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SH3KBP1 promotes skeletal myofiber formation and functionality through ER/SR architecture integrity

ABSTRACT: Dynamic changes in the arrangement of myonuclei and the organization of the sarcoplasmic reticulum are important determinants of myofiber formation and muscle function. To find factors associated to muscle integrity, we perform an siRNA screen and identify SH3KBP1 as a new factor controlling myoblast fusion, myonuclear positioning and myotube elongation. We find that the N-terminus of SH3KBP1 binds to dynamin-2 while the C-terminus associates with the endoplasmic reticulum through calnexin, which in turn control myonuclei dynamics and ER integrity, respectively. Additionally, in mature muscle fibers, SH3KBP1 contributes to the formation of triads and modulates the Excitation-Contraction Coupling process efficiency. In Dnm2R465W/+ mice, a model for centronuclear myopathy (CNM), depletion of Sh3kbp1 expression aggravates CNM-related atrophic phenotypes and impaired autophagic flux in mutant skeletal muscle fiber. Altogether our results identify SH3KBP1 as a new regulator of myofiber integrity and function.

SUBMITTER: Alexandre Guiraud

PROVIDER: S-SCDT-10_1038-S44319-025-00413-9 | biostudies-other |

REPOSITORIES: biostudies-other

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Project description:BackgroundAberrations to endoplasmic/sarcoplasmic reticulum (ER/SR) calcium concentration can result in the departure of endogenous proteins in a phenomenon termed exodosis. Redistribution of the ER/SR proteome can have deleterious effects to cell function and cell viability, often contributing to disease pathogenesis. Many proteins prone to exodosis reside in the ER/SR via an ER retention/retrieval sequence (ERS) and are involved in protein folding, protein modification, and protein trafficking. While the consequences of their extracellular presence have yet to be fully delineated, the proteins that have undergone exodosis may be useful for biomarker development. Skeletal muscle cells rely upon tightly coordinated ER/SR calcium release for muscle contractions, and perturbations to calcium homeostasis can result in myopathies. Ryanodine receptor type-1 (RYR1) is a calcium release channel located in the SR. Mutations to the RYR1 gene can compromise calcium homeostasis leading to a vast range of clinical phenotypes encompassing hypotonia, myalgia, respiratory insufficiency, ophthalmoplegia, fatigue and malignant hyperthermia (MH). There are currently no FDA approved treatments for RYR1-related myopathies (RYR1-RM).ResultsHere we examine the exodosis profile of skeletal muscle cells following ER/SR calcium depletion. Proteomic analysis identified 4,465 extracellular proteins following ER/SR calcium depletion with 1,280 proteins significantly different than vehicle. A total of 54 ERS proteins were identified and 33 ERS proteins significantly increased following ER/SR calcium depletion. Specifically, ERS protein, mesencephalic astrocyte-derived neurotrophic factor (MANF), was elevated following calcium depletion, making it a potential biomarker candidate for human samples. Despite no significant elevation of MANF in plasma levels among healthy volunteers and RYR1-RM individuals, MANF plasma levels positively correlated with age in RYR1-RM individuals, presenting a potential biomarker of disease progression. Selenoprotein N (SEPN1) was also detected only in extracellular samples following ER/SR calcium depletion. This protein is integral to calcium handling and SEPN1 variants have a causal role in SEPN1-related myopathies (SEPN1-RM). Extracellular presence of ER/SR membrane proteins may provide new insight into proteomic alterations extending beyond ERS proteins. Pre-treatment of skeletal muscle cells with bromocriptine, an FDA approved drug recently found to have anti-exodosis effects, curbed exodosis of ER/SR resident proteins.ConclusionChanges to the extracellular content caused by intracellular calcium dysregulation presents an opportunity for biomarker development and drug discovery.

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SH3KBP1 promotes skeletal myofiber formation and functionality through ER/SR architecture integrity

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