Osteonectin promoter. DNA sequence analysis and S1 endonuclease site potentially associated with transcriptional control in bone cells.
To understand the basis of osteonectin (SPARC) transcriptional regulation, we have isolated a bovine genomic clone (lambda Og15) encoding exon 1 and 15 kilobase pairs (kb) of flanking DNA. Direct RNA sequencing of the 5' end of the osteonectin message showed it contained a sequence identical to that of a 2.4-kb EcoRI-BamHI fragment located midway in the clone lambda Og15. The results indicate exon 1 is located 10 kb away from exon 2 in the bovine genome. The DNA sequence unit CCTG is repeated five times in exon 1 which is composed exclusively of untranslated sequence. Sequence analysis of the 5'-flanking DNA revealed the presence of many regulatory motifs including a "GC" box with four overlapping SP1 consensus sequences. Immediately downstream from the GC box is a 72-base pair purine-rich stretch composed primarily of direct repeats of the sequence motifs GGGGA and GGA (GAGA box). Digestion of the flanking DNA in vitro with S1 endonuclease showed a site for the enzyme at position -55 which is just 3' to the GAGA box. Chimeric chloramphenicol acetyltransferase constructs were prepared containing the S1-sensitive site and showed substantial transcriptional activity in UMR-106 and fetal and adult human bone cells which are known to be high producers of the protein. The results indicate a potential regulatory activity of the S1 site in osteonectin gene activation.
Young
M F
MF
Bone Research Branch, National Institute of Dental Research, Bethesda, Maryland 20892.
Findlay
D M
DM
Dominguez
P
P
Burbelo
P D
PD
McQuillan
C
C
Kopp
J B
JB
Robey
P G
PG
Termine
J D
JD
D000818:Animals; D001483:Base Sequence; D002352:Carrier Proteins / Q000235:genetics*; D002417:Cattle; D002460:Cell Line; D004247:DNA / Q000235:genetics; D004720:Endonucleases; D005786:Gene Expression Regulation; D005796:Genes; D008099:Liver / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000235:genetics*; D008969:Molecular Sequence Data; D015676:Osteonectin; D011401:Promoter Regions, Genetic; D015183:Restriction Mapping; D015719:Single-Strand Specific DNA and RNA Endonucleases; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection
D002352:Carrier Proteins; D008562:Membrane Glycoproteins; D015676:Osteonectin; D004247:DNA; D004720:Endonucleases; D015719:Single-Strand Specific DNA and RNA Endonucleases