Osteocalcin gene promoter-binding factors are tissue-specific nuclear matrix components.

Abstract

The nuclear matrix appears to play an important role in developmental gene expression during osteoblast differentiation. To better understand this role, we examined nuclear matrix DNA-binding proteins that are sequencespecific and interact with the osteocalcin gene promoter. Multiple protein-DNA interactions involving two distinct nuclear matrix proteins occur within the 5' regulatory sequences (nt -640 to -430). One of these proteins, NMP-1, is a ubiquitous, cell growth-regulated protein that is related to the transcription factor ATF and resides in both the nuclear matrix and the nonmatrix nuclear compartment. The other protein, NMP-2, is a cell type-specific, 38-kDa promoter factor that recognizes binding sites resembling the consensus site for the CCAAT/enhancer-binding protein C/EBP and is localized exclusively on the nuclear matrix. NMP-1 and NMP-2 each interact with two nuclear matrix protein-binding elements. These elements are present near key regulatory sites of the osteocalcin gene promoter, such as the principal steroid hormone (vitamin D)-responsive sequences. Binding in this region of the osteocalcin gene promoter suggests transient associations with the nuclear matrix that are distinct from the stable interactions of matrix attachment regions. Our results are consistent with involvement of the nuclear matrix in concentrating and/or localizing transcription factors that mediate the basal and steroid hormone responsiveness of osteocalcin gene transcription.Promoter-binding factors associate with DNA regulatory elements to initiate RNA transcription. This interaction cannot be simply through diffusion, since factor concentration in the nucleus is far too low. Very likely, DNA-binding factors and DNA are brought intojuxtaposition by the nonchromatin scaffolding of the nucleus termed the nuclear matrix. By regulating this interaction, the nuclear matrix can play a fundamental role in transcriptional control.The nuclear matrix is the nonchromatin nuclear substructure (1), which is organized as a proteinaceous network of polymorphic anastomosing fibers (2). Some nuclear matrix components are clearly cell-and tissue-type specific, including some matrix proteins and the heterogeneous nuclear RNA, an integral part of matrix structure (3-7). The linkage of nuclear matrix composition to cell and tissue type strongly suggests nuclear matrix participation in the regulation ofgene expression. There is other evidence of nuclear matrix contribution to the regulation of gene expression. This includes the preferential association of actively transcribed genes with the matrix (8-12), the localization of RNA synthesis and pre-mRNA splicing on the matrix (13-15), and the presence of steroid receptors in the matrix fraction (16)(17)(18). Our recent studies showed that a cell cycle-regulated histone gene interacts with the nuclear matrix when actively transcribed in proliferating cells and that a sequence-specific, activating transcription factor (ATF)-related histone gene promoterbinding factor associates with t...

Engineering gene expression and protein synthesis by modulation of nuclear shape

“…Therefore, changes in the nuclear matrix architecture could affect the degree of promoter supercoiling and bending and result in altered transcriptional activity of OC. Similarly, NMP-1 (or YY1), which is not osteoblast-specific but binds the OC promoter, is also found in the nuclear matrix fractions of osteoblasts and is recognized as a complex DNA binding͞bending protein with multiple roles (13). Both NMP-1 and NMP-2 have been characterized as DNA binding and DNA bending proteins whose involvement in OC expression has been identified but not fully elaborated.…”

“…The osteoblast phenotype has been influenced by morphology, external forces, cytoskeletal organization, the extracellular matrix, and nuclear matrix architecture (14,25). Additionally, several nuclear matrixspecific proteins such as NMP-2 (also identified as Cbfa1 or Osf2) have been shown to bind to the promoter region of the OC gene (13). Therefore, changes in the nuclear matrix architecture could affect the degree of promoter supercoiling and bending and result in altered transcriptional activity of OC.…”

“…Both NMP-1 and NMP-2 have been characterized as DNA binding and DNA bending proteins whose involvement in OC expression has been identified but not fully elaborated. However, the continuous link between the extracellular matrix, cytoskeleton, nuclear matrix, and promoter region of the OC gene indicates the importance of cell͞nuclear morphology and intracellular signaling in OC gene activation (13,26). Potentially, engagement of ␣ 2 integrins with CollI and subsequent activation of the mitogen-activated protein kinase pathway (MAPK)͞MAPK kinase leads to NMP-2 activation and expression of the OC gene (26).…”

“…Furthermore, cells can be forced to enter the apoptotic cascade when the area on which the cell is allowed to spread is constrained (1). One proposed mechanism for the transduction of cell shape information into gene expression is through mechanical forces transmitted by means of the direct link of the cytoskeleton to the nucleus (9,12), and in particular to nuclear matrix proteins (NMPs) such as NMP-1 and NMP-2 (13). These architectural transcription factors, which are components of the nuclear scaffold, induce changes in DNA supercoiling and can interact directly with gene promoter sequences (14).…”

“…Because of our interest in bone, we patterned primary bone-derived cells from rat calvaria and probed for type I collagen (CollI) synthesis and osteocalcin (OC) mRNA expression to determine whether constraining the projected area and nuclear shape of a bone-derived cell altered the time required for differentiation into the osteoblast phenotype. We chose CollI and OC because they are markers of osteoblast differentiation, and NMPs (e.g., Cbfa1, a transcriptional activator of osteoblast differentiation) have been shown to bind to the promoter region of genes for both OC and CollI (13).…”