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Termination codon readthrough of NNAT mRNA regulates calcium-mediated neuronal differentiation

Pandit, M and Akhtar, MN and Sundaram, S and Sahoo, S and Manjunath, LE and Eswarappa, SM (2023) Termination codon readthrough of NNAT mRNA regulates calcium-mediated neuronal differentiation. In: Journal of Biological Chemistry, 299 (9).

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Official URL: https://doi.org/10.1016/j.jbc.2023.105184


Termination codon readthrough (TCR) is a process in which ribosomes continue to translate an mRNA beyond a stop codon generating a C-terminally extended protein isoform. Here, we demonstrate TCR in mammalian NNAT mRNA, which encodes NNAT, a proteolipid important for neuronal differentiation. This is a programmed event driven by cis-acting RNA sequences present immediately upstream and downstream of the canonical stop codon and is negatively regulated by NONO, an RNA-binding protein known to promote neuronal differentiation. Unlike the canonical isoform NNAT, we determined that the TCR product (NNATx) does not show detectable interaction with the sarco/endoplasmic reticulum Ca2+-ATPase isoform 2 Ca2+ pump, cannot increase cytoplasmic Ca2+ levels, and therefore does not enhance neuronal differentiation in Neuro-2a cells. Additionally, an antisense oligonucleotide that targets a region downstream of the canonical stop codon reduced TCR of NNAT and enhanced the differentiation of Neuro-2a cells to cholinergic neurons. Furthermore, NNATx-deficient Neuro-2a cells, generated using CRISPR-Cas9, showed increased cytoplasmic Ca2+ levels and enhanced neuronal differentiation. Overall, these results demonstrate regulation of neuronal differentiation by TCR of NNAT. Importantly, this process can be modulated using a synthetic antisense oligonucleotide. © 2023 The Authors

Item Type: Journal Article
Publication: Journal of Biological Chemistry
Publisher: American Society for Biochemistry and Molecular Biology Inc.
Additional Information: The copyright for this article belongs to the Authors.
Keywords: Cell proliferation; Mammals; Neurons; Oligonucleotides; Proteins; RNA, Antisense oligonucleotides; Ca 2+; Down-stream; Isoforms; Neuro-2A cells; Neuronal differentiation; NNAT; SERCA2; Stop codons; Translational readthrough, Calcium
Department/Centre: Division of Biological Sciences > Biochemistry
UG Programme
Date Deposited: 22 Nov 2023 03:59
Last Modified: 22 Nov 2023 03:59
URI: https://eprints.iisc.ac.in/id/eprint/83197

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