Mpl ligand enhances the transcription of the cyclin D3 gene: A potential role for Sp1 transcription factor

Zack Z. Wang, Ying Zhang, Jun Lu, Shinnshin Sun, Katya Ravid

Research output: Contribution to journalArticle

Abstract

Cyclin D3 plays a major role in the development of polyploidy in megakaryocytes. The expression of cyclin D3 gene and the level of cyclin D3 protein are increased by the Mpl ligand in the Y10/L8057 megakaryocytic cell line, as indicated by Northern and Western blot analyses, and by nuclear run- on assays and transfection experiments with cyclin D3 promoter constructs. DNase I footprinting of the promoter region showed protected segments, at - 75 to -60 bp and at -134 to -92 bp, which display binding sites for the Sp family of transcription factors. Gel mobility shift assay and supershifts with specific antibodies indicate that Sp1 binds to these regions in the cyclin D3 promoter and that Sp1 binding activity is significantly increased by Mpl ligand. Mutation of either Sp1 site both decreases the basal promoter activity and eliminates the induction by Mpl ligand. We find that the nonphosphorylated form of SP1 has greater affinity for the cyclin D3 promoter and that the majority of Sp1 in the cells is nonphosphorylated. Mpl ligand treatment results in increased levels of Sp1 protein, which also appears as nonphosphorylated. Okadaic acid, which inhibits protein phosphatase 1 (PP1) and shifts Sp1 to a phosphorylated form, decreases cyclin D3 gene expression and suppresses Mpl ligand induction. Our data point to the potential of Mpl ligand to activate at once several Sp1-dependent genes during megakaryopoiesis.

Original languageEnglish (US)
Pages (from-to)4208-4221
Number of pages14
JournalBlood
Volume93
Issue number12
StatePublished - Jun 15 1999
Externally publishedYes

Fingerprint

Cyclin D3
Sp1 Transcription Factor
Thrombopoietin
Transcription
Genes
Assays
Sp Transcription Factors
Protein Phosphatase 1
Okadaic Acid
Polyploidy
Megakaryocytes
Deoxyribonuclease I
Electrophoretic Mobility Shift Assay
Genetic Promoter Regions
Gene expression
Northern Blotting
Transfection
Proteins
Transcription Factors
Western Blotting

ASJC Scopus subject areas

  • Hematology

Cite this

Mpl ligand enhances the transcription of the cyclin D3 gene : A potential role for Sp1 transcription factor. / Wang, Zack Z.; Zhang, Ying; Lu, Jun; Sun, Shinnshin; Ravid, Katya.

In: Blood, Vol. 93, No. 12, 15.06.1999, p. 4208-4221.

Research output: Contribution to journalArticle

Wang, ZZ, Zhang, Y, Lu, J, Sun, S & Ravid, K 1999, 'Mpl ligand enhances the transcription of the cyclin D3 gene: A potential role for Sp1 transcription factor', Blood, vol. 93, no. 12, pp. 4208-4221.
Wang, Zack Z. ; Zhang, Ying ; Lu, Jun ; Sun, Shinnshin ; Ravid, Katya. / Mpl ligand enhances the transcription of the cyclin D3 gene : A potential role for Sp1 transcription factor. In: Blood. 1999 ; Vol. 93, No. 12. pp. 4208-4221.
@article{d2a4d09466a946ab9955c0bce787019e,
title = "Mpl ligand enhances the transcription of the cyclin D3 gene: A potential role for Sp1 transcription factor",
abstract = "Cyclin D3 plays a major role in the development of polyploidy in megakaryocytes. The expression of cyclin D3 gene and the level of cyclin D3 protein are increased by the Mpl ligand in the Y10/L8057 megakaryocytic cell line, as indicated by Northern and Western blot analyses, and by nuclear run- on assays and transfection experiments with cyclin D3 promoter constructs. DNase I footprinting of the promoter region showed protected segments, at - 75 to -60 bp and at -134 to -92 bp, which display binding sites for the Sp family of transcription factors. Gel mobility shift assay and supershifts with specific antibodies indicate that Sp1 binds to these regions in the cyclin D3 promoter and that Sp1 binding activity is significantly increased by Mpl ligand. Mutation of either Sp1 site both decreases the basal promoter activity and eliminates the induction by Mpl ligand. We find that the nonphosphorylated form of SP1 has greater affinity for the cyclin D3 promoter and that the majority of Sp1 in the cells is nonphosphorylated. Mpl ligand treatment results in increased levels of Sp1 protein, which also appears as nonphosphorylated. Okadaic acid, which inhibits protein phosphatase 1 (PP1) and shifts Sp1 to a phosphorylated form, decreases cyclin D3 gene expression and suppresses Mpl ligand induction. Our data point to the potential of Mpl ligand to activate at once several Sp1-dependent genes during megakaryopoiesis.",
author = "Wang, {Zack Z.} and Ying Zhang and Jun Lu and Shinnshin Sun and Katya Ravid",
year = "1999",
month = "6",
day = "15",
language = "English (US)",
volume = "93",
pages = "4208--4221",
journal = "Blood",
issn = "0006-4971",
publisher = "American Society of Hematology",
number = "12",

