CHIP phosphorylation by protein kinase G enhances protein quality control and attenuates cardiac ischemic injury.

Proteotoxicity from insufficient clearance of misfolded/damaged proteins underlies many diseases. Carboxyl terminus of Hsc70-interacting protein (CHIP) is an important regulator of proteostasis in many cells, having E3-ligase and chaperone functions and often directing damaged proteins towards proteasome recycling. While enhancing CHIP functionality has broad therapeutic potential, prior efforts have all relied on genetic upregulation. Here we report that CHIP-mediated protein turnover is markedly post-translationally enhanced by direct protein kinase G (PKG) phosphorylation at S20 (mouse, S19 human). This increases CHIP binding affinity to Hsc70, CHIP protein half-life, and consequent clearance of stress-induced ubiquitinated-insoluble proteins. PKG-mediated CHIP-pS20 or expressing CHIP-S20E (phosphomimetic) reduces ischemic proteo- and cytotoxicity, whereas a phospho-silenced CHIP-S20A amplifies both. In vivo, depressing PKG activity lowers CHIP-S20 phosphorylation and protein, exacerbating proteotoxicity and heart dysfunction after ischemic injury. CHIP-S20E knock-in mice better clear ubiquitinated proteins and are cardio-protected. PKG activation provides post-translational enhancement of protein quality control via CHIP.

Pulseless Electrical Activity Cardiac Arrest.

Audience: This simulation-based scenario is appropriate for senior level emergency medicine residents. Introduction: Pulseless electrical activity (PEA) accounts for up to 25% of sudden cardiac arrest;1 therefore the ability to recognize and care for this condition is an essential skill of emergency medicine physicians. Management of PEA arrest in the emergency department centers on Advanced Cardiac Life Support (ACLS) algorithms and the identification and treatment of potentially reversible causes. Massive pulmonary embolism (PE) is one of several causes of PEA cardiac arrest.2 However, diagnosis by CT-angiographic or nuclear imaging may not be obtainable in the hemodynamically unstable patient, requiring physicians to have a high index of suspicion. Systemic thrombolytic therapy is indicated in cardiac arrest due to known or presumed massive pulmonary embolism.3,4,5. Educational Objectives: After competing this simulation-based session, the learner will be able to:Identify PEA arrestReview the ACLS commonly recognized PEA arrest etiologies via the H &T mnemonicReview and discuss the risks and benefits of tissue plasminogen activator (tPA) for massive PE. Educational Methods: This is a high-fidelity simulation that allows learners to evaluate and treat a PEA arrest secondary to massive PE in a safe environment. The learners will demonstrate their ability to recognize a PEA arrest, sort through possible etiologies, and demonstrate treatment of a massive PE with tPA. Debriefing will focus on diagnosis and management of the PEA arrest. Research Methods: This case was piloted with 12 PGY-2 and PGY-3 residents. Group and individual debriefing occurred post-case. Results: Post-simulation feedback from the faculty suggested two potential issues. First was fidelity, which we increased by using our ultrasound simulator. Second, the elevated presenting glucose with lactic acidosis could be a poor cue, leading some towards diabetic ketoacidosis (DKA). Discussion: Learners felt more confident about running a PEA arrest. The simulation improved resident awareness of the value of point of care ultrasound (POCUS) in cardiac arrest. It also clarified the dosing of tPA in massive PE. Faculty felt simulating the actual US without breaking simulation would be more challenging without our US simulator. Although there was concern about results pointing towards possible DKA, this did not occur in any of the pilot simulations. The presenting glucose was reduced to make this less likely in future simulations. Topics: Pulseless electrical activity (PEA), syncope, cardiac arrest, Hs and Ts from ACLS PEA instruction, tPA for massive PE, critical care medicine, simulation.

1H, 15N and 13C resonance assignments for free and IEEVD peptide-bound forms of the tetratricopeptide repeat domain from the human E3 ubiquitin ligase CHIP.

The ubiquitin ligase CHIP catalyzes covalent attachment of ubiquitin to unfolded proteins chaperoned by the heat shock proteins Hsp70/Hsc70 and Hsp90. CHIP interacts with Hsp70/Hsc70 and Hsp90 by binding of a C-terminal IEEVD motif found in Hsp70/Hsc70 and Hsp90 to the tetratricopeptide repeat (TPR) domain of CHIP. Although recruitment of heat shock proteins to CHIP via interaction with the CHIP-TPR domain is well established, alterations in structure and dynamics of CHIP upon binding are not well understood. In particular, the absence of a structure for CHIP-TPR in the free form presents a significant limitation upon studies seeking to rationally design inhibitors that may disrupt interactions between CHIP and heat shock proteins. Here we report the 1H, 13C, and 15N backbone and side chain chemical shift assignments for CHIP-TPR in the free form, and backbone chemical shift assignments for CHIP-TPR in the IEEVD-bound form. The NMR resonance assignments will enable further studies examining the roles of dynamics and structure in regulating interactions between CHIP and the heat shock proteins Hsp70/Hsc70 and Hsp90.