CaMKII protein expression and phosphorylation in human skeletal muscle by immunoblotting: Isoform specificity.

Autor: Martinez-Canton M; Department of Physical Education, University of Las Palmas de Gran Canaria, Campus Universitario de Tafira S/n, Las Palmas de Gran Canaria, 35017, Spain; Research Institute of Biomedical and Health Sciences (IUIBS), University of Las Palmas de Gran Canaria, Paseo Blas Cabrera Felipe 'Físico' s/n, 35017, Las Palmas de Gran Canaria, Spain., Gallego-Selles A; Department of Physical Education, University of Las Palmas de Gran Canaria, Campus Universitario de Tafira S/n, Las Palmas de Gran Canaria, 35017, Spain; Research Institute of Biomedical and Health Sciences (IUIBS), University of Las Palmas de Gran Canaria, Paseo Blas Cabrera Felipe 'Físico' s/n, 35017, Las Palmas de Gran Canaria, Spain., Galvan-Alvarez V; Department of Physical Education, University of Las Palmas de Gran Canaria, Campus Universitario de Tafira S/n, Las Palmas de Gran Canaria, 35017, Spain; Research Institute of Biomedical and Health Sciences (IUIBS), University of Las Palmas de Gran Canaria, Paseo Blas Cabrera Felipe 'Físico' s/n, 35017, Las Palmas de Gran Canaria, Spain., Garcia-Gonzalez E; Department of Physical Education, University of Las Palmas de Gran Canaria, Campus Universitario de Tafira S/n, Las Palmas de Gran Canaria, 35017, Spain; Research Institute of Biomedical and Health Sciences (IUIBS), University of Las Palmas de Gran Canaria, Paseo Blas Cabrera Felipe 'Físico' s/n, 35017, Las Palmas de Gran Canaria, Spain., Garcia-Perez G; Department of Physical Education, University of Las Palmas de Gran Canaria, Campus Universitario de Tafira S/n, Las Palmas de Gran Canaria, 35017, Spain; Research Institute of Biomedical and Health Sciences (IUIBS), University of Las Palmas de Gran Canaria, Paseo Blas Cabrera Felipe 'Físico' s/n, 35017, Las Palmas de Gran Canaria, Spain., Santana A; Research Institute of Biomedical and Health Sciences (IUIBS), University of Las Palmas de Gran Canaria, Paseo Blas Cabrera Felipe 'Físico' s/n, 35017, Las Palmas de Gran Canaria, Spain; Complejo Hospitalario Universitario Insular-Materno Infantil de Las Palmas de Gran Canaria, Clinical Genetics Unit, 35016, Las Palmas de Gran Canaria, Spain., Martin-Rincon M; Department of Physical Education, University of Las Palmas de Gran Canaria, Campus Universitario de Tafira S/n, Las Palmas de Gran Canaria, 35017, Spain; Research Institute of Biomedical and Health Sciences (IUIBS), University of Las Palmas de Gran Canaria, Paseo Blas Cabrera Felipe 'Físico' s/n, 35017, Las Palmas de Gran Canaria, Spain. Electronic address: marcos.martinrincon@ulpgc.es., Calbet JAL; Department of Physical Education, University of Las Palmas de Gran Canaria, Campus Universitario de Tafira S/n, Las Palmas de Gran Canaria, 35017, Spain; Research Institute of Biomedical and Health Sciences (IUIBS), University of Las Palmas de Gran Canaria, Paseo Blas Cabrera Felipe 'Físico' s/n, 35017, Las Palmas de Gran Canaria, Spain; School of Kinesiology, Faculty of Education, The University of British Columbia, Vancouver, BC, Canada; Department of Physical Performance, The Norwegian School of Sport Sciences, Postboks, 4014 Ulleval Stadion, 0806, Oslo, Norway. Electronic address: jose.calbet@ulpgc.es.
Jazyk: angličtina
Zdroj: Free radical biology & medicine [Free Radic Biol Med] 2024 Nov 01; Vol. 224, pp. 182-189. Date of Electronic Publication: 2024 Aug 24.
DOI: 10.1016/j.freeradbiomed.2024.08.030
Abstrakt: Calcium (Ca 2+ )/calmodulin-dependent protein kinase II (CaMKII) is activated during exercise by reactive oxygen species (ROS) and Ca 2+ transients initiating muscle contraction. CaMKII modulates antioxidant, inflammatory, metabolic and autophagy signalling pathways. CaMKII is coded by four homologous genes (α, β, γ, and δ). In rat skeletal muscle, δ D, δ A , γ D , γ B and β M have been described while different characterisations of human skeletal muscle CaMKII isoforms have been documented. Precisely discerning between the various isoforms is pivotal for understanding their distinctive functions and regulatory mechanisms in response to exercise and other stimuli. This study aimed to optimize the detection of the different CaMKII isoforms by western blotting using eight different CaMKII commercial antibodies in human skeletal muscle. Exercise-induced posttranslational modifications, i.e. phosphorylation and oxidations, allowed the identification of specific bands by multitargeting them with different antibodies after stripping and reprobing. The methodology proposed has confirmed the molecular weight of β M CaMKII and allows distinguishing between γ/δ and δ D CaMKII isoforms. The corresponding molecular weight for the CaMKII isoforms resolved were: δ D , at 54.2 ± 2.1 kDa; γ/δ, at 59.0 ± 1.2 kDa and 61.6 ± 1.3 kDa; and β M isoform, at 76.0 ± 1.8 kDa. Some tested antibodies showed high specificity for the δ D , the most responsive isoform to ROS and intracellular Ca 2+ transients in human skeletal muscle, while others, despite the commercial claims, failed to show such specificity.
Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
(Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)
Databáze: MEDLINE
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