Phosphoproteomic Profiling of Human Myocardial Tissues Distinguishes Ischemic from Non-Ischemic End Stage Heart Failure

Abstract

The molecular differences between ischemic (IF) and non-ischemic (NIF) heart failure are poorly defined. A better understanding of the molecular differences between these two heart failure etiologies may lead to the development of more effective heart failure therapeutics. In this study extensive proteomic and phosphoproteomic profiles of myocardial tissue from patients diagnosed with IF or NIF were assembled and compared.Proteins extracted from left ventricular sections were proteolyzed and phosphopeptides were enriched using titanium dioxide resin. Gel- and label-free nanoscale capillary liquid chromatography coupled to high resolution accuracy mass tandem mass spectrometry allowed for the quantification of 4,436 peptides (corresponding to 450 proteins) and 823 phosphopeptides (corresponding to 400 proteins) from the unenriched and phospho-enriched fractions, respectively.Protein abundance did not distinguish NIF from IF. In contrast, 37 peptides (corresponding to 26 proteins) exhibited a ≥2-fold alteration in phosphorylation state (p<0.05) when comparing IF and NIF. The degree of protein phosphorylation at these 37 sites was specifically dependent upon the heart failure etiology examined. Proteins exhibiting phosphorylation alterations were grouped into functional categories: transcriptional activation/RNA processing; cytoskeleton structure/function; molecular chaperones; cell adhesion/signaling; apoptosis; and energetic/metabolism.Phosphoproteomic analysis demonstrated profound post-translational differences in proteins that are involved in multiple cellular processes between different heart failure phenotypes. Understanding the roles these phosphorylation alterations play in the development of NIF and IF has the potential to generate etiology-specific heart failure therapeutics, which could be more effective than current therapeutics in addressing the growing concern of heart failure.

Myocardial expression analysis of osteopontin and its splice variants in patients affected by end-stage idiopathic or ischemic dilated cardiomyopathy

“…We have observed that levels of OPN-a were inversely related to myocardial fibrosis in the presence of similar decay of systolic function. We have detected larger myocardial deposits of type I collagen, fibronectin and fibroblasts in ICM rather than DCM hearts [31], in accord with previous studies [34][35][36][37][38]. Our data are also consistent with proteomic analysis performed by us on the same LV samples [31], which show the up-regulation of pro-apoptotic and pro-fibrotic factors in ICM failing hearts with overt fibrosis (ICM).…”

POPDC1S201F causes muscular dystrophy and arrhythmia by affecting protein trafficking

“…Experiments with a TREK-1 mutant, which lacks the PKA-dependent phosphorylation site, or pharmacological studies using PKA inhibitors will be informative in this context (32). Interestingly, POPDC1 is also subject to β-adrenergic-dependent phosphorylation (33,34). Thus, the regulation of POPDC1 involving cAMP binding, subcellular localization, and phosphorylation of itself and of interacting proteins deserves further investigations.…”

The Arabidopsis Proteins AtNHR2A and AtNHR2B Are Multi-Functional Proteins Integrating Plant Immunity With Other Biological Processes

“…Proteins bound to the beads were pre-digested with 0.2% of RapiGest SF (Waters Corporation, Milford, MA, United States) in 50 mM ammonium bicarbonate (Schechter et al, 2014). After pre-digestion, samples were reduced with 10 mM Tris (2-carboxyethyl) phosphine hydrochloride (TCEP-HCl) and alkylated with 25 mM iodoacetamide, followed by overnight digestion at 37 • C with sequencing grade trypsin (Promega, Madison, WI, United States) in 1:50 trypsin/protein ratio.…”