Changes in Na,K-ATPase gene expression during granulocytic differentiation of HL60 cells.
During granulocytic differentiation of HL60 cells, immediate reduction of ouabain-sensitive potassium flux is observed within the first 12 hours of addition of dimethyl sulfoxide (DMSO). We show that gene expression of the alpha 3 isoform of Na+,K(+)-ATPase, which encodes an ouabain-inhibitable Na+,K(+)-ATPase activity, significantly declines during the first 24 hours of granulocytic differentiation by DMSO of HL60 cells. The more common alpha 1 isoform decreases, but more gradually over 72 hours of DMSO induction. Loss of alpha 3 and alpha 1 messenger RNA (mRNA) are due to changes in mRNA decay; their transcription is not altered. alpha 3 mRNA half-life is 3 hours in HL60 cells; upon induction by 16 hours of DMSO, it decreases to approximately 2 hours. alpha 1 transcripts are less sensitive to DMSO induction, with their half-life being 3.5 hours in HL60 cells; upon induction, their half-life decreases to 3 hours. Experiments measuring protein stability confirm that alpha 3 protein is more labile than alpha 1. In uninduced HL60 cells, alpha 3 membrane protein comprises 30% of the total alpha isoforms, and is less stable than alpha 1, with a protein half-life of only 9 hours. Upon DMSO induction, steady-state alpha 3 protein decreases markedly within 10 hours, whereas alpha 1 protein remains stable. These results show that posttranscriptional changes during induction play a major role in the differential regulation of alpha 1 and alpha 3 isoforms of Na+,K(+)-ATPase; regulation of the latter may be important for early granulocytic differentiation, or for one of the differentiated functions of mature granulocytes.
Chambers
S K
SK
Department of Obstetrics and Gynecology, Yale University School of Medicine, New Haven, CT 06510.
Gilmore-Hebert
M
M
Kacinski
B M
BM
Benz
E J
EJ
D002454:Cell Differentiation; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D004355:Drug Stability; D015870:Gene Expression; D006098:Granulocytes / Q000166:cytology*; D006801:Humans; D007536:Isomerism; D007942:Leukemia, Experimental / Q000473:pathology; D015473:Leukemia, Promyelocytic, Acute / Q000235:genetics; D008565:Membrane Proteins / Q000032:analysis; D012333:RNA, Messenger / Q000592:standards; D000254:Sodium-Potassium-Exchanging ATPase / Q000235:genetics*; D014407:Tumor Cells, Cultured / Q000166:cytology*
D008565:Membrane Proteins; D012333:RNA, Messenger; D000254:Sodium-Potassium-Exchanging ATPase; D004121:Dimethyl Sulfoxide