Complete immunoglobulin heavy chain (Ig(H)) genes (γ and μ) containing the intronic Ig(H) enhancer and mutations in the upstream promoter region were constructed in vitro and introduced into murine J558L myeloma cells by protoplast fusion. S1-nuclease mapping experiments demonstrated that Ig(H) gene expression was extremely sensitive to mutation in an upstream region containing the octanucleotide sequence ATGCAAAT. Significant Ig(H) mRNA levels were detected in RNA from cells transfected with Ig(H) gene constructs in which all upstream sequences on the 5' proximal side of this element were deleted. Similar results were obtained using the precise inverse of the Ig(H) octamer, which is found in the upstream promoter region of immunoglobulin light chain genes. Deletion of the Ig(H) octamer, or point mutation of adenine to guanine at position 6, resulted in the loss of correctly initiated Ig(H) mRNA. A DNA binding factor from J558L nuclear extracts was identified that appeared to recognize the octamer on the basis of differential binding to homologous restriction fragments containing the various mutations and that bound preferentially with octamer DNA fragments derived from functional relative to nonfunctional Ig(H) constructs. Collectively, these data suggest that the octamer element contains residues that are critical to accurate immunoglobulin gene transcription and that may serve as part of a recognition locus for nuclear factors important to B-cell-specific immunoglobulin expression.
|Original language||English (US)|
|Number of pages||5|
|Journal||Proceedings of the National Academy of Sciences of the United States of America|
|State||Published - 1986|
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