Negative transcriptional regulation of connexin 43 by Tbx2 in rat immature coronal sutures and ROS 17/2.8 cells in culture.
BACKGROUND
Tbx2 is a member of the T-box family of transcriptional regulatory genes with an extensive but not yet fully understood role in embryonic development. This study explores the potential role of Tbx2 in calvarial morphogenesis.


OBJECTIVE
To explore the hypothesis that Tbx2 has a negative regulatory effect on the expression of connexin 43 (Cx43), a protein necessary for cell-to-cell communication; document the presence of Tbx2 protein in the developing cranial sutures; and determine the spatial pattern of expression of this developmentally regulated transcription factor in calvariae.


METHODS
The osteoblast-like cell line ROS 17/2.8 was stably transfected with sense or antisense Tbx2. Immunohistochemistry and Western blotting was used to study Tbx2 and Cx43 expression in these cells and sections of embedded developing coronal sutures.


RESULTS
The ROS 17/2.8 cells transfected with antisense Tbx2 showed a decrease in expression of Tbx2 protein and an increase in expression of endogenous Cx43. The reverse is seen with sense-transfected cells. Both of these proteins are expressed in rat developing coronal sutures. The pattern of Tbx2 expression in the developing was also reciprocal to the pattern of Cx43 expression. Tbx2 protein is concentrated in the center of the sutural blastema, an area devoid of Cx43 protein localization. Conversely, Tbx2 protein expression is low in the periphery of the sutures, in which there is high Cx43 protein expression.


CONCLUSIONS
Taken together, these studies suggest that Tbx2 protein is a negative regulator of Cx43 expression at the transcriptional level in cranial sutures in vivo.
Department of Oral Biology and Maxillofacial Pathology, Medical College of Georgia, School of Dentistry, Augusta, GA, USA.
James L
JL
Borke
Jung-Ren
JR
Chen
Jack C
JC
Yu
Roni J
RJ
Bollag
Maria F
MF
Orellana
Carlos M
CM
Isales
D000818:Animals; D000906:Antibodies; D015153:Blotting, Western; D018031:Connexin 43 / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D003393:Cranial Sutures / Q000737:chemistry / Q000378:metabolism*; D018507:Gene Expression Regulation, Developmental; D007124:Immunoenzyme Techniques; D010006:Osteoblasts / Q000378:metabolism; D012516:Osteosarcoma; D051381:Rats; D020825:T-Box Domain Proteins / Q000096:biosynthesis*; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured
D000906:Antibodies; D018031:Connexin 43; C097348:T-Box Domain Protein 2; D020825:T-Box Domain Proteins