}

TY - JOUR

T1 - Mpl ligand enhances the transcription of the cyclin D3 gene

T2 - A potential role for Sp1 transcription factor

AU - Wang, Zack Z.

AU - Zhang, Ying

AU - Lu, Jun

AU - Sun, Shinnshin

AU - Ravid, Katya

PY - 1999/6/15

Y1 - 1999/6/15

N2 - Cyclin D3 plays a major role in the development of polyploidy in megakaryocytes. The expression of cyclin D3 gene and the level of cyclin D3 protein are increased by the Mpl ligand in the Y10/L8057 megakaryocytic cell line, as indicated by Northern and Western blot analyses, and by nuclear run- on assays and transfection experiments with cyclin D3 promoter constructs. DNase I footprinting of the promoter region showed protected segments, at - 75 to -60 bp and at -134 to -92 bp, which display binding sites for the Sp family of transcription factors. Gel mobility shift assay and supershifts with specific antibodies indicate that Sp1 binds to these regions in the cyclin D3 promoter and that Sp1 binding activity is significantly increased by Mpl ligand. Mutation of either Sp1 site both decreases the basal promoter activity and eliminates the induction by Mpl ligand. We find that the nonphosphorylated form of SP1 has greater affinity for the cyclin D3 promoter and that the majority of Sp1 in the cells is nonphosphorylated. Mpl ligand treatment results in increased levels of Sp1 protein, which also appears as nonphosphorylated. Okadaic acid, which inhibits protein phosphatase 1 (PP1) and shifts Sp1 to a phosphorylated form, decreases cyclin D3 gene expression and suppresses Mpl ligand induction. Our data point to the potential of Mpl ligand to activate at once several Sp1-dependent genes during megakaryopoiesis.

AB - Cyclin D3 plays a major role in the development of polyploidy in megakaryocytes. The expression of cyclin D3 gene and the level of cyclin D3 protein are increased by the Mpl ligand in the Y10/L8057 megakaryocytic cell line, as indicated by Northern and Western blot analyses, and by nuclear run- on assays and transfection experiments with cyclin D3 promoter constructs. DNase I footprinting of the promoter region showed protected segments, at - 75 to -60 bp and at -134 to -92 bp, which display binding sites for the Sp family of transcription factors. Gel mobility shift assay and supershifts with specific antibodies indicate that Sp1 binds to these regions in the cyclin D3 promoter and that Sp1 binding activity is significantly increased by Mpl ligand. Mutation of either Sp1 site both decreases the basal promoter activity and eliminates the induction by Mpl ligand. We find that the nonphosphorylated form of SP1 has greater affinity for the cyclin D3 promoter and that the majority of Sp1 in the cells is nonphosphorylated. Mpl ligand treatment results in increased levels of Sp1 protein, which also appears as nonphosphorylated. Okadaic acid, which inhibits protein phosphatase 1 (PP1) and shifts Sp1 to a phosphorylated form, decreases cyclin D3 gene expression and suppresses Mpl ligand induction. Our data point to the potential of Mpl ligand to activate at once several Sp1-dependent genes during megakaryopoiesis.

UR - http://www.scopus.com/inward/record.url?scp=0033564952&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0033564952&partnerID=8YFLogxK

M3 - Article

C2 - 10361118

AN - SCOPUS:0033564952

VL - 93

SP - 4208

EP - 4221

JO - Blood

JF - Blood

SN - 0006-4971

IS - 12

ER -