Regulation of Tcf7 by Runx2 in chondrocyte maturation and proliferation.
Runx2 plays important roles in the regulation of chondrocyte differentiation and proliferation; however, the Runx2 target molecules still remain to be investigated. We searched the genes upregulated by the introduction of Runx2 into Runx2(-/-) chondrocytes using microarray and found that Tcf7 is upregulated by Runx2. Thus, we examined the functions of Runx2 in the regulation of the Tcf/Lef family of transcription factors. Runx2 induced Tcf7 and Lef1 strongly, but Tcf7l1 and Tcf7l2 only slightly in Runx2(-/-) chondrocytes; the expressions of Tcf7 and Tcf7l2 were reduced in Runx2(-/-) cartilaginous skeletons and calvaria, and Tcf7 showed a similar expression pattern to Runx2. In reporter assays, Runx2 mildly activated the 8.6 and 1.8 kb Tcf7 promoter constructs. The reporter assays using the deletion constructs of the 1.8-kb fragment showed that the 0.3-kb promoter region is responsible for the Runx2-dependent transcriptional activation. To investigate the function of Tcf7 in skeletal development, we generated dominant-negative (dn) Tcf7 transgenic mice using the Col2a1 promoter. Dn-Tcf7 transgenic embryos showed dwarfism, and mineralization was retarded in limbs, ribs, and vertebrae in a manner dependent on the expression levels of the transgene. In situ hybridization analysis showed that endochondral ossification is retarded in dn-Tcf7 transgenic embryos due to the decelerated chondrocyte maturation. Further, BrdU labeling showed a reduction in chondrocyte proliferation in the proliferating layer of the growth plate in dn-Tcf7 transgenic embryos. These findings indicate that Runx2 regulates chondrocyte maturation and proliferation at least partly through the induction of Tcf7.
Mikasa
Masaki
M
Department of Cell Biology, Unit of Basic Medical Sciences, Nagasaki University Graduate School of Biomedical Sciences, 1-7-1 Sakamoto, Nagasaki 852-8588, Japan.
Rokutanda
Satoshi
S
Komori
Hisato
H
Ito
Kosei
K
Tsang
Ying Sze
YS
Date
Yuki
Y
Yoshida
Carolina A
CA
Komori
Toshihisa
T
D000818:Animals; D001483:Base Sequence; D001842:Bone and Bones / Q000378:metabolism / Q000473:pathology; D001973:Bromodeoxyuridine / Q000378:metabolism; D002454:Cell Differentiation / Q000235:genetics*; D049109:Cell Proliferation; D019902:Chondrocytes / Q000378:metabolism* / Q000473:pathology*; D047369:Chromatin Immunoprecipitation; D050659:Core Binding Factor Alpha 1 Subunit / Q000378:metabolism*; D005121:Extremities / Q000473:pathology; D005786:Gene Expression Regulation; D017930:Genes, Reporter / Q000235:genetics; D051538:Hepatocyte Nuclear Factor 1-alpha; D051379:Mice; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D010012:Osteogenesis / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics; D051822:T Cell Transcription Factor 1 / Q000235:genetics* / Q000378:metabolism
D050659:Core Binding Factor Alpha 1 Subunit; D051538:Hepatocyte Nuclear Factor 1-alpha; C496488:Hnf1a protein, mouse; C493716:Runx2 protein, mouse; D051822:T Cell Transcription Factor 1; D001973:Bromodeoxyuridine