P300 or CBP are required for calorie restriction-mediated enhancement of skeletal muscle insulin sensitivity

Abstract

A calorie-restricted diet robustly improves skeletal muscle insulin sensitivity and is a cornerstone lifestyle intervention for preventing and treating clinical hyperglycemia. While studies demonstrate that insulin-stimulated activation of phosphatidylinositol 3-kinase (PI3K) and RAC-beta serine/threonine-protein kinase (AKT2) are fundamental to enhanced muscle insulin sensitivity after calorie restriction (CR), the downstream signaling steps remain to be fully elucidated. We have previously demonstrated that the lysine acetyltransferases, E1A binding protein (p300) or cAMP response element binding protein binding protein (CBP) are required for insulin-stimulated glucose uptake by skeletal muscle, at both sub-maximal and maximal concentrations of insulin. Moreover, p300/CBP are phosphorylated by AKT, which increases their acetyltransferase activity. Considering these findings together, the objective of this study was to determine whether p300 or CBP are required for the ability of CR to enhance skeletal muscle insulin sensitivity. We hypothesized that combined inhibition of p300 and CBP acetyltransferase activity would prevent CR-induced enhancement of skeletal muscle insulin sensitivity. To address this objective, 8-week-old male C57BL/6NJ mice were either fed ad libitum (AL; n=16) or were fed a CR diet (60% of AL intake; n=16) diet for 20 days. Then, an ex vivo [3H]-2-deoxyglucose (2DOG) approach was used to assess basal and insulin-stimulated (60μU/mL [0.36nM]) 2DOG uptake in paired extensor digitorum longus (EDL) and soleus from fasted (4 h) mice. To define the role of p300/CBP acetyltransferase activity, paired muscles from AL- or CR-fed mice were pre-incubated for 60 minutes with either DMSO (experimental control; n=8/dietary group [AL-DMSO, CR-DMSO]) or the p300/CBP acetyltransferase activity inhibitor, ip300w (25mM; n=8/dietary group [AL-ip300w, CR-ip300w]). As expected, insulin-stimulated 2DOG uptake (i.e. Insulin 2DOG minus basal 2DOG) in DMSO incubated muscles was ~2.3-fold and ~3.3-fold higher in the soleus and EDL, respectively, from CR- vs. AL-fed mice. Remarkably, ip300w not only blocked insulin-stimulated 2DOG uptake in AL-fed mice, but it also abrogated the insulin-sensitizing effects of CR on muscle insulin sensitivity. To further define the importance of p300 and CBP to CR-induced enhancement of skeletal muscle insulin sensitivity, we studied mice with tamoxifen-inducible and skeletal muscle-specific knockout of CBP and heterozygous (HZ) loss of p300 (referred to as, i-mCKO/PHZ) or knockout of p300 and HZ loss of CBP (referred to as, i-mPKO/CHZ). In these mice, the AL diet or 20 days of CR was initiated 3 weeks after ~10 week-old mice were dosed with tamoxifen (5 consecutive days, 2 mg/day); Cre recombinase-negative littermates, who were also dosed with tamoxifen, were the experimental controls (referred to as wildtype [WT]). Interestingly, oral glucose tolerance and EDL insulin sensitivity were comparably enhanced by CR in WT and both i-mCKO/PHZ and i-mPKO/CHZ mice, thus suggesting that just one allele of p300 or CBP is suffcient for CR to enhance skeletal muscle insulin sensitivity. Taken together, these results demonstrate that p300 or CBP are essential for the ability of calorie restriction to enhance skeletal muscle insulin sensitivity. This work was supported by a grant from the National Institutes of Health (R56 DK1311211), and the Dr. Maryam Ahmadian Memorial Summer Fellowship Program (L.M.L.) and the UC San Diego Medical Scientist Training Program (L.S.F.). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.