Membrane Micropeptides and Calcium Pump Allostery

Loyola University Chicago, Maywood IL

Investigators

Linked publications & trials

Abstract

Project Summary/Abstract: The lead investigator, Seth Robia, and co-PI Gianluigi Veglia, share a mutual interest in SERCA regulation and have a track record of successful collaboration utilizing complementary approaches. Our prior collaboration received support from an NIH multi-PI R01, and this current proposal seeks to renew that grant. The proposed project aims to determine the influence of membrane micropeptides on the allosteric regulation of the SERCA Ca pump. These small proteins are expressed differently in various tissues, and each has a distinct way of regulating SERCA, but the structural determinants that contribute to these regulatory characteristics are still unknown. Our approach will involve using a combination of fluorescence methods and NMR spectroscopy to investigate the structure of the micropeptide/SERCA protein complex and dynamic protein-protein interactions. Subsequently, we will analyze how various micropeptides modulate SERCA function to govern cardiac contraction and relaxation. The planned experiments will uncover how SERCA activity is finely tuned in the healthy heart, enabling it to adapt to exercise or other physiological stressors. Additionally, we aim to uncover the pathogenic mechanisms associated with genetic variants that disrupt micropeptide regulatory processes, ultimately leading to heart failure. The research plan comprises two major aims. Aim 1 will identify unique determinants of function and more general features that are shared across different members of the membrane micropeptide family. In Aim 1a, we will determine how micropeptides modulate allosteric pathways of communication within the structure of the Ca pump. We hypothesize that micropeptides tune cellular calcium handling by modulating the pathway of communication between ATP- and Ca-binding sites on SERCA. Aim 1b will test our specific hypothesis that the tilt angle of the transmembrane domain dictates micropeptide inhibitory potency. Aim 2 is focused on mechanisms of heart failure. Aim 2a will examine the binding dynamics of the prototypical micropeptide phospholamban. We will measure how these interactions are changed by genetic variants associated with dilated cardiomyopathy. We hypothesize that pathogenic variants impair cardiac calcium handling through excessive stabilization of phospholamban oligomers. Aim 2b will test the specific hypothesis that a genetic variant linked to heart failure changes phospholamban structural dynamics, increasing its structural disorder and disrupting its inhibitory function.