Ablation of skeletal muscle triadin impairs FKBP12/RyR1 channel interactions essential for maintaining resting cytoplasmic Ca2+

Jose M. Eltit, Wei Feng, Jose R. Lopez, Isela T. Padilla, Isaac N Pessah, Tadeusz F. Molinski, Bradley R. Fruen, Paul D. Allen, Claudio F. Perez

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14 Scopus citations

Abstract

Previously, we have shown that lack of expression of triadins in skeletal muscle cells results in significant increase of myoplasmic resting free Ca 2+([Ca2+ rest), suggesting a role for triadins in modulating global intracellular Ca2+ homeostasis. To understand this mechanism, we study here how triadin alters [Ca2+ rest, Ca2+ release, and Ca2+ entry pathways using a combination of Ca2+ microelectrodes, channels reconstituted in bilayer lipid membranes (BLM), Ca2+, and Mn2+ imaging analyses of myotubes and RyR1 channels obtained from triadin-null mice. Unlike WT cells, triadin-null myotubes had chronically elevated [Ca2+] rest that was sensitive to inhibition with ryanodine, suggesting that triadin-null cells have increased basal RyR1 activity. Consistently, BLM studies indicate that, unlike WT-RyR1, triadin-null channels more frequently display atypical gating behavior with multiple and stable subconductance states. Accordingly, pulldown analysis and fluorescent FKBP12 binding studies in triadin-null muscles revealed a significant impairment of the FKBP12/RyR1 interaction. Mn2+ quench rates under resting conditions indicate that triadin-null cells also have higher Ca2+ entry rates and lower sarcoplasmic reticulum Ca2+ load than WT cells. Overexpression of FKBP12.6 reverted the null phenotype, reducing resting Ca2+ entry, recovering sarcoplasmic reticulum Ca2+ content levels, and restoring near normal [Ca2+]rest. Exogenous FKBP12.6 also reduced the RyR1 channel Po but did not rescue subconductance behavior. In contrast, FKBP12 neither reduced Po nor recovered multiple subconductance gating. These data suggest that elevated [Ca2+] rest in triadin-null myotubes is primarily driven by dysregulated RyR1 channel activity that results in part from impaired FKBP12/RyR1 functional interactions and a secondary increased Ca2+ entry at rest.

Original languageEnglish (US)
Pages (from-to)38453-38462
Number of pages10
JournalJournal of Biological Chemistry
Volume285
Issue number49
DOIs
StatePublished - Dec 3 2010

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ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Cite this

Eltit, J. M., Feng, W., Lopez, J. R., Padilla, I. T., Pessah, I. N., Molinski, T. F., Fruen, B. R., Allen, P. D., & Perez, C. F. (2010). Ablation of skeletal muscle triadin impairs FKBP12/RyR1 channel interactions essential for maintaining resting cytoplasmic Ca2+ Journal of Biological Chemistry, 285(49), 38453-38462. https://doi.org/10.1074/jbc.M110.164525