Phospho‐Proteomic Analysis of Cardiac Dyssynchrony and Resynchronization Therapy

Abstract

Cardiac dyssynchrony arises from conduction abnormalities during heart failure and worsens morbidity and mortality. Cardiac resynchronization therapy (CRT) re-coordinates contraction using bi-ventricular pacing, but the cellular and molecular mechanisms involved remain largely unknown. The aim is to determine how dyssynchronous heart failure (HF ) alters the phospho-proteome and how CRT interacts with this unique phospho-proteome by analyzing Ser/Thr and Tyr phosphorylation. Phospho-enriched myocardium from dog models of Control, HF , and CRT is analyzed via MS. There were 209 regulated phospho-sites among 1761 identified sites. Compared to Con and CRT, HF is hyper-phosphorylated and tyrosine phosphorylation is more likely to be involved in signaling that increased with HF and was exacerbated by CRT. For each regulated site, the most-likely targeting-kinase is predicted, and CK2 is highly specific for sites that are "fixed" by CRT, suggesting activation of CK2 signaling occurs in HF that is reversed by CRT, which is supported by western blot analysis. These data elucidate signaling networks and kinases that may be involved and deserve further study. Importantly, a possible role for CK2 modulation in CRT has been identified. This may be harnessed in the future therapeutically to compliment CRT, improving its clinical effects.

Hyperphosphorylation of hepatic proteome characterizes nonalcoholic fatty liver disease in S-adenosylmethionine deficiency

“…Titanium Dioxide (TiO2) phosphorylation affinity enrichment was performed as previously described. 58 Briefly, 400 μg of digested peptides were incubated in 50 μL titanium dioxide (TiO2) slurry (30 mg/mL, Glygen Corp, Columbia, MD) at room temperature on a shaker for 16 hrs. TiO2 beads were washed twice with 200 μL of 80% ACN, 5% TFA, once with 200 μL of 80% ACN, 0.1% TFA, and eluted in 180 μL of 30% ACN/ 1% NH4OH and neutralized with 200 μL of 10% FA.…”

PINE: An Automation Tool to Extract and Visualize Protein-Centric Functional Networks

“…Titanium dioxide (TiO 2 ) phospho enrichment was performed as previously described . Briefly, 400 μg of sample was digested as described above, individually desalted on Oasis HLB cartridges (Water, 10 mg), and eluted in 300 μL of 80% ACN, 5% TFA, 1 mM glycolic acid.…”

Inflammation and Heart Failure: Searching for the Enemy—Reaching the Entelechy

“…In other words, the body tries to protect itself from itself. Indeed, when mitochondrial morphology and function are disturbed (lack of fission, fusion and hence mitophagy), mitochondrial DNA is released into cytosol, and along with the misfolded proteins and the activation of the mitochondria-associated endoplasmic reticulum membranes (MAMs), promotes the enhancement of a self-destruction process, that might involve the entire body [ 33 , 34 ]. In case of a cardiac harmful event, there is an activation of danger-associated molecular patterns (DAMP) released by the nucleus (e.g., DNA, RNA), the mitochondria (e.g., DNA) and the cytosol (e.g., RNA).…”