TRNT1 MUTATIONS LINKED TO EARLY ONSET AUTOSOMAL RECESSIVE RETINITIS PIGMENTOSA

Abstract

Purpose: Retinitis pigmentosa (RP), a heterogeneous group of monogenic disorders, is characterized by a variety of symptoms including night blindness and progressive death of photoreceptors. Through whole-exome sequencing, we recently identified novel mutations in the tRNA Nucleotidyl Transferase, CCA-Adding, 1 (TRNT1) gene in a proband displaying early-onset RP. Mutations in TRNT1 have been previously implicated in congenital sideroblastic anemia. The purpose of this study was 1) to demonstrate that TRNT1 mutations are causative of nonsyndromic autosomal recessive RP and 2) to elucidate the pathophysiologic mechanism of newly identified TRNT1 mutations in photoreceptor cell dysfunction and death.

Methods: Next generation whole exome sequencing was used to identify potential disease-causing mutations in a proband with clinically diagnosed early onset autosomal recessive RP. Zebrafish, human induced pluripotent stem cells (iPSC), gene addition transfer, and CRISPR/Cas9-mediated genome editing technologies were used to confirm pathogenicity and develop potential treatment strategies.

Results: Whole exome sequencing revealed two potential disease-causing mutations in the gene TRNT1. Morpholino-mediated knockdown of TRNT1 in zebrafish caused a visual function defect that could in part be rescued by injection of wild-type RNA. Dermal fibroblasts from the proband were expanded and targeted for iPSC generation. IPSC pluripotency was confirmed using standard rt-PCR, immunoblotting, immunocytochemistry and teratoma formation. The genetic mutations identified via whole exome sequencing were confirmed to alter retinal TRNT1 transcript and cause loss of full-length TRNT1 protein in iPSC-derived retinal progenitor cells.

Conclusions: Mutations in TRNT1 are capable of causing early onset, non-syndromic autosomal recessive RP. We have generated patient-specific iPSC-derived retinal cells to model disease in vitro. These cells will allow us to learn more about the pathophysiology and test multiple avenues of treatment for TRNT1-associated RP.