Identification of a TAAT-containing motif required for high level expression of the COL1A1 promoter in differentiated osteoblasts of transgenic mice

被引:78
作者
Dodig, M
Kronenberg, MS
Bedalov, A
Kream, BE
Gronowicz, G
Clark, SH
Mack, K
Liu, YH
Maxon, R
Pan, ZZ
Upholt, WB
Rowe, DW
Lichtler, AC
机构
[1] UNIV CONNECTICUT,CTR HLTH,DEPT PEDIAT,FARMINGTON,CT 06030
[2] UNIV CONNECTICUT,CTR HLTH,DEPT ORTHOPAED SURG,FARMINGTON,CT 06030
[3] UNIV CONNECTICUT,CTR HLTH,DEPT BIOSTRUCT & FUNCT,FARMINGTON,CT 06030
[4] UNIV CONNECTICUT,CTR HLTH,DEPT MED,DIV RHEUMAT DIS,FARMINGTON,CT 06030
[5] DEPT VET AFFAIRS MED CTR,NEWINGTON,CT 06111
[6] UNIV SO CALIF,SCH MED,KENNETH R NORRIS HOSP & INST,DEPT BIOCHEM & MOLEC BIOL,LOS ANGELES,CA 90033
关键词
D O I
10.1074/jbc.271.27.16422
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Our previous studies have shown that the 49-base pair region of promoter DNA between -1719 and -1670 base pairs is necessary for transcription of the rat COL1A1 gene in transgenic mouse calvariae, In this study, we further define this element to the 13-base pair region between -1683 and -1670. This element contains a TAAT motif that binds homeodomain-containing proteins, Site-directed mutagenesis of this element in the context of a COL1A1-chloramphenicol acetyltransferase construct extending to -3518 base pairs decreased the ratio of reporter gene activity in calvariae to tendon from 3:1 to 1:1, suggesting a preferential effect on activity in calvariae, Moreover, chloramphenicol acetyltransferase-specific immunofluorescence microscopy of transgenic calvariae showed that the mutation preferentially reduced levels of chloramphenicol acetyltransferase-protein in differentiated osteoblasts, Gel mobility shift assays demonstrate that differentiated osteoblasts contain a nuclear factor that binds to this site, This binding activity is not present in undifferentiated osteoblasts, Ne show that Msx2, a homeodomain protein, binds to this motif; however, Northern blot analysis revealed that Msx2 mRNA is present in undifferentiated bone cells but not in fully differentiated osteoblasts, In addition, cotransfection studies in ROS 17/2.8 osteosarcoma cells using an Msx2 expression vector showed that Msx2 inhibits a COL1A1 promoter-chloramphenicol acetyltransferase construct, Our results suggest that high COL1A1 expression in bone is mediated by a protein that is induced during osteoblast differentiation. This protein may contain a homeodomain; however, it is distinct from homeodomain proteins reported previously to be present in bone.
引用
收藏
页码:16422 / 16429
页数:8
相关论文
共 50 条
  • [1] BEDALOV A, 1995, J BONE MINER RES, V10, P1443
  • [2] GENOMIC STRUCTURE, CHROMOSOMAL LOCATION, AND EVOLUTION OF THE MOUSE HOX-8 GENE
    BELL, JR
    NOVEEN, A
    LIU, YH
    MA, L
    DOBIAS, S
    KUNDU, R
    LUO, W
    XIA, YR
    LUSIS, AJ
    SNEAD, ML
    MAXSON, R
    [J]. GENOMICS, 1993, 16 (01) : 123 - 131
  • [3] CHANGES IN INSULIN-LIKE GROWTH FACTOR-BINDING PROTEIN EXPRESSION AND SECRETION DURING THE PROLIFERATION, DIFFERENTIATION, AND MINERALIZATION OF PRIMARY CULTURES OF RAT OSTEOBLASTS
    BIRNBAUM, RS
    WIREN, KM
    [J]. ENDOCRINOLOGY, 1994, 135 (01) : 223 - 230
  • [4] BOGDANOVIC Z, 1994, J BONE MINER RES, V9, P285
  • [5] NUCLEOTIDES FLANKING A CONSERVED TAAT CORE DICTATE THE DNA-BINDING SPECIFICITY OF 3 MURINE HOMEODOMAIN PROTEINS
    CATRON, KM
    ILER, N
    ABATE, C
    [J]. MOLECULAR AND CELLULAR BIOLOGY, 1993, 13 (04) : 2354 - 2365
  • [6] ISOLATION OF THE HUMAN-GENE FOR BONE GLA PROTEIN UTILIZING MOUSE AND RAT CDNA CLONES
    CELESTE, AJ
    ROSEN, V
    BUECKER, JL
    KRIZ, R
    WANG, EA
    WOZNEY, JM
    [J]. EMBO JOURNAL, 1986, 5 (08) : 1885 - 1890
  • [7] LOCALIZATION OF BONE SIALOPROTEIN (BSP) EXPRESSION TO SITES OF MINERALIZED TISSUE FORMATION IN FETAL-RAT TISSUES BY INSITU HYBRIDIZATION
    CHEN, JK
    SHAPIRO, HS
    WRANA, JL
    REIMERS, S
    HEERSCHE, JNM
    SODEK, J
    [J]. MATRIX, 1991, 11 (02): : 133 - 143
  • [8] CHOMCZYNSKI P, 1993, BIOTECHNIQUES, V15, P532
  • [9] CSERJESI P, 1992, DEVELOPMENT, V115, P1087
  • [10] ACCURATE TRANSCRIPTION INITIATION BY RNA POLYMERASE-II IN A SOLUBLE EXTRACT FROM ISOLATED MAMMALIAN NUCLEI
    DIGNAM, JD
    LEBOVITZ, RM
    ROEDER, RG
    [J]. NUCLEIC ACIDS RESEARCH, 1983, 11 (05) : 1475 - 1489