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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Abstract
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 |
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| 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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