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Olfactory marker protein is unlikely to be cleaved by calpain 5

Olfactory maturation marker protein (OMP) is expressed in olfactory receptor neurons and hypothalamic neurons. OMP is a nested gene located in the intron of calpain 5 (CAPN5), a Ca(2+)-dependent cysteine protease. Despite being located at the same genomic locus, genetic regulation of the reciprocal...

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Autores principales: Nakashima, Noriyuki, Nakashima, Kie, Nakashima, Akiko, Takano, Makoto
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9618205/
https://www.ncbi.nlm.nih.gov/pubmed/36309704
http://dx.doi.org/10.1186/s13041-022-00971-2
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author Nakashima, Noriyuki
Nakashima, Kie
Nakashima, Akiko
Takano, Makoto
author_facet Nakashima, Noriyuki
Nakashima, Kie
Nakashima, Akiko
Takano, Makoto
author_sort Nakashima, Noriyuki
collection PubMed
description Olfactory maturation marker protein (OMP) is expressed in olfactory receptor neurons and hypothalamic neurons. OMP is a nested gene located in the intron of calpain 5 (CAPN5), a Ca(2+)-dependent cysteine protease. Despite being located at the same genomic locus, genetic regulation of the reciprocal expression of OMP and CAPN5 has been suggested. By performing a motif search, we detected possible calpain cleavage sites in OMP. However, the direct proteolytic regulation of OMP by CAPN5 is unclear. Here, we generated OMP fused with Myc-tag and His-tag at its N- and C-termini and examined whether CAPN5 cleaves OMP into fragments by detecting immunoreactivity against Myc, OMP and His. Western blotting demonstrated that OMP was unlikely to be cleaved even in the presence of Ca(2+) in vitro. We expressed OMP and CAPN5 in HEK293T cells and applied a calcium ionophore under physiological conditions in cellulo, which resulted in no apparent fragmentation of OMP. We also applied liquid chromatography/mass spectrometry to the electrophoresed fractions smaller than the uncut Myc-OMP-His signals, which demonstrated no significant fragmentation of OMP. These results collectively indicate that OMP is unlikely to be cleaved by CAPN5. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s13041-022-00971-2.
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spelling pubmed-96182052022-10-31 Olfactory marker protein is unlikely to be cleaved by calpain 5 Nakashima, Noriyuki Nakashima, Kie Nakashima, Akiko Takano, Makoto Mol Brain Micro Report Olfactory maturation marker protein (OMP) is expressed in olfactory receptor neurons and hypothalamic neurons. OMP is a nested gene located in the intron of calpain 5 (CAPN5), a Ca(2+)-dependent cysteine protease. Despite being located at the same genomic locus, genetic regulation of the reciprocal expression of OMP and CAPN5 has been suggested. By performing a motif search, we detected possible calpain cleavage sites in OMP. However, the direct proteolytic regulation of OMP by CAPN5 is unclear. Here, we generated OMP fused with Myc-tag and His-tag at its N- and C-termini and examined whether CAPN5 cleaves OMP into fragments by detecting immunoreactivity against Myc, OMP and His. Western blotting demonstrated that OMP was unlikely to be cleaved even in the presence of Ca(2+) in vitro. We expressed OMP and CAPN5 in HEK293T cells and applied a calcium ionophore under physiological conditions in cellulo, which resulted in no apparent fragmentation of OMP. We also applied liquid chromatography/mass spectrometry to the electrophoresed fractions smaller than the uncut Myc-OMP-His signals, which demonstrated no significant fragmentation of OMP. These results collectively indicate that OMP is unlikely to be cleaved by CAPN5. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s13041-022-00971-2. BioMed Central 2022-10-29 /pmc/articles/PMC9618205/ /pubmed/36309704 http://dx.doi.org/10.1186/s13041-022-00971-2 Text en © The Author(s) 2022 https://creativecommons.org/licenses/by/4.0/Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) . The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/ (https://creativecommons.org/publicdomain/zero/1.0/) ) applies to the data made available in this article, unless otherwise stated in a credit line to the data.
spellingShingle Micro Report
Nakashima, Noriyuki
Nakashima, Kie
Nakashima, Akiko
Takano, Makoto
Olfactory marker protein is unlikely to be cleaved by calpain 5
title Olfactory marker protein is unlikely to be cleaved by calpain 5
title_full Olfactory marker protein is unlikely to be cleaved by calpain 5
title_fullStr Olfactory marker protein is unlikely to be cleaved by calpain 5
title_full_unstemmed Olfactory marker protein is unlikely to be cleaved by calpain 5
title_short Olfactory marker protein is unlikely to be cleaved by calpain 5
title_sort olfactory marker protein is unlikely to be cleaved by calpain 5
topic Micro Report
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9618205/
https://www.ncbi.nlm.nih.gov/pubmed/36309704
http://dx.doi.org/10.1186/s13041-022-00971-2
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