Identification of a homeodomain binding element in the bone sialoprotein gene promoter that is required for its osteoblast-selective expression.
Bone sialoprotein is a 70-kDa extracellular matrix component that is intimately associated with biomineralization, yet the cis-acting elements of the Bsp gene that restrict its expression to mineralizing cells remain uncharacterized. To identify such elements, we analyzed a 2472-base pair fragment of the murine promoter that directs osteoblast-selective expression of a luciferase reporter gene and found that the region between -338 and -178 relative to the transcriptional start is crucial for its osteoblast-selective activity. We identified an element within this region that binds a protein complex in the nuclear extracts of osteoblastic cells and is required for its transcriptional activity. Introduction of a mutation that disrupts a homeodomain binding site within this sequence eliminates both its in vitro binding and nearly all of the osteoblastic-selective activity of the 2472-base pair promoter. We further found that the Dlx5 homeoprotein, which is able to regulate the osteoblast-specific osteocalcin promoter, can bind this element and stimulate its enhancer activity when overexpressed in COS7 cells. These data represent the first description of an osteoblast-specific element within the bone sialoprotein promoter and demonstrate its regulation by a member of a family of factors known to be involved in skeletogenesis.
Benson
M D
MD
Department of Biological Chemistry, School of Medicine, University of Michigan, Ann Arbor, Michigan 48109-1078, USA.
Bargeon
J L
JL
Xiao
G
G
Thomas
P E
PE
Kim
A
A
Cui
Y
Y
Franceschi
R T
RT
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D005786:Gene Expression Regulation; D018398:Homeodomain Proteins / Q000235:genetics*; D058574:Integrin-Binding Sialoprotein; D051379:Mice; D008969:Molecular Sequence Data; D009928:Organ Specificity; D010006:Osteoblasts / Q000502:physiology*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012795:Sialoglycoproteins / Q000235:genetics
D018398:Homeodomain Proteins; C546872:Ibsp protein, mouse; D058574:Integrin-Binding Sialoprotein; D012795:Sialoglycoproteins