Translational control in heart and lung disease by RNase A family

Univ Of Massachusetts Med Sch Worcester, Worcester MA

Investigators

Abstract

PROJECT SUMMARY Cardiovascular disease is the leading cause of death in the United States. Despite decades of research, it is unclear how the RNase A family nuclease angiogenin stimulates blood vessel formation, and why angiogenin dysfunction is associated with heart failure and poor cardiovascular health. Recent work revealed that angiogenin’s nuclease activity, which is required for its angiogenic function, is stimulated by binding to the ribosome, but it remains unclear whether angiogenin’s ribosome-dependent mechanism is involved in angiogenesis. A bacterial nuclease named ribocin with a strikingly angiogenin-like structure and ribosome- dependent activity holds similar potential for understanding lung health. Nearly all Cystic Fibrosis patients experience Pseudomonas aeruginosa bacterial pneumonia and subsequently suffer from lung tissue inflammation and lasting damage long after the infection has cleared. Ribocin encoded by P. aeruginosa damages human ribosomes specifically at central helix 69 of the 28S rRNA and inhibits translation. We hypothesize that like other ribosome-inactivating proteins, ribocin induces a ribotoxic stress response that causes inflammation and cell death in human lung tissues. To inform future therapeutic studies aimed at treating cardiovascular disease and post-infection pulmonary damage, the mechanisms of translation control by these RNase A-family nucleases must be elucidated in the context of their cellular functions. The goal of this project is to determine the structural basis of translation control by angiogenin during angiogenesis and determine the impact of translation control by ribocin on ribotoxic stress response and cell death. With guidance from the sponsor, an expert in biochemical and structural basis of translation, and collaborators, who are experts in the RNA developmental biology and cryo-EM method development, the trainee will apply cutting edge methods for in-cell cryogenic electron microscopy (cryo-EM) complemented by cell assay and biochemical approaches to visualize structural changes to actively translating ribosomes during angiogenin- stimulated vascularization and ribocin-mediated tissue damage. Aim 1 will determine the contribution of angiogenin’s ribosome-specific activity on tube formation (angiogenesis) in human umbilical vascular endothelial cells (HUVEC). Aim 2 will elucidate the structural mechanism of translation inhibition by ribocin and investigate the impact of selective ribosome damage by ribocin on ribotoxic stress response in human lung cells (IB3). The results of this study will reveal details of mechanisms underlying fundamental cardiovascular function and novel components of pulmonary disfunction, necessary for future work in developing therapeutics for cardiovascular and lung disease.