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Characterization of Spi-B, a transcription factor related to the putative oncoprotein Spi-1/PU.1
D Ray1, R Bosselut, J Ghysdael
1Faculté de Médecine Lariboisière Saint-Louis, INSERM U-248, Paris, France.
Abstract:
We have cloned a human cDNA from a new gene, spi-B, on the basis of its homology with the DNA-binding domain of the Spi-1/PU.1 putative oncogene product. spi-B codes for a protein of 262 amino acids presenting 43% overall identity with Spi-1. Its highly basic carboxy-terminal region exhibits 34% sequence identity with the DNA-binding domain of the Ets-1 protein. We showed that the Spi-B protein is able to bind the purine-rich sequence (PU box) recognized by Spi-1/PU.1 and to activate transcription of a reporter plasmid containing PU boxes. Chromosome in situ hybridization allowed us to map spi-B to the 19q13.3-19q13.4 region of the human genome. spi-B, like spi-1, was found to be expressed in various murine and human hematopoietic cell lines except T lymphoid cell lines.
Insights
Researchers identified a new gene, spi-B, which codes for a protein similar to Spi-1/PU.1. This Spi-B protein binds DNA and activates transcription, with expression found in hematopoietic cells.
Area of Science:
- Molecular Biology
- Genetics
- Oncology
Background:
- The Spi-1/PU.1 gene product is a putative oncogene involved in hematopoietic regulation.
- Homology searches identified a novel gene, spi-B, related to Spi-1/PU.1.
Purpose of the Study:
- To clone and characterize the newly identified spi-B gene and its protein product.
- To investigate the DNA-binding and transcriptional activation capabilities of the Spi-B protein.
- To determine the genomic location and expression pattern of the spi-B gene.
Main Methods:
- Human cDNA cloning based on homology to Spi-1/PU.1 DNA-binding domain.
- Amino acid sequence analysis and comparison with Spi-1 and Ets-1.
- DNA-binding assays using PU box sequences.
- Reporter gene assays to assess transcriptional activation.
- Chromosome in situ hybridization for gene mapping.
- Analysis of gene expression in various cell lines.
Main Results:
- A human cDNA for spi-B was cloned, encoding a 262-amino acid protein with 43% identity to Spi-1.
- The Spi-B protein shares sequence identity in its DNA-binding domain with Ets-1.
- Spi-B protein demonstrated binding to PU boxes and activated transcription.
- The spi-B gene was mapped to chromosome region 19q13.3-19q13.4.
- Spi-B expression was detected in murine and human hematopoietic cell lines, excluding T lymphoid cells.
Conclusions:
- Spi-B represents a novel gene encoding a transcription factor with DNA-binding and activation functions similar to Spi-1/PU.1.
- The characterization of Spi-B provides insights into the regulation of hematopoietic cell differentiation.
- The genomic location and expression profile of Spi-B suggest its potential role in specific hematopoietic lineages.