Induced TRPC1 expression increases protein phosphatase 2A sensitizing intestinal epithelial cells to apoptosis through inhibition of NF-kappaB activation.
Transient receptor potential canonical-1 (TRPC1) functions as a store-operated Ca2+ channel in intestinal epithelial cells (IECs), and induced TRPC1 expression sensitizes IECs to apoptosis by inhibiting NF-kappaB activation. However, the exact mechanism by which increased TRPC1 results in NF-kappaB inactivation remains elusive. Protein phosphatase 2A (PP2A) is a widely conserved protein serine/threonine phosphatase that is implicated in the regulation of a wide array of cellular functions including apoptosis. The present study tests the hypothesis that induced TRPC1 expression inhibits NF-kappaB activation by increasing PP2A activity through Ca2+ influx in IECs. The expression of TRPC1 induced by stable transfection with the wild-type TRPC1 gene increased PP2A activity as indicated by increases in levels of PP2A proteins and their phosphatase activity. Increased levels of PP2A activity in stable TRPC1-transfected IEC-6 cells (IEC-TRPC1) were associated with decreased nuclear levels of NF-kappaB proteins and a reduction in NF-kappaB-dependent transcriptional activity, although there were no changes in total NF-kappaB protein levels. Inhibition of PP2A activity by treatment with okadaic acid or PP2A silencing with small interfering RNA not only enhanced NF-kappaB transactivation but also prevented the increased susceptibility of IEC-TRPC1 cells to apoptosis induced by treatment with tumor necrosis factor-alpha (TNF-alpha)/cycloheximide (CHX). Decreasing Ca2+ influx by exposure to the Ca2+-free medium reduced PP2A mRNA levels, destabilized PP2A proteins, and induced NF-kappaB activation, thus blocking the increased sensitivity of IEC-TRPC1 cells to TNF-alpha/CHX-induced apoptosis. These results indicate that induced TRPC1 expression increases PP2A activity through Ca2+ influx and that increased PP2A sensitizes IECs to apoptosis as a result of NF-kappaB inactivation.
Marasa
Bernard S
BS
Cell Biology Group, Department of Surgery, University of Maryland School of Medicine, Baltimore, Maryland, USA.
Xiao
Lan
L
Rao
Jaladanki N
JN
Zou
Tongtong
T
Liu
Lan
L
Wang
Jian
J
Bellavance
Emily
E
Turner
Douglas J
DJ
Wang
Jian-Ying
JY
D000818:Animals; D017209:Apoptosis; D002460:Cell Line; D005786:Gene Expression Regulation; D007413:Intestinal Mucosa / Q000166:cytology* / Q000502:physiology*; D016328:NF-kappa B / Q000037:antagonists & inhibitors*; D016133:Polymerase Chain Reaction; D054648:Protein Phosphatase 2 / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics; D051381:Rats; D050052:TRPC Cation Channels / Q000235:genetics*
D016328:NF-kappa B; D034741:RNA, Small Interfering; D050052:TRPC Cation Channels; C096351:transient receptor potential cation channel, subfamily C, member 1; C514014:Ppp2ca protein, rat; D054648:Protein Phosphatase 2