCI27438 [
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Figure 3Detection of 14-3-3 binding sites of HDAC4 in response to CaMKI and CaMKII. (
A and
B) Coimmunoprecipitation assays with COS cell lysates were analyzed with an antibody directed against endogenous 14-3-3 protein. The effects of CaMKI and CaMKII on various FLAG-HDAC4 mutants were tested. HDAC4-input, HDAC4 present in the COS cell lysate before IP was performed; CaMK-input, CaMKI or CaMKIIδB-T287D present in the COS cell lysate before IP was performed. (
B) Fold-increase in 14-3-3 binding in response to CaMKII or CaMKI as compared with baseline. (
C) The N terminal half of HDAC4 (amino acids 1–740) was fused to the GAL4 DNA-binding domain, and 14-3-3 was fused to the VP16 transcription activation domain. If GAL4-HDAC4 is not phosphorylated, it cannot recruit VP16–14-3-3 and cannot activate the GAL4-dependent luciferase reporter. (
D) As indicated, different GAL4-HDAC constructs were used in this assay in the absence and presence of CaMKIIδB-T287D. COS cells were transfected with the indicated constructs. Increase in 14-3-3 binding is expressed as compared with control conditions without kinase. (
E) CaMKI and CaMKII phosphorylation sites of HDAC4 are shown. ND, not detectable; NES, nuclear export signal.
