RhoA enhances store-operated Ca2+ entry and intestinal epithelial restitution by interacting with TRPC1 after wounding.
Early mucosal restitution occurs as a consequence of epithelial cell migration to resealing of superficial wounds after injury. Our previous studies show that canonical transient receptor potential-1 (TRPC1) functions as a store-operated Ca(2+) channel (SOC) in intestinal epithelial cells (IECs) and plays an important role in early epithelial restitution by increasing Ca(2+) influx. Here we further reported that RhoA, a small GTP-binding protein, interacts with and regulates TRPC1, thus enhancing SOC-mediated Ca(2+) entry (SOCE) and epithelial restitution after wounding. RhoA physically associated with TRPC1 and formed the RhoA/TRPC1 complexes, and this interaction increased in stable TRPC1-transfected IEC-6 cells (IEC-TRPC1). Inactivation of RhoA by treating IEC-TRPC1 cells with exoenzyme C3 transferase (C3) or ectopic expression of dominant negative RhoA (DNMRhoA) reduced RhoA/TRPC1 complexes and inhibited Ca(2+) influx after store depletion, which was paralleled by an inhibition of cell migration over the wounded area. In contrast, ectopic expression of wild-type (WT)-RhoA increased the levels of RhoA/TRPC1 complexes, induced Ca(2+) influx through activation of SOCE, and promoted cell migration after wounding. TRPC1 silencing by transfecting stable WT RhoA-transfected cells with siRNA targeting TRPC1 (siTRPC1) reduced SOCE and repressed epithelial restitution. Moreover, ectopic overexpression of WT-RhoA in polyamine-deficient cells rescued the inhibition of Ca(2+) influx and cell migration induced by polyamine depletion. These findings indicate that RhoA interacts with and activates TRPC1 and thus stimulates rapid epithelial restitution after injury by inducing Ca(2+) signaling.
Cell Biology Group, Department of Surgery, University of Maryland School of Medicine, Baltimore, Maryland; Baltimore Veterans Affairs Medical Center, Baltimore, Maryland.
Hee Kyoung
HK
Chung
Cell Biology Group, Department of Surgery, University of Maryland School of Medicine, Baltimore, Maryland; Baltimore Veterans Affairs Medical Center, Baltimore, Maryland.
Navneeta
N
Rathor
Cell Biology Group, Department of Surgery, University of Maryland School of Medicine, Baltimore, Maryland;
Shelley R
SR
Wang
Cell Biology Group, Department of Surgery, University of Maryland School of Medicine, Baltimore, Maryland; Department of Pathology, University of Maryland School of Medicine, Baltimore, Maryland; and Baltimore Veterans Affairs Medical Center, Baltimore, Maryland.
Jian-Ying
JY
Wang
Cell Biology Group, Department of Surgery, University of Maryland School of Medicine, Baltimore, Maryland; Baltimore Veterans Affairs Medical Center, Baltimore, Maryland rjaladanki@smail.umaryland.edu.
Jaladanki N
JN
Rao
D000818:Animals; D020013:Calcium Signaling; D002460:Cell Line; D002465:Cell Movement; D049109:Cell Proliferation; D004847:Epithelial Cells / Q000201:enzymology* / Q000473:pathology; D006801:Humans; D007413:Intestinal Mucosa / Q000201:enzymology* / Q000473:pathology; D034622:RNA Interference; D051381:Rats; D062326:Re-Epithelialization; D050052:TRPC Cation Channels / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D014945:Wound Healing; D020742:rhoA GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*
D050052:TRPC Cation Channels; C096351:transient receptor potential cation channel, subfamily C, member 1; C066737:RHOA protein, human; D020742:rhoA GTP-Binding Protein
Ca2+ influx; GTP-binding proteins; cyclopiazonic acid; epithelial restitution; polyamines