Deficiency of Src family kinases Fgr and Hck results in activation of erythrocyte K/Cl cotransport.
Src-family kinases play a central role in regulation of hematopoietic cell functions. We found that mouse erythrocytes express the Src-family kinases Fgr and Hck, as well as Lyn. To directly test whether Fgr and Hck play any role in erythrocyte function, we analyzed red cells isolated from fgr-/-, hck-/-, and fgr-/- hck-/- knock-out mice. Mean corpuscular hemoglobin concentration and median density are increased, while K content is decreased, in fgr-/- hck-/- double-mutant erythrocytes compared with wild-type, fgr-/-, or hck-/- erythrocytes. Na/K pump and Na/K/Cl cotransport were not altered, but K/Cl cotransport activity was significantly and substantially higher (approximately three-fold) in fgr-/- hck-/- double-mutant erythrocytes. This enhanced K/Cl cotransport activity did not depend on cell age. In fact, in response to bleeding, K/Cl cotransport activity increased in parallel with reticulocytosis in wild-type erythrocytes, while abnormal K/Cl cotransport did not change as a consequence of reticulocytosis in fgr-/- hck-/- double-mutant erythrocytes. Okadaic acid, an inhibitor of a phosphatase that has been implicated in activation of the K/Cl cotransporter, inhibited K/Cl cotransport in wild-type and fgr-/- hck-/- double-mutant erythrocytes to a comparable extent. In contrast, staurosporine, an inhibitor of a kinase that has been suggested to negatively regulate this same phosphatase enhanced K/Cl cotransport in wild-type but not in fgr-/- hck-/- double-mutant erythrocytes. On the basis of these findings, we propose that Fgr and Hck are the kinases involved in the negative regulation of the K/Cl cotransporter-activating phosphatase. Abnormality of erythrocyte K/Cl cotransport in fgr-/- hck-/- double-mutant animals represents the first demonstration that Src-family kinases may be involved in regulation of membrane transport.
De Franceschi
L
L
Institute of Internal Medicine, University of Verona, Italy.
Fumagalli
L
L
Olivieri
O
O
Corrocher
R
R
Lowell
C A
CA
Berton
G
G
D000818:Animals; D001692:Biological Transport / Q000235:genetics; D002352:Carrier Proteins / Q000378:metabolism; D002412:Cations / Q000032:analysis; D002712:Chlorides / Q000378:metabolism*; D004912:Erythrocytes / Q000378:metabolism*; D005260:Female; D016623:Ion Pumps / Q000378:metabolism; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D008954:Models, Biological; D011188:Potassium / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism; D051572:Proto-Oncogene Proteins c-hck; D017701:Reticulocyte Count; D028021:Sodium-Potassium-Chloride Symporters; D027981:Symporters; D019061:src-Family Kinases / Q000235:genetics / Q000378:metabolism*; D000096922:K Cl- Cotransporters
D002352:Carrier Proteins; D002412:Cations; D002712:Chlorides; D016623:Ion Pumps; D011188:Potassium; D011505:Protein-Tyrosine Kinases; D011518:Proto-Oncogene Proteins; D051572:Proto-Oncogene Proteins c-hck; D028021:Sodium-Potassium-Chloride Symporters; D027981:Symporters; D019061:src-Family Kinases; D000096922:K Cl- Cotransporters; C496632:Hck protein, mouse; C053102:proto-oncogene proteins c-fgr