Na+-K+-Cl- cotransport in human fibroblasts is inhibited by cytomegalovirus infection.
We examined the effects of human cytomegalovirus (HCMV) infection on the Na+-K+-Cl- cotransporter (NKCC) in a human fibroblast cell line. Using the Cl--sensitive dye MQAE, we showed that the mock-infected MRC-5 cells express a functional NKCC. 1) Intracellular Cl- concentration ([Cl-]i) was significantly reduced from 53.4 +/- 3.4 mM to 35.1 +/- 3.6 mM following bumetanide treatment. 2) Net Cl- efflux caused by replacement of external Cl- with gluconate was bumetanide sensitive. 3) In Cl--depleted mock-infected cells, the Cl- reuptake rate (in HCO-3-free media) was reduced in the absence of external Na+ and by treatment with bumetanide. After HCMV infection, we found that although [Cl-]i increased progressively [24 h postexposure (PE), 65.2 +/- 4.5 mM; 72 h PE, 80.4 +/- 5.0 mM], the bumetanide and Na+ sensitivities of [Cl-]i and net Cl- uptake and loss were reduced by 24 h PE and abolished by 72 h PE. Western blots using the NKCC-specific monoclonal antibody T4 showed an approximately ninefold decrease in the amount of NKCC protein after 72 h of infection. Thus HCMV infection resulted in the abolition of NKCC function coincident with the severe reduction in the amount of NKCC protein expressed.
Maglova
L M
LM
Department of Physiology, Allegheny University of the Health Sciences, Philadelphia, Pennsylvania 19129, USA.
Crowe
W E
WE
Smith
P R
PR
Altamirano
A A
AA
Russell
J M
JM
D001692:Biological Transport / Q000187:drug effects; D002034:Bumetanide / Q000494:pharmacology; D002352:Carrier Proteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000378:metabolism*; D002460:Cell Line; D002712:Chlorides / Q000378:metabolism*; D004396:Coloring Agents; D003587:Cytomegalovirus / Q000502:physiology*; D003586:Cytomegalovirus Infections; D004622:Embryo, Mammalian; D005347:Fibroblasts; D006801:Humans; D007700:Kinetics; D008168:Lung; D012964:Sodium / Q000378:metabolism*; D028021:Sodium-Potassium-Chloride Symporters; D013997:Time Factors
D002352:Carrier Proteins; D002712:Chlorides; D004396:Coloring Agents; D028021:Sodium-Potassium-Chloride Symporters; D002034:Bumetanide; D012964:Sodium