This proposal aims to delineate molecular mechanisms that link laminar flow mediated signaling with gene expression, mitochondrial homeostasis and endothelial cell (EC) function. Carotid intima-media thickening (IMT) is caused by intima growth, and is a significant risk factor for cardiovascular diseases (CVD). Intima growth is mediated by EC dysfunction, vascular smooth muscle cell (VSMC) growth as well as inflammatory cell accumulation and activation. These pathological processes are stimulated by a disturbed flow pattern (d- flow), while being minimized by steady s-flow. Using congenic mouse strains, we identified a QTL for intima in C3H/F (no intima) and SJL (high intima) mice on chromosome 11 (Im2) that overlapped with a vascular inflammation QTL. Transcriptomic and bioinformatic analyses revealed significant differences in inflammation, cell cycle and RNA degradation. Using KEGG pathway analysis, a focus on genes in Im2 with polymorphisms that were differentially expressed between C3H/F and SJL identified a single gene: polyribonucleotide nucleotidyltransferase 1 (Pnpt1), a 3'-5' exoribonuclease that is required for import and processing of RNA in mitochondria. High level Pnpt1 expression correlated with decreased intima growth and inflammation in the carotid ligation model suggesting it was protective. The goal of this proposal is to understand how Pnpt1 restricts inflammation and atherosclerosis, focusing on novel transcriptional programs and mechanisms that link d-flow-mediated signaling through mitochondrial homeostasis, mitophagy/autophagy and cellular RNA processing pathways to EC dysfunction and CVD. While intima growth is primarily due to proliferation of VSMC and fibroblast-like cells, we focused on d-flow-mediated effects on EC because we believe these signals are initiating events, and are likely more specific and better therapeutic targets. We hypothesize that Pnpt1 is a mechanoresponsive enzyme that is critical to mitochondrial homeostasis and acts as a negative regulator of vascular inflammation and intima growth, thereby limiting CVD. Exciting preliminary data in support of the hypothesis include 1) inducible EC-specific Cre-loxP Pnpt1 mice that exhibit increased intima formation upon loss of Pnpt1; 2) RNA-Seq analyses of altered Pnpt1 expression under different flow patterns identified a novel and significant role for the TFAP2b/c transcription factor; 3) d-flow inhibited Pnpt1 function in EC; 4) Pnpt1 expression regulated EC inflammatory and apoptotic signaling both in vivo and in vitro and 5) Pnpt1 deficiency exacerbated mitochondrial-stress, as measured by ROS generation and autophagy. Proposed experiments will study changes in vascular remodeling and atherosclerosis in transgenic mouse models; determine the transcriptional program regulated by Pnpt1 focusing on the TFAP2b/c transcription factor; and the mechanisms by which flow regulates Pnpt1 function assayed by expression and enzyme activity. This proposal will characterize for the first time the role of Pnpt1, a major enzyme for mitochondrial RNA import and processing, in mouse models of atherosclerosis and vascular remodeling and in human carotid endarterectomy specimens.

Public Health Relevance

We are studying how one important gene (Polyribonucleotide Nucleotidyltransferase 1, Pnpt1) controls molecules inside endothelial cells (cells that line the interior of the blood vessel) that regulate inflammation and cellular function. If we determine how this gene controls these processes, we can design drugs that will mimic Pnpt1?s function and enhance its activity. In summary, our study will identify new ways to treat cardiovascular diseases such as aneurysms, atherosclerosis and hypertension.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL140958-02
Application #
9618545
Study Section
Atherosclerosis and Inflammation of the Cardiovascular System Study Section (AICS)
Program Officer
Olive, Michelle
Project Start
2018-01-01
Project End
2021-11-30
Budget Start
2019-01-01
Budget End
2019-12-31
Support Year
2
Fiscal Year
2019
Total Cost
Indirect Cost
Name
University of Rochester
Department
Internal Medicine/Medicine
Type
School of Medicine & Dentistry
DUNS #
041294109
City
Rochester
State
NY
Country
United States
Zip Code
14627