Development of a Rat Model to Investigate the Physiology of Telomerase Reverse Transcriptase alternative splicing isoforms

University Of Michigan At Ann Arbor, Ann Arbor MI

Investigators

Abstract

Aging is the greatest risk factor for cardiovascular diseases (CVD) and cancer development. Key primary aging pathways have been identified including mitochondrial dysfunction, reactive oxygen species damage, genome instability/telomere dysfunction, and inflammation underpin these two age related diseases. Development of interventions to slow/attenuate primary aging are of primary importance to healthy life extension. The catalytic subunit of telomerase, telomerase reverse transcriptase (TERT), has been well recognized for its nuclear role in telomere elongation. Recently, the importance of TERT splice isoforms for cellular health have been established. Over the past decade non-canonical mitochondrial functions of TERT have been described to protect cells from external stressors in a variety of disease pathologies, however no animal model that allows the study of physiological functions of existing splice variants is available. Understanding the physiological roles of TERT alternative splicing isoforms is of critical importance to aging, CVD, and cancer development because of their roles in mitochondrial biology. In direct response to an RFA from the Office of the Director (PAR21-167 - Development of Animal Models and Related Biological Materials for Research), we propose to generate and phenotype a transgenic rat model expressing a catalytically inactive splice variant, β-deletion variant (β-del- TERT) in comparison to a new full-length TERT (FL-TERT) transgenic rat model, and an already generated rat TERT knockout model. We will use these models to investigate the overarching hypothesis that independent of telomere lengthening or maintenance (human TERT will not function to elongate rat telomeres), FL-TERT will maintain mitochondrial functioning to a greater extent compared to β-del-TERT improving function, resilience, and ultimately slowed primary aging and reduced risk of disease progression. This model will represent a cross- cutting animal model that will be useful for a variety of research questions in the future well beyond the scope of these first two aims that we will test. To test our hypothesis the following two aims have been developed. Aim 1 will characterize and compare the newly generated transgenic rats to wildtype rats and existing (Beyer laboratory) rat TERT knockout animals for mitochondrial function, mitochondrial DNA damage, nuclear DNA damage, telomere associated DNA damage foci, and markers of cellular senescence in liver, intestine, skeletal muscle, heart, and the brain. Aim 2 will focus on the vasculature and hematopoietic system and directly test if β- del-TERT is sufficient to induce mitochondrial dysfunction that impairs endothelial function and promotes myeloid bias resulting in acceleration of cardiovascular disease progression via reduced endothelial cell dilation, macrophage polarization, and increased inflammatory stress. Successful completion of proposed work will provide strong proof of concept that enhancing mitochondrial TERT functions is a potential means to slow primary aging (reduced mitochondrial dysfunction, oxidative damage, and inflammation) and enhance healthy life expectation and aging resilience.