Loss of calcineurin homologous protein-1 in chicken B lymphoma DT40 cells destabilizes Na+/H+ exchanger isoform-1 protein.
NHE1/SLC9A1 is a ubiquitous isoform of vertebrate Na(+)/H(+) exchangers (NHEs) functioning in maintaining intracellular concentrations of Na(+) and H(+) ions. Calcineurin homologous protein-1 (CHP1) binds to the hydrophilic region of NHE1 and regulates NHE1 activity but reportedly does not play a role in translocating NHE1 from the endoplasmic reticulum to the plasma membrane. However, an antiport function of NHE1 requiring CHP1 remains to be clarified. Here we established CHP1-deficient chicken B lymphoma DT40 cells by gene targeting to address CHP1 function. CHP1-deficient cells showed extensive decreases in Na(+)/H(+) activities in intact cells. Although NHE1 mRNA levels were not affected, NHE1 protein levels were significantly reduced not only in the plasma membrane but in whole cells. The expression of a CHP1 transgene in CHP1-deficient cells rescued NHE1 protein expression. Expression of mutant forms of CHP1 defective in Ca(2+) binding or myristoylation also partially decreased NHE1 protein levels. Knockdown of CHP1 also caused a moderate decrease in NHE1 protein in HeLa cells. These data indicate that CHP1 primarily plays an essential role in stabilization of NHE1 for reaching of NHE1 to the plasma membrane and its exchange activity.
Matsushita
Masafumi
M
Dept. of Biological Sciences, Graduate School of Science, Osaka University, Machikaneyama-cho 1-1, Toyonaka City, Osaka, Japan 560-0043.
Sano
Yoshie
Y
Yokoyama
Shunsuke
S
Takai
Tomoyo
T
Inoue
Hiroki
H
Mitsui
Keiji
K
Todo
Kagefumi
K
Ohmori
Hitoshi
H
Kanazawa
Hiroshi
H
D000584:Amiloride / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D027682:Cation Transport Proteins / Q000235:genetics / Q000378:metabolism; D002645:Chickens; D015536:Down-Regulation; D017353:Gene Deletion; D006367:HeLa Cells; D006801:Humans; D016393:Lymphoma, B-Cell / Q000378:metabolism* / Q000473:pathology; D009154:Mutation; D019814:Myristic Acid / Q000378:metabolism; D009893:Opossums; D011485:Protein Binding; D020033:Protein Isoforms / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D012964:Sodium / Q000378:metabolism; D000074082:Sodium-Hydrogen Exchanger 1; D017923:Sodium-Hydrogen Exchangers / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*
C469743:CHP1 protein, human; D002135:Calcium-Binding Proteins; D027682:Cation Transport Proteins; D020033:Protein Isoforms; D012333:RNA, Messenger; D034741:RNA, Small Interfering; C490998:SLC9A1 protein, human; D000074082:Sodium-Hydrogen Exchanger 1; D017923:Sodium-Hydrogen Exchangers; D019814:Myristic Acid; D000584:Amiloride; D012964:Sodium; D002118:Calcium; C039614:ethylisopropylamiloride