The structure and expression of the murine gene encoding granulocyte-macrophage colony stimulating factor: evidence for utilisation of alternative promoters.

Abstract

Two overlapping genomic clones containing the murine granulocyte‐macrophage colony stimulating factor (GM‐CSF) gene have been isolated. On the basis of transfection experiments, we have established that a 9‐kb BamHI fragment from one of these recombinants encodes biologically active GM‐CSF. As deduced from nucleotide sequence analysis, the GM‐CSF gene comprises four exons encompassing 2.5 kb of genomic DNA. Primer extension analysis of GM‐CSF mRNA identifies a transcriptional initiation site 35 bp upstream of a single translational initiation codon in‐frame with the GM‐CSF coding sequences and 28 bp downstream of a TATA promoter consensus sequence. Pre‐GM‐CSF molecules encoded by mRNAs originating from this promoter would include a hydrophobic leader sequence typical for a secreted protein. Intriguingly, sequences present at the 5′ end of a GM‐CSF cDNA clone previously isolated in our laboratory are not contained within either of the genomic clones and must therefore be transcribed from a promoter located at least 10 kb 5′ of the main body of the gene. mRNAs transcribed from this alternative upstream promoter possess an additional initiating codon and potentially encode a pre‐GM‐CSF polypeptide with an atypical NH2‐terminal leader peptide. Comparison of the nucleotide sequence of the GM‐CSF gene with that of other haemopoietic growth factor genes has revealed a common decanucleotide (5′‐GPuGPuTTPyCAPy‐3′) within their respective 5′‐flanking regions which may be involved in their co‐ordinate regulation.