Identification of an N-terminal TRPC2 splice variant which inhibits calcium influx

Xin Chu, Qin Tong, Jocelyn Wozney, Wenyi Zhang, Joseph Y. Cheung, Kathleen Conrad, Virginia Mazack, Richard Stahl, Dwayne L. Barber, Barbara Miller

Research output: Contribution to journalArticle

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Abstract

TRPC2 is a member of the transient receptor potential (TRP) superfamily of Ca2+-permeable channels expressed in nonexcitable cells. TRPC2 is involved in a number of physiological processes including sensory activation of the vomeronasal organ, sustained Ca2+ entry in sperm, and regulation of calcium influx by erythropoietin. Here, a new splice variant of TRPC2, called "Similar to mouse TRPC2"; (smTRPC2), was identified consisting of 213 amino acids, largely coincident with the N-terminus of TRPC2 clone 17. This splice variant lacks all six TRPC2 transmembrane domains and the calcium pore. Expression of smTRPC2 was found in all tissues examined by RT-PCR and in primary erythroid cells by RT-PCR and Western blotting. Confocal microscopy of CHO-S cells transfected with TRPC2 clone 14 and smTRPC2 demonstrated that TRPC2 clone 14 and smTRPC2 both localize at or near the plasma membrane and in the perinuclear region. Cell surface localization of TRPC2 was confirmed with biotinylation, and was not substantially affected by smTRPC2 expression. Coassociation of TRPC2 c14 and α with smTRPC2 was confirmed by immunoprecipitation. To examine the functional significance of smTRPC2 expression, a CHO-S model was used to study its effect on calcium influx stimulated by Epo through TRPC2. Single CHO-S cells which express transfected Epo-R were identified by detection of green fluorescent protein (GFP). Cells that express transfected TRPC2 c14 or α were identified by detection of blue fluorescent protein (BFP). [Ca]i was quantitiated with Fura Red fluorescence using digital video imaging. Epo stimulated calcium influx through TRPC2 isoforms c14 and α, which was inhibited by coexpression of smTRPC2. These data demonstrate that a short splice variant of TRPC2 exists in many cell types, which associates with and modifies the activity of functional TRPC2 splice variants.

Original languageEnglish (US)
Pages (from-to)173-182
Number of pages10
JournalCell Calcium
Volume37
Issue number2
DOIs
StatePublished - Jan 1 2005

Fingerprint

Calcium
CHO Cells
Clone Cells
Vomeronasal Organ
Physiological Phenomena
Biotinylation
Polymerase Chain Reaction
Erythroid Cells
Erythropoietin
Green Fluorescent Proteins
Immunoprecipitation
Confocal Microscopy
Spermatozoa
Protein Isoforms
Fluorescence
Western Blotting
Cell Membrane
Amino Acids
Proteins
fura red

All Science Journal Classification (ASJC) codes

  • Physiology
  • Molecular Biology
  • Cell Biology

Cite this

Chu, Xin ; Tong, Qin ; Wozney, Jocelyn ; Zhang, Wenyi ; Cheung, Joseph Y. ; Conrad, Kathleen ; Mazack, Virginia ; Stahl, Richard ; Barber, Dwayne L. ; Miller, Barbara. / Identification of an N-terminal TRPC2 splice variant which inhibits calcium influx. In: Cell Calcium. 2005 ; Vol. 37, No. 2. pp. 173-182.
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abstract = "TRPC2 is a member of the transient receptor potential (TRP) superfamily of Ca2+-permeable channels expressed in nonexcitable cells. TRPC2 is involved in a number of physiological processes including sensory activation of the vomeronasal organ, sustained Ca2+ entry in sperm, and regulation of calcium influx by erythropoietin. Here, a new splice variant of TRPC2, called {"}Similar to mouse TRPC2{"}; (smTRPC2), was identified consisting of 213 amino acids, largely coincident with the N-terminus of TRPC2 clone 17. This splice variant lacks all six TRPC2 transmembrane domains and the calcium pore. Expression of smTRPC2 was found in all tissues examined by RT-PCR and in primary erythroid cells by RT-PCR and Western blotting. Confocal microscopy of CHO-S cells transfected with TRPC2 clone 14 and smTRPC2 demonstrated that TRPC2 clone 14 and smTRPC2 both localize at or near the plasma membrane and in the perinuclear region. Cell surface localization of TRPC2 was confirmed with biotinylation, and was not substantially affected by smTRPC2 expression. Coassociation of TRPC2 c14 and α with smTRPC2 was confirmed by immunoprecipitation. To examine the functional significance of smTRPC2 expression, a CHO-S model was used to study its effect on calcium influx stimulated by Epo through TRPC2. Single CHO-S cells which express transfected Epo-R were identified by detection of green fluorescent protein (GFP). Cells that express transfected TRPC2 c14 or α were identified by detection of blue fluorescent protein (BFP). [Ca]i was quantitiated with Fura Red fluorescence using digital video imaging. Epo stimulated calcium influx through TRPC2 isoforms c14 and α, which was inhibited by coexpression of smTRPC2. These data demonstrate that a short splice variant of TRPC2 exists in many cell types, which associates with and modifies the activity of functional TRPC2 splice variants.",
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Chu, X, Tong, Q, Wozney, J, Zhang, W, Cheung, JY, Conrad, K, Mazack, V, Stahl, R, Barber, DL & Miller, B 2005, 'Identification of an N-terminal TRPC2 splice variant which inhibits calcium influx', Cell Calcium, vol. 37, no. 2, pp. 173-182. https://doi.org/10.1016/j.ceca.2004.08.005

Identification of an N-terminal TRPC2 splice variant which inhibits calcium influx. / Chu, Xin; Tong, Qin; Wozney, Jocelyn; Zhang, Wenyi; Cheung, Joseph Y.; Conrad, Kathleen; Mazack, Virginia; Stahl, Richard; Barber, Dwayne L.; Miller, Barbara.

In: Cell Calcium, Vol. 37, No. 2, 01.01.2005, p. 173-182.

Research output: Contribution to journalArticle

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AU - Chu, Xin

AU - Tong, Qin

AU - Wozney, Jocelyn

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Chu X, Tong Q, Wozney J, Zhang W, Cheung JY, Conrad K et al. Identification of an N-terminal TRPC2 splice variant which inhibits calcium influx. Cell Calcium. 2005 Jan 1;37(2):173-182. https://doi.org/10.1016/j.ceca.2004.08.005