Diabetes is a major risk factor for heart failure, and no mechanism-specific therapies targeting the diabetic heart are currently available. A better understanding of diabetes-associated remodelling of cardiac proteome and post-translational modification (PTM) landscape may provide critical insights into disease mechanisms. A PTM significantly altered in the diabetic myocardium is protein O-GlcNAcylation; a highly dynamic form of glycosylation which regulates cell signalling through attachment of monosaccharide N-acetyl glucosamine to serine and threonine residues. Although excessive O-GlcNAcylation is associated with cardiac dysfunction, the identity and functional association of these modified proteins in the human diabetic myocardium remain unknown.
Human right atrial appendage (RAA) and left ventricle (LV) biopsies were obtained from diabetic and non-diabetic patients and investigated for differences in global, regional, and O-GlcNAc post translationally modified proteomes. O-GlcNAc modified proteins were isolated from tissue lysates through an approach combining enzymatic labelling, click chemistry, and affinity enrichment. Global lysates and O-GlcNAc and control captures underwent single-pot, solid-phase-enhanced sample preparation (SP3) and data independent acquisition (DIA) MS/MS analysis.
Global proteomic profiling revealed greater differences in diabetic RAA than LV, with diabetic RAA exhibiting dysregulated protein networks associated with necroptosis, energy metabolism, spliceosome, and membrane trafficking, and diabetic LV displayed alterations to proteoglycan, energy metabolism, and stress response components. Moreover, paired inter-tissue comparison revealed alterations to regional protein expression in diabetic patients. Investigation of O-GlcNAc modified proteins further delineated significant differences between diabetic and non-diabetic tissues. Differentially O-GlcNAcylated proteins in diabetic RAA were associated with cardiac contraction, cell junctions, autophagy, immune response, energy metabolism, translation, glycosylation, and cell death, and those in diabetic LV were associated with cardiomyopathies, contraction, ECM organization, adrenergic signalling, and inflammation.
Here, we provide critical insights into diabetes associated changes to the global and O-GlcNAc-modified proteome in human RAA and LV. These data reveal significant regional differences of the cardiac proteome and its modification with O-GlcNAc during diabetes, shedding light on potential molecular drivers of myocardial dysfunction.