Protein Tyrosine Phosphatase-Dependent Activation of β-Globin and δ-Aminolevulinic Acid Synthase Genes in the Camptothecin-Induced IW32 Erythroleukemia Cell Differentiation
- 1Institute of Biochemistry, College of Life Sciences, National Yang-Ming University, Taipei, Taiwan 112, Republic of China (M.-C.W., F.-F.W.) and 2Department of Oncology, Veteran General Hospital, Taipei, Taiwan 112, Republic of China (J.H.L.)
Abstract
Camptothecin, an antitumor drug that specifically targets topoisomerase I, induced IW32 erythroleukemia cells to differentiate along the erythroid pathway, as demonstrated by the increased mRNA and protein expression of hemoglobin. Unlike other chemically induced erythroleukemia cell differentiation, no c-myc mRNA down-regulation was observed in the early phases of drug treatment. Among the heme-synthesizing enzyme mRNAs that were analyzed, only that of the erythroid-specific δ-aminolevulinic acid synthase (ALAS-E) was stimulated. Vanadate or benzylphosphonic acid, which inhibited protein tyrosine phosphatases (PTPase), blocked the camptothecin-induced differentiation. Maximal inhibition was attained if vanadate was added within the first 6 hr of camptothecin treatment, after which vanadate gradually lost its effectiveness. Camptothecin-induced expression of β-globin or ALAS-E transcript levels was inhibited in the presence of cycloheximide or vanadate. It was also shown that vanadate blocked differentiation of IW32 cells induced by sodium butyrate, VM-26, and p53. Increased PTPase activity could be observed 48 hr after cells were treated with camptothecin, VM-26, or sodium butyrate. Analysis of PTPase activity in the course of camptothecin treatment showed elevated levels of PTPase in the cytosol and the nucleus, with a greater increase demonstrated in the cytosol than in the nucleus. Our results suggest that by stimulating the β-globin and ALAS-E gene expression, PTPase plays a critical role in the induced differentiation of IW32 erythroleukemia cells.
Footnotes
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Send reprint requests to: Fung-Fang Wang, Ph.D., Institute of Biochemistry, National Yang-Ming University, Shih-Pai, Taipei, Taiwan 112, Republic of China.
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This work was supported by Grants NSC-82–0412-B010–035 from the National Science Council, Republic of China, and VGHYM 85-S4–19 from the Veteran General Hospital-Yang Ming Joint Foundation.
- Abbreviations:
- PTPase
- protein tyrosine phosphatase
- ALAS-E
- erythroid-specific δ-aminolevulinic acid synthase
- BPA
- benzylphosphonic acid
- TPA
- 12-O-tetradecanoylphorbol-13-acetate
- MEL
- murine erythroleukemia
- PMSF
- phenylmethylsulfonyl fluoride
- p-Npp
- para-nitrophenyl phosphate
- TBST
- Tris-buffered saline/Tween 20
- PBG-D
- porphobilinogen deaminase
- URO-D
- uroporphyrinogen III decarboxylase
- EGTA
- ethylene glycol bis(α-aminoethyl ether)-N,N,N′,N′-tetraacetic acid
- HEPES
- 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid
- DMSO
- dimethylsulfoxide
- DTT
- dithiothreitol
- GAPDH
- glyceraldehyde-3-phosphate dehydrogenase
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- Received August 13, 1996.
- Accepted December 12, 1996.
- The American Society for Pharmacology and Experimental Therapeutics
