Functional coupling between TRPC3 and RyR1 regulates the expressions of key triadic proteins

Hui Lee Eun, Gennady Cherednichenko, Isaac N Pessah, P. D. Allen

Research output: Contribution to journalArticle

75 Citations (Scopus)

Abstract

We have shown that TRPC3(transient receptor potential channel canonical type 3) is sharply up-regulated during the early part of myotube differentiation and remains elevated in mature myotubes compared with myoblasts. To examine its functional roles in muscle, TRPC3 was "knocked down" in mouse primary skeletal myoblasts using retroviral-delivered small interference RNAs and single cell cloning. TRPC3 knockdown myoblasts (97.6 ± 1.9% reduction in mRNA) were differentiated into myotubes (TRPC3 KD) and subjected to functional and biochemical assays. By measuring rates of Mn2+ influx with Fura-2 and Ca2+ transients with Fluo-4, we found that neither excitation-coupled Ca2+ entry nor thapsigargin-induced store-operated Ca2+ entry was significantly altered in TRPC3 KD, indicating that expression of TRPC3 is not required for engaging either Ca2+ entry mechanism. In Ca2+ imaging experiments, the gain of excitation-contraction coupling and the amplitude of the Ca2+ release seen after direct RyR1 activation with caffeine was significantly reduced in TRPC3 KD. The decreased gain appears to be due to a decrease in RyR1 Ca 2+ release channel activity, because sarcoplasmic reticulum (SR) Ca2+ content was not different between TRPC3 KD and wild-type myotubes. Immunoblot analysis demonstrated that TRPC1, calsequestrin, triadin, and junctophilin 1 were up-regulated (1.46 ± 1.91-, 1.42 ± 0.08-, 2.99 ± 0.32-, and 1.91 ± 0.26-fold, respectively) in TRPC3 KD. Based on these data, we conclude that expression of TRPC3 is tightly regulated during muscle cell differentiation and propose that functional interaction between TRPC3 and RyR1 may regulate the gain of SR Ca2+ release independent of SR Ca2+ load.

Original languageEnglish (US)
Pages (from-to)10042-10048
Number of pages7
JournalJournal of Biological Chemistry
Volume281
Issue number15
DOIs
StatePublished - Apr 14 2006

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Transient Receptor Potential Channels
Ryanodine Receptor Calcium Release Channel
Proteins
Skeletal Muscle Fibers
Sarcoplasmic Reticulum
Myoblasts
Muscle
Calsequestrin
Skeletal Myoblasts
Excitation Contraction Coupling
Thapsigargin
Fura-2
Cloning
RNA Interference
Caffeine
Muscle Cells
Organism Cloning
Cell Differentiation
Assays
Chemical activation

ASJC Scopus subject areas

  • Biochemistry

Cite this

Functional coupling between TRPC3 and RyR1 regulates the expressions of key triadic proteins. / Eun, Hui Lee; Cherednichenko, Gennady; Pessah, Isaac N; Allen, P. D.

In: Journal of Biological Chemistry, Vol. 281, No. 15, 14.04.2006, p. 10042-10048.

Research output: Contribution to journalArticle

Eun, Hui Lee ; Cherednichenko, Gennady ; Pessah, Isaac N ; Allen, P. D. / Functional coupling between TRPC3 and RyR1 regulates the expressions of key triadic proteins. In: Journal of Biological Chemistry. 2006 ; Vol. 281, No. 15. pp. 10042-10048.
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