PMID- 10455146
OWN - NLM
STAT- MEDLINE
DCOM- 19990930
LR  - 20210209
IS  - 0021-9258 (Print)
IS  - 0021-9258 (Linking)
VI  - 274
IP  - 35
DP  - 1999 Aug 27
TI  - cAMP-induced phosphorylation and inhibition of Na(+)/H(+) exchanger 3 (NHE3) are 
      dependent on the presence but not the phosphorylation of NHE regulatory factor.
PG  - 24753-8
AB  - The members of the regulatory factor (RF) gene family, Na(+)/H(+) exchanger
      (NHE)-RF and NHE3 kinase A regulatory factor (E3KARP) are necessary for cAMP to
      inhibit the epithelial brush border NHE isoform 3 (NHE3). The mechanism of their 
      action was studied using PS120 fibroblasts stably transfected with rabbit NHE3
      and wild type rabbit NHE-RF or wild type human E3KARP. 8-Bromo-cAMP (8-Br-cAMP)
      had no effect on Na(+)/H(+) exchange activity in cells expressing NHE3 alone. In 
      contrast, in cells co-expressing NHE-RF, 8-Br-cAMP inhibited NHE3 by 39%. In vivo
      phosphorylation of NHE3 demonstrated that cAMP increased phosphorylation in two
      chymotrypsin-generated phosphopeptides of NHE3 in cells containing NHE-RF or
      E3KARP but not in cells lacking these proteins. The requirement for
      phosphorylation of NHE-RF in this cAMP-induced inhibition of NHE3 was examined by
      studying a mutant NHE-RF in which serines 287, 289, and 290 were mutated to
      alanines. Wild type NHE-RF was a phosphorylated protein under basal conditions,
      but treatment with 8-Br-cAMP did not alter its phosphorylation. Mutant NHE-RF was
      not phosphorylated either under basal conditions or after 8-Br-cAMP. 8-Br-cAMP
      inhibited NHE3 similarly in PS120/NHE3 cells containing wild type or mutant
      NHE-RF. NHE-RF and NHE3 co-precipitated and did so similarly with and without
      cAMP. Mutant NHE-RF also similarly immunoprecipitated NHE3 in the presence and
      absence of 8-Br-cAMP. This study shows that members of the regulatory factor gene
      family, NHE-RF and E3KARP, are necessary for cAMP inhibition of NHE3 by allowing 
      NHE3 to be phosphorylated. This inhibition is not dependent on the
      phosphorylation of NHE-RF.
FAU - Zizak, M
AU  - Zizak M
AD  - Department of Medicine, Gl Division, Johns Hopkins University School of Medicine,
      Baltimore, Maryland 21205, USA.
FAU - Lamprecht, G
AU  - Lamprecht G
FAU - Steplock, D
AU  - Steplock D
FAU - Tariq, N
AU  - Tariq N
FAU - Shenolikar, S
AU  - Shenolikar S
FAU - Donowitz, M
AU  - Donowitz M
FAU - Yun, C H
AU  - Yun CH
FAU - Weinman, E J
AU  - Weinman EJ
LA  - eng
GR  - P01 DK44484/DK/NIDDK NIH HHS/United States
GR  - R01 DK26523/DK/NIDDK NIH HHS/United States
GR  - R01 DK37319/DK/NIDDK NIH HHS/United States
GR  - etc.
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PT  - Research Support, U.S. Gov't, Non-P.H.S.
PT  - Research Support, U.S. Gov't, P.H.S.
PL  - United States
TA  - J Biol Chem
JT  - The Journal of biological chemistry
JID - 2985121R
RN  - 0 (Phosphopeptides)
RN  - 0 (Phosphoproteins)
RN  - 0 (Sodium-Hydrogen Exchangers)
RN  - 0 (sodium-hydrogen exchanger regulatory factor)
RN  - 23583-48-4 (8-Bromo Cyclic Adenosine Monophosphate)
RN  - E0399OZS9N (Cyclic AMP)
RN  - EC 3.4.21.1 (Chymotrypsin)
SB  - IM
MH  - 8-Bromo Cyclic Adenosine Monophosphate/pharmacology
MH  - Animals
MH  - Blotting, Western
MH  - Cell Line
MH  - Chymotrypsin
MH  - Cyclic AMP/*pharmacology
MH  - Humans
MH  - Microvilli/metabolism
MH  - Mutation
MH  - Phosphopeptides/analysis
MH  - Phosphoproteins/genetics/*metabolism
MH  - Phosphorylation
MH  - Precipitin Tests
MH  - Rabbits
MH  - Sodium-Hydrogen Exchangers/*antagonists & inhibitors
EDAT- 1999/08/24 00:00
MHDA- 1999/08/24 00:01
CRDT- 1999/08/24 00:00
PHST- 1999/08/24 00:00 [pubmed]
PHST- 1999/08/24 00:01 [medline]
PHST- 1999/08/24 00:00 [entrez]
AID - 10.1074/jbc.274.35.24753 [doi]
AID - S0021-9258(19)55412-6 [pii]
PST - ppublish
SO  - J Biol Chem. 1999 Aug 27;274(35):24753-8. doi: 10.1074/jbc.274.35.24753.