Nuclear matrix architectural transcription factors that contribute to the regulation of the alpha1(I) collagen promoter in osteoblasts

In connective tissue, cell structure contributes to type 1 collagen expression, the major protein found in bone. An increase in collagen expression is an indicator of bone formation. Parathyroid hormone (PTH) increases bone formation when administered intermittently.;The tissue matrix may couple the changes in osteoblast shape to collagen expression. The tissue matrix is the dynamic structural linkages between the extracellular matrix, the cytoplasm, and the nuclear matrix (NM), which connects DNA to the cell periphery. Architectural transcription factors bind to the minor groove of AT-rich DNA and bend it, altering interactions between other trans-acting proteins. Similarly, NM proteins bind to the minor groove of AT-rich matrix attachment regions regulating transcription by altering DNA structure. We therefore propose that PTH-responsive osteoblast NM architectural transcription factors link cell structure to the type I collagen (alpha1(I) (COL1A1) polypeptide chain promoter geometry and COL1A1 transcription.;Our objectives were (i) to identify sequence-specific, PTH-responsive, NM protein interactions along the COL1A1 promoter, (ii) to characterize the NM protein-DNA interactions biochemically, and (iii) to determine the functional significance of these binding sites. NM and soluble nuclear proteins were obtained as separate subfractions from rat bone and rat osteosarcoma cells (ROS 17/2.8 and UMR 106-01). Electrophoretic mobility shift analysis (EMSA) was used to identify COL1A1-protein interactions in the nuclear subfractions. NMP3 is a NM protein-COL1A1 interaction between -2149 and -2106 nt. NP/NMP4 is a NM protein-COL1A1 complex observed at -3469/-3450 nt, site A, and -1574/-1555 nt, site B. PTH treatment of ROS 17/2.8 cells increased NP/NMP4-COL1A1 binding, but had no effect on NMP3-COL1A1 binding. Biochemical analyses revealed that NP/NMP4 binding to the COL1A1 promoter localizes to the minor groove and bends the DNA. The activity of promoter-reporter constructs, consisting of the rat COL1A1 promoter fused to the chloramphenicol acetyltransferase (CAT) gene, were sensitive to mutations in sites A and/or B when stably transfected into UMR 106-01 cells. In conclusion, NP/NMP4 are NM architectural transcription factors that contribute to the regulation of the COL1A1 promoter. We propose that NP/NMP4 act to transduce changes in cell shape and cell adhesion into changes in collagen expression in bone.