Characterization of cis-acting elements of the gene for macrophage- stimulating protein from the human. The involvement of positive and negative regulatory elements

Atsuhisa Ueda, Teizo Yoshimura

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

To analyze the promotor region of the human macrophage-stimulating protein (MSP) gene, the 5'-flanking region of this gene was cloned. The major initiation site was determined at T located 49 base pairs upstream of the translation initiation site by primer extention with mRNA from HepG2 and Hep3B cells. There was no TATA sequence in this region. Transient transfection assay with 5'-deletion constructs showed that the transcription of this gene was regulated by positive and negative regulatory elements (PRE and NRE). The PRE (-34 to +2) was essential for the maximal transcription of this gene, and the NRE (-141 to -34) appeared to be responsible for the tissue-specific expression of the gene. The PRE contained the CCAAT sequence and a mutation from CCAAT to CTGAT resulted in a significant loss of the transcriptional activity. Electrophoretic mobility shift assay suggested that two different proteins bound to the PRE (MSP-PRE-binding protein-1 (MSP- PREB1) and 2). MSP-PREB1 and 2 were detected in various cell types, and the CCAAT sequence was involved in these bindings. These findings indicate that MSP-PREB1 and 2 are positive regulators. Further characterization also revealed that MSP-PREB2 was identical to CCAAT binding transcription factor, also known as NF-Y.

Original languageEnglish
Pages (from-to)20265-20272
Number of pages8
JournalJournal of Biological Chemistry
Volume271
Issue number34
DOIs
Publication statusPublished - 1996
Externally publishedYes

Fingerprint

Genes
Transcription
Assays
CCAAT-Binding Factor
Electrophoretic mobility
5' Flanking Region
Hep G2 Cells
Electrophoretic Mobility Shift Assay
Genetic Promoter Regions
Protein Binding
Base Pairing
Transfection
Carrier Proteins
macrophage stimulating protein
Tissue
Gene Expression
Messenger RNA
Mutation
Proteins

ASJC Scopus subject areas

  • Biochemistry

Cite this

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abstract = "To analyze the promotor region of the human macrophage-stimulating protein (MSP) gene, the 5'-flanking region of this gene was cloned. The major initiation site was determined at T located 49 base pairs upstream of the translation initiation site by primer extention with mRNA from HepG2 and Hep3B cells. There was no TATA sequence in this region. Transient transfection assay with 5'-deletion constructs showed that the transcription of this gene was regulated by positive and negative regulatory elements (PRE and NRE). The PRE (-34 to +2) was essential for the maximal transcription of this gene, and the NRE (-141 to -34) appeared to be responsible for the tissue-specific expression of the gene. The PRE contained the CCAAT sequence and a mutation from CCAAT to CTGAT resulted in a significant loss of the transcriptional activity. Electrophoretic mobility shift assay suggested that two different proteins bound to the PRE (MSP-PRE-binding protein-1 (MSP- PREB1) and 2). MSP-PREB1 and 2 were detected in various cell types, and the CCAAT sequence was involved in these bindings. These findings indicate that MSP-PREB1 and 2 are positive regulators. Further characterization also revealed that MSP-PREB2 was identical to CCAAT binding transcription factor, also known as NF-Y.",
author = "Atsuhisa Ueda and Teizo Yoshimura",
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AU - Ueda, Atsuhisa

AU - Yoshimura, Teizo

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AB - To analyze the promotor region of the human macrophage-stimulating protein (MSP) gene, the 5'-flanking region of this gene was cloned. The major initiation site was determined at T located 49 base pairs upstream of the translation initiation site by primer extention with mRNA from HepG2 and Hep3B cells. There was no TATA sequence in this region. Transient transfection assay with 5'-deletion constructs showed that the transcription of this gene was regulated by positive and negative regulatory elements (PRE and NRE). The PRE (-34 to +2) was essential for the maximal transcription of this gene, and the NRE (-141 to -34) appeared to be responsible for the tissue-specific expression of the gene. The PRE contained the CCAAT sequence and a mutation from CCAAT to CTGAT resulted in a significant loss of the transcriptional activity. Electrophoretic mobility shift assay suggested that two different proteins bound to the PRE (MSP-PRE-binding protein-1 (MSP- PREB1) and 2). MSP-PREB1 and 2 were detected in various cell types, and the CCAAT sequence was involved in these bindings. These findings indicate that MSP-PREB1 and 2 are positive regulators. Further characterization also revealed that MSP-PREB2 was identical to CCAAT binding transcription factor, also known as NF-Y.

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