Cloning and characterization of the proximal murine Phex promoter.
Phex is an endopetidase that regulates systemic phosphate homeostasis. We investigated Phex gene transcription by cloning and performing functional analysis of the 2736 bp of the 5' flanking region of the mouse Phex gene containing its promoter. We identified a transcription start site, a consensus TATA-box, and multiple potential cis-acting regulator elements. To determine whether the promoter directs cell-type restricted Phex expression, we transfected full-length and 5'-deleted Phex luciferase reporter constructs into various cell lines. Phex-expressing C5.18 chondrocytes displayed the highest activity of the transfected Phex promoter constructs compared with non-Phex-expressing COS-7 cells, whereas promoter activity was intermediate in ROS 17/2.8 osteoblasts and maturation stage-dependent in MC3T3-E1 osteoblasts. Analysis of sequential 5'-deletion mutants of the Phex promoter in ROS 17/2.8 cells revealed bimodal activity, suggesting that both positive and negative cis-acting regions may be present. The chondrogenic factor SOX9 markedly stimulated Phex promoter activity, whereas Cbfa1, PTH, and 1,25(OH)(2)D(3) had no effect. Our findings are consistent with the predominant expression of Phex in bone and cartilage. Additional studies will be needed to confirm the regulatory regions in the Phex promoter that function in a cell-restricted manner.
Liu
S
S
Department of Medicine, Duke University Medical Center, Durham, North Carolina 27710, USA.
Guo
R
R
Quarles
L D
LD
D020121:5' Untranslated Regions / Q000235:genetics; D000595:Amino Acid Sequence / Q000235:genetics; D000818:Animals; D001483:Base Sequence / Q000235:genetics; D002460:Cell Line; D016922:Cellular Senescence / Q000502:physiology; D003001:Cloning, Molecular; D005347:Fibroblasts / Q000378:metabolism; D006609:High Mobility Group Proteins / