Comparative analysis of two alternative first exons reported for the mouse osteopontin gene.
Two conflicting conclusions regarding the structure of the mouse osteopontin (OPN) gene were tested for their validity. Miyazaki et al. (Miyazaki, Y., Setoguchi, M., Yoshida, S., Higuchi, Y., Akizuki, S., and Yamamoto, S. (1990) J. Biol. Chem. 265, 14432-14438) state that the OPN gene is composed of six exons and spans approximately 4.8 kilobases. Craig and Denhardt (Craig, A. M., and Denhardt, D. T. (1991) Gene (Amst.) 100, 163-171) independently reported an additional exon 5' to the region designated as "exon 1" by Miyazaki and colleagues. To investigate this discrepancy, we generated oligodeoxynucleotide probes to three regions of these reported sequences and used them to hybridize to Northern and Southern blots of RNA and DNA from mouse fibroblasts and macrophages. Two of these regions (probes "A" and "B") represent sequences that are disputed, while one of these regions (probe "C") is predicted to be in the mRNA of both sequences. Our results are consistent only with the OPN gene structure reported by Craig and Denhardt and show that a significant portion of the exon 1 reported by Miyazaki et al. is present in murine genomic DNA but is not found in cytoplasmic message. Our results also show that OPN RNA molecules from mouse fibroblasts and macrophages do not differ significantly in the regions tested.
Behrend
E I
EI
London Regional Cancer Centre, Ontario, Canada.
Chambers
A F
AF
Wilson
S M
SM
Denhardt
D T
DT
D016475:3T3 Cells; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D015139:Blotting, Southern; D002460:Cell Line; D004247:DNA / Q000235:genetics / Q000302:isolation & purification; D005091:Exons; D005347:Fibroblasts / Q000502:physiology; D011905:Genes, ras; D006801:Humans; D008264:Macrophages / Q000502:physiology; D008545:Melanoma; D051379:Mice; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D015345:Oligonucleotide Probes; D053495:Osteopontin; D010750:Phosphoproteins / Q000235:genetics*; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D012795:Sialoglycoproteins / Q000235:genetics*; D014162:Transfection; D014407:Tumor Cells, Cultured
D009838:Oligodeoxyribonucleotides; D015345:Oligonucleotide Probes; D010750:Phosphoproteins; C506895:SPP1 protein, human; D012795:Sialoglycoproteins; C506896:Spp1 protein, mouse; D053495:Osteopontin; D012313:RNA; D004247:DNA