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Molecular cloning, sequence analysis, and functional expression of a novel growth regulator, oncostatin M
N Malik1, J C Kallestad, N L Gunderson
1Oncogen, Seattle, Washington 98121.
Abstract:
Oncostatin M is a polypeptide of Mr approximately 28,000 that acts as a growth regulator for many cultured mammalian cells. We report the cDNA and genomic cloning, sequence analysis, and functional expression in heterologous cells of oncostatin M. cDNA clones were isolated from mRNA of U937 cells that had been induced to differentiate into macrophagelike cells by treatment with phorbol 12-myristate 13-acetate, and a genomic clone was also isolated from human brain DNA. Sequence analysis of these clones established the 1,814-base-pair cDNA sequence as well as exon boundaries. This sequence predicted that oncostatin M is synthesized as a 252-amino-acid polypeptide, with a 25-residue hydrophobic sequence resembling a signal peptide at the N terminus. The predicted oncostatin M amino acid sequence shared no homology with other known proteins, but the sequence of the 3' noncoding region of the cDNA contained an A + T-rich stretch with sequence motifs found in the 3' untranslated regions of many cytokine and lymphokine cDNAs. Oncostatin M mRNA of approximately 2 kilobase pairs was detected in phorbol 12-myristate 13-acetate-treated U937 cells and in activated human T cells. Transfection of cDNA encoding the oncostatin M precursor into COS cells resulted in the secretion of proteins with the structural and functional properties of oncostatin M. The unique amino acid sequence, expression by lymphoid cells, and growth-regulatory activities of oncostatin M suggest that it is a novel cytokine.
Insights
Oncostatin M, a novel cytokine, regulates mammalian cell growth. Researchers cloned its cDNA and genomic DNA, revealing its unique amino acid sequence and function when expressed in other cells.
Area of Science:
- Molecular Biology
- Cell Biology
- Immunology
Background:
- Oncostatin M (M) is a polypeptide involved in regulating mammalian cell growth.
- Its precise molecular characteristics and genetic basis were previously uncharacterized.
Purpose of the Study:
- To clone and sequence the cDNA and genomic DNA of Oncostatin M.
- To analyze its predicted amino acid sequence and functional expression.
Main Methods:
- cDNA library screening from U937 cells induced with phorbol 12-myristate 13-acetate.
- Genomic DNA isolation and cloning from human brain.
- Sequence analysis of cDNA and genomic clones.
- Functional expression in COS cells via cDNA transfection.
Main Results:
- Established the 1,814-bp cDNA sequence and exon boundaries of Oncostatin M.
- Predicted a 252-amino-acid precursor with a signal peptide; no homology to known proteins.
- Detected Oncostatin M mRNA in treated U937 cells and activated T cells.
- Demonstrated secretion of functional Oncostatin M from transfected COS cells.
Conclusions:
- Oncostatin M possesses a unique amino acid sequence and is expressed by lymphoid cells.
- Its growth-regulatory activities suggest it is a novel cytokine.
- The cloning and expression data provide a foundation for further research into Oncostatin M's biological roles.