Na+/H+ antiporter gene expression during monocytic differentiation of HL60 cells

Cardiology Division, Emory University School of Medicine, Atlanta, Georgia 30322.
Journal of Biological Chemistry (Impact Factor: 4.57). 08/1991; 266(21):13485-8.
Source: PubMed


During differentiation of human promyelocytic HL60 cells into monocytes there are sustained increases in intracellular pH and Na+/H+ antiporter activity. Here we show that increased transcription and expression of the gene for the Na+/H+ antiporter precedes phorbol 12-myristate 13-acetate (PMA)-induced HL60 cell differentiation. PMA increased steady-state Na+/H+ antiporter mRNA levels approximately 50-fold within 8 h (at which time less than 15% of cells had differentiated). This increase was due to an increased transcriptional rate as determined by nuclear run on. Immunoprecipitation of [35S]methionine-labeled Na+/H+ antiporter using an antiporter fusion protein antibody (RP1-c28) showed an equivalent increase in Na+/H+ antiporter protein synthesis. The synthetic diacylglycerol, 1-oleolyl-2-acetylglycerol, an activator of protein kinase C, which unlike PMA did not cause differentiation, failed to induce Na+/H+ antiporter mRNA. Furthermore, inhibition of PMA-induced differentiation by either sphingosine or cycloheximide prevented accumulation of Na+/H+ antiporter mRNA. Together, these findings strongly suggest a close association of Na+/H+ antiporter induction with HL60 cell differentiation. The HL60 cell system is a promising model to study the mechanisms of Na+/H+ antiporter gene regulation and its function in differentiation.

Download full-text


Available from: Jacques Pouysségur
  • Source
    • "Growth Inhibition of Lymphoma Cells by Monensin 405 human lymphoma cells by inducing cell cycle arrest and apoptosis, although there was a slight variation in susceptibility to monensin among the cell lines. It has long been considered that activation of the Na + /H + antiporter and the resulting intracellular alkalization are prerequisites for various cellular functions such as cell proliferation, differentiation and apoptosis (Dixon et al, 1987; Grinstein et al, 1989; Rao et al, 1991; Bental & Deutsch, 1994; Zhu & Loh, 1995; Cobo et al, 1998). In particular, monensin, a Na + ionophore, is known to transport one molecule of Na + into cells per molecule of H + transported out of cells, resulting in intracellular alkalinization (Pressman & Fashin, 1982; Nakazato & Hatano, 1991). "
    [Show abstract] [Hide abstract]
    ABSTRACT: Monensin, a Na+ ionophore, regulates many cellular functions, including apoptosis. We investigated the in vitro antiproliferative effect of monensin on nine human lymphoma cell lines. Monensin significantly inhibited the proliferation of all the lymphoma cell lines examined with a 50% inhibition concentration of about 0.5 micromol/l, and induced a G1 and/or a G2-M phase arrest in these cell lines. To address the antiproliferative mechanism of monensin, we examined the effect of this drug on cell-cycle-related proteins in CA46 cells (both G1 and G2 arrest) and Molt-4 cells (G2 arrest). Treatment with monensin for 72 h decreased CDK4 and cyclin A levels in CA46 cells, and cdc2 levels in Molt-4 cells. In monensin-treated CA46 cells, increased p21-CDK2, p27-CDK2 and p27-CDK4 complex forms were observed. And, in monensin-treated Molt-4 cells, increased p21-cdc2 complex form was detected. Furthermore, the activities of CDK2- and CDK4-associated kinases were reduced in association with Rb hypophosphorylation in monensin-treated CA46 cells. The activity of cdc2-associated kinase was decreased in both cell lines, which was accompanied by induction of Wee1. Also, monensin induced apoptosis in these cell lines, as evidenced by annexin V binding assay and flow cytometric detection of sub-G1 DNA content. This apoptotic process was associated with loss of mitochondria transmembrane potential (Delta(psi)m). Taken together, these results demonstrated for the first time that monensin potently inhibits the proliferation of human lymphoma cell lines via cell cycle arrest and apoptosis.
    Full-text · Article · Dec 2002 · British Journal of Haematology
  • [Show abstract] [Hide abstract]
    ABSTRACT: During differentiation of human leukemic HL60 cells into granulocytes, sustained increases in intracellular pH and Na+/H+ antiporter activity have been observed. In the present study we report that retinoic acid (RA)-induced granulocytic differentiation of HL60 cells causes an approximately 18-fold increase in the steady-state mRNA levels for the Na+/H+ antiporter. This was due to an increase in the rate of Na+/H+ antiporter gene transcription as measured by nuclear run-on analysis. Antiporter protein synthesis increased by seven-fold during RA-induced granulocytic differentiation of HL60 cells as measured by immunoprecipitation of 35S-methionine-labeled proteins with the RP1-c28 Na+/H+ antiporter antibody. No increase in antiporter mRNA was observed in response to etretinate, an analogue of retinoic acid, which did not induce differentiation. Thus, Na+/H+ antiporter gene expression is associated with RA-induced granulocytic differentiation of HL60 cells. The present findings and our previous data (Rao et al., 1991) demonstrate that Na+/H+ antiporter gene expression is a generalized feature of HL60 cell differentiation.
    No preview · Article · Jun 1992 · Journal of Cellular Physiology
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Chronic incubation of cultured renal tubular epithelial cells in acid medium causes an increase in Na/H antiporter activity that persists after removal from acid, is dependent on protein synthesis, and is associated with an increase in Na/H antiporter mRNA. Chronic activation of protein kinase C has similar effects in these cells. The present studies examined the role of protein kinase C in the effect of acid incubation. Incubation of MCT cells in acid for 24 h caused a 50% increase in Na/H antiporter activity. This was prevented by inhibition of protein kinase C, either with sphingosine or by protein kinase C downregulation. Pertussis toxin pretreatment did not prevent the increase in antiporter activity. Acid incubation caused an increase in transcription factor AP-1 activity, as shown by an increase in expression from a reporter gene containing six tandem AP-1 binding sites. This was associated with transient increases in c-fos and c-jun mRNAs. This response is typical of that for gene activation by protein kinase C. These studies demonstrate that acid activation of the Na/H antiporter requires protein kinase C and is associated with c-fos and c-jun expression and increased AP-1 activity.
    Full-text · Article · Jul 1992 · Proceedings of the National Academy of Sciences
Show more