Coordinate expression and developmental role of Id2 protein and TAL1/E2A heterodimer in erythroid progenitor differentiation

Gianluigi Condorelli, Luigi Vitelli, Mauro Valtieri, Isabella Marta, Elisabetta Montesoro, Valentine Lulli, Richard Baer, Cesare Peschle

Research output: Contribution to journalArticle

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Abstract

The Id proteins and basic helix-loop-helix (bHLH) proteins play major roles in specifying cell fate decisions in diverse biologic settings. A potential role for Id and TAL1/E2A bHLH genes in hematopoiesis has been suggested by studies on immortalized cell lines. However, it is uncertain whether these observations reflect normal hematopoiesis. We have investigated the expression pattern of Id2 and TAL1/E2A genes in liquid suspension culture of purified hematopoietic progenitor cell (HPCs) undergoing erythroid or granulopoietic differentiation in the first culture week and maturation to terminal cells in the second week. In quiescent, freshly purified HPCs, Id2 mRNA is detected by reverse transcriptase-polymerase chain reaction (RT- PCR), whereas TAL1 and E2A mRNAs are not. At the onset of erythroid differentiation, Id2 mRNA is downregulated, while E2A and TAL1 mRNAs are concomitantly upregulated: their expression is further increased at erythroblast level. Conversely, Id2 is not downmodulated in granulopoietic culture, except for a late decline at day 10 to 12, while TAL1 and E2A are only transiently induced in the first week of granulopoietic differentiation. The expression pattern of the TAL1/E2A heterodimer, as evaluated by mobility shift assay, is consistent with RT-PCR results (except for lower levels of the heterodimer in late erythroid maturation). TAL1 protein level, analyzed by Western blot, shows a pattern consistent with gel-shift results. Functional experiments were performed on purified HPCs treated with phosphorothioate antisense oligodeoxynucleotides to Id2 or TAL1 mRNA. The results are strictly consistent with the expression studies: anti-Id2 oligomer (α-Id2) causes a significant dose-dependent increase of erythroid colony formation, whereas α-TAL1 induces a selective dose-related inhibitory effect on erythroid colonies, as compared with untreated or scrambled oligomer-treated control HPCs. Finally, murine and human glutathione-S- transferase (GST)-Id2 polypeptides compete the TAL1/E2A-specific DNA binding activity when added to the nuclear extracts derived from erythroid culture cells, thus indicating biochemical and suggesting functional interaction of Id2 with the TAL1/E2A complex. These novel observations indicate a coordinate expression and function of an inhibitory Id protein (Id2) and a stimulatory bHLH/bHLH heterodimer (TAL1/E2A) in normal erythroid differentiation.

Original languageEnglish
Pages (from-to)164-175
Number of pages12
JournalBlood
Volume86
Issue number1
Publication statusPublished - Jul 1 1995

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Hematopoietic Stem Cells
Messenger RNA
Hematopoiesis
Proteins
Cell culture
Oligomers
Genes
Erythroblasts
Erythroid Cells
Oligodeoxyribonucleotides
Polymerase chain reaction
RNA-Directed DNA Polymerase
Electrophoretic Mobility Shift Assay
Glutathione Transferase
Reverse Transcriptase Polymerase Chain Reaction
Assays
Suspensions
Down-Regulation
Western Blotting
Gels

ASJC Scopus subject areas

  • Hematology

Cite this

Condorelli, G., Vitelli, L., Valtieri, M., Marta, I., Montesoro, E., Lulli, V., ... Peschle, C. (1995). Coordinate expression and developmental role of Id2 protein and TAL1/E2A heterodimer in erythroid progenitor differentiation. Blood, 86(1), 164-175.

Coordinate expression and developmental role of Id2 protein and TAL1/E2A heterodimer in erythroid progenitor differentiation. / Condorelli, Gianluigi; Vitelli, Luigi; Valtieri, Mauro; Marta, Isabella; Montesoro, Elisabetta; Lulli, Valentine; Baer, Richard; Peschle, Cesare.

In: Blood, Vol. 86, No. 1, 01.07.1995, p. 164-175.

Research output: Contribution to journalArticle

Condorelli, G, Vitelli, L, Valtieri, M, Marta, I, Montesoro, E, Lulli, V, Baer, R & Peschle, C 1995, 'Coordinate expression and developmental role of Id2 protein and TAL1/E2A heterodimer in erythroid progenitor differentiation', Blood, vol. 86, no. 1, pp. 164-175.
Condorelli, Gianluigi ; Vitelli, Luigi ; Valtieri, Mauro ; Marta, Isabella ; Montesoro, Elisabetta ; Lulli, Valentine ; Baer, Richard ; Peschle, Cesare. / Coordinate expression and developmental role of Id2 protein and TAL1/E2A heterodimer in erythroid progenitor differentiation. In: Blood. 1995 ; Vol. 86, No. 1. pp. 164-175.
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N2 - The Id proteins and basic helix-loop-helix (bHLH) proteins play major roles in specifying cell fate decisions in diverse biologic settings. A potential role for Id and TAL1/E2A bHLH genes in hematopoiesis has been suggested by studies on immortalized cell lines. However, it is uncertain whether these observations reflect normal hematopoiesis. We have investigated the expression pattern of Id2 and TAL1/E2A genes in liquid suspension culture of purified hematopoietic progenitor cell (HPCs) undergoing erythroid or granulopoietic differentiation in the first culture week and maturation to terminal cells in the second week. In quiescent, freshly purified HPCs, Id2 mRNA is detected by reverse transcriptase-polymerase chain reaction (RT- PCR), whereas TAL1 and E2A mRNAs are not. At the onset of erythroid differentiation, Id2 mRNA is downregulated, while E2A and TAL1 mRNAs are concomitantly upregulated: their expression is further increased at erythroblast level. Conversely, Id2 is not downmodulated in granulopoietic culture, except for a late decline at day 10 to 12, while TAL1 and E2A are only transiently induced in the first week of granulopoietic differentiation. The expression pattern of the TAL1/E2A heterodimer, as evaluated by mobility shift assay, is consistent with RT-PCR results (except for lower levels of the heterodimer in late erythroid maturation). TAL1 protein level, analyzed by Western blot, shows a pattern consistent with gel-shift results. Functional experiments were performed on purified HPCs treated with phosphorothioate antisense oligodeoxynucleotides to Id2 or TAL1 mRNA. The results are strictly consistent with the expression studies: anti-Id2 oligomer (α-Id2) causes a significant dose-dependent increase of erythroid colony formation, whereas α-TAL1 induces a selective dose-related inhibitory effect on erythroid colonies, as compared with untreated or scrambled oligomer-treated control HPCs. Finally, murine and human glutathione-S- transferase (GST)-Id2 polypeptides compete the TAL1/E2A-specific DNA binding activity when added to the nuclear extracts derived from erythroid culture cells, thus indicating biochemical and suggesting functional interaction of Id2 with the TAL1/E2A complex. These novel observations indicate a coordinate expression and function of an inhibitory Id protein (Id2) and a stimulatory bHLH/bHLH heterodimer (TAL1/E2A) in normal erythroid differentiation.

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