TY - JOUR
T1 - Asymmetric signal transduction in polarized ileal Na+-absorbing cells
T2 - Carbachol activates brush-border but not basolateral-membrane PIP2-PLC and translocates PLC-γ1 only to the brush border
AU - Khurana, Seema
AU - Kreydiyyeh, Sawsan
AU - Aronzon, Anna
AU - Hoogerwerf, W. Alexandra
AU - Rhee, Sue Goo
AU - Donowitz, Mark
AU - Cohen, Michael E.
PY - 1996/1/15
Y1 - 1996/1/15
N2 - In ileal Na+ absorptive cells, carbachol inhibits NaCl absorption and its component brush-border Na+/H+ exchanger, acting via basolateral membrane (BLM) receptors. This carbachol effect involves brush-border but not BLM protein kinase C. In the present work we describe another asymmetric aspect of signal transduction in these epithelial cells, this time involving phosphatidylinositol 4,5-bisphosphate (PIP2)-specific phospholipase C (PLC). Thirty seconds and 1 min after carbachol treatment, brush-border PIP2-specific PLC activity increased, returning to control levels by 2.5 min. Involvement of brush-border tyrosine kinase(s) in this effect was suggested by inhibition of the carbachol effect on NaCl absorption by the tyrosine kinase inhibitor genistein, added to the mucosal but not the serosal surface. Luminal genistein pretreatment also prevented the carbachol-induced increase in brush-border PLC activity. In contrast, carbachol exposure did not change the BLM PIP2-specific PLC activity. Western analysis and immunoprecipitation demonstrated that PLC-γ1 is present in the brush border and that carbachol increases the PLC-γ1 amount in the brush border. Both the brush border and BLM contain PLC-β3 and a small amount of PLC-δ1 but no PLC-β1, whereas BLM lacks detectable PLC-γ1. No change in PLC-β3 or PLC-δ1 amount in the brush border occurred with carbachol exposure. No change in tyrosine phosphorylation of brush-border PLC-γ1 occurred with carbachol treatment. The Ca2+ ionophore A23187 did not alter PIP2-specific PLC activity in either the brush border or the BLM. These studies demonstrate that carbachol but not Ca2+ ionophore effects on brush-border NaCl absorption are associated with increases in brush-border but not BLM PIP2-specific PLC activity and in the amount of brush-border PLC-γ1, and involve tyrosine phosphorylation. This asymmetric aspect of epithelial signal transduction, together with the previous demonstration of localization of high-sensitivity IP3 stores to the apical membrane area in intestinal epithelial cells, shows that different aspects of signal transduction occur at the apical and basolateral membranes in epithelia and requires studies in both domains to define mechanisms of intracellular signalling.
AB - In ileal Na+ absorptive cells, carbachol inhibits NaCl absorption and its component brush-border Na+/H+ exchanger, acting via basolateral membrane (BLM) receptors. This carbachol effect involves brush-border but not BLM protein kinase C. In the present work we describe another asymmetric aspect of signal transduction in these epithelial cells, this time involving phosphatidylinositol 4,5-bisphosphate (PIP2)-specific phospholipase C (PLC). Thirty seconds and 1 min after carbachol treatment, brush-border PIP2-specific PLC activity increased, returning to control levels by 2.5 min. Involvement of brush-border tyrosine kinase(s) in this effect was suggested by inhibition of the carbachol effect on NaCl absorption by the tyrosine kinase inhibitor genistein, added to the mucosal but not the serosal surface. Luminal genistein pretreatment also prevented the carbachol-induced increase in brush-border PLC activity. In contrast, carbachol exposure did not change the BLM PIP2-specific PLC activity. Western analysis and immunoprecipitation demonstrated that PLC-γ1 is present in the brush border and that carbachol increases the PLC-γ1 amount in the brush border. Both the brush border and BLM contain PLC-β3 and a small amount of PLC-δ1 but no PLC-β1, whereas BLM lacks detectable PLC-γ1. No change in PLC-β3 or PLC-δ1 amount in the brush border occurred with carbachol exposure. No change in tyrosine phosphorylation of brush-border PLC-γ1 occurred with carbachol treatment. The Ca2+ ionophore A23187 did not alter PIP2-specific PLC activity in either the brush border or the BLM. These studies demonstrate that carbachol but not Ca2+ ionophore effects on brush-border NaCl absorption are associated with increases in brush-border but not BLM PIP2-specific PLC activity and in the amount of brush-border PLC-γ1, and involve tyrosine phosphorylation. This asymmetric aspect of epithelial signal transduction, together with the previous demonstration of localization of high-sensitivity IP3 stores to the apical membrane area in intestinal epithelial cells, shows that different aspects of signal transduction occur at the apical and basolateral membranes in epithelia and requires studies in both domains to define mechanisms of intracellular signalling.
UR - http://www.scopus.com/inward/record.url?scp=0030032796&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0030032796&partnerID=8YFLogxK
U2 - 10.1042/bj3130509
DO - 10.1042/bj3130509
M3 - Article
C2 - 8573085
AN - SCOPUS:0030032796
SN - 0264-6021
VL - 313
SP - 509
EP - 518
JO - Biochemical Journal
JF - Biochemical Journal
IS - 2
ER -