Caffeine induces hyperacetylation of histones at the MEF2 site on theGlut4promoter and increases MEF2A binding to the site via a CaMK-dependent mechanism

Abstract

Mukwevho E, Kohn TA, Lang D, Nyatia E, Smith J, Ojuka EO. Caffeine induces hyperacetylation of histones at the MEF2 site on the Glut4 promoter and increases MEF2A binding to the site via a CaMK-dependent mechanism. Am J Physiol Endocrinol Metab 294: E582-E588, 2008. First published January 15, 2008 doi:10.1152/ajpendo.00312.2007.-This study was conducted to explore the mechanism by which caffeine increases GLUT4 expression in C2C12 myotubes. Myoblasts were differentiated in DMEM containing 2% horse serum for 13 days and the resultant myotubes exposed to 10 mM caffeine in the presence or absence of 25 M KN93 or 10 mM dantrolene for 2 h. After the treatment, cells were kept in serum-free medium and harvested between 0 and 6 h later, depending on the assay. Chromatin immunoprecipitation (ChIP) assays revealed that caffeine treatment caused hyperacetylation of histone H3 at the myocyte enhancer factor 2 (MEF2) site on the Glut4 promoter (P Ͻ 0.05) and increased the amount of MEF2A that was bound to this site ϳ2.2-fold (P Ͻ 0.05) 4 h posttreatment compared with controls. These increases were accompanied by an ϳ1.8-fold rise (P Ͻ 0.05 vs. control) in GLUT4 mRNA content at 6 h post-caffeine treatment. Both immunoblot and immunocytochemical analyses showed reduced nuclear content of histone deacetylase-5 in caffeinetreated myotubes compared with controls at 0 -2 h posttreatment. Inclusion of 10 mM dantrolene in the medium to prevent the increase in cytosolic Ca 2ϩ , or 25 M KN93 to inhibit Ca 2ϩ /calmodulindependent protein kinase (CaMK II), attenuated all the above caffeine-induced changes. These data indicate that caffeine increases GLUT4 expression by acetylating the MEF2 site to increase MEF2A binding via a mechanism that involves CaMK II. myocyte enhancer factor 2; glucose transporter 4; histone deacetylase; chromatin immunoprecipitation assay; Ca 2ϩ /calmodulin-dependent protein kinase II; histone H3 REGULAR EXERCISE INCREASES THE CONTENT of the GLUT4 glucose transporter that increases glucose disposal and protects against type II diabetes (8, 28). There is mounting evidence that the increase in cytosolic Ca 2ϩ

Chronic caffeine intake reverses age-induced insulin resistance in the rat: effect on skeletal muscle Glut4 transporters and AMPK activity

“…Our results suggest that caffeine administration restored Glut4 expression in the elderly group, but it was not able to increase Glut4 expression above a maximal level in the 12 M group. The mechanism by which caffeine increases Glut4 expression is still unclear, but the effect appears to be extant to several cell types: rat skeletal muscle primary cultures (Egawa et al 2009) and myoblasts (Mukwevho et al 2008), although contrasting results have been obtained in adipose tissue cells where caffeine suppressed insulininduced Glut4 translocation (Akiba et al 2004). …”

“…Recently, caffeine has also been implicated in skeletal muscle regulation of glucose and lipid metabolism. Caffeine stimulates insulin-independent muscle glucose transport (Wright et al 2004;Jensen et al 2007) and increases Glut4 mRNA and/or protein expression and fatty acid metabolism (Mukwevho et al 2008), having metabolic effects similar to the ones of 5′-AMP activated protein kinase (AMPK).…”

CaMK activation during exercise is required for histone hyperacetylation and MEF2A binding at the MEF2 site on theGlut4gene

“…First, Smith et al (33) infected C 2 C 12 myotubes with adenoviruses that expressed constitutively active (CA) or dominant negative (DN) CaMK IV and observed a twofold increase of Glut4 promoter-bound MEF2A in the CA CaMK IV-expressing myotubes compared with the DN-expressing controls. Second, Mukwevho et al (25) incubated C 2 C 12 myotubes with caffeine to activate CaMK II and found an ϳ2.2-fold increase in the amount of MEF2A that bound to the Glut4 gene compared with untreated controls, but inclusion of a CaMK II inhibitor (KN93) in the culture medium abolished the increase by caffeine. In both studies, CaMK activation also increased GLUT4 expression.…”

The role of CaMKII in regulating GLUT4 expression in skeletal muscle

“…Once bound, they prevent new activation of the subunit but have little effect on the activity of subunits that have already been activated by phosphorylation (102,110). KN93 has been widely used to inhibit CaMKII activity in cultured muscle cells, in isolated muscles, and in live animals (8,73,78). It has, however, been reported that, in addition to inhibiting CaMKII, KN93 also inhibits voltage-dependent K ϩ currents (87) and L-type calcium channel activity (30).…”

“…Mukwehvo et al (73) explored the mechanism by which caffeine increased GLUT4 expression in C 2 C 12 myotubes and tested whether these mechanisms were also CaMKII dependent. They used KN-93 to inhibit CaMKII and dantrolene to prevent Ca 2ϩ release from the sarcoplasmic reticulum.…”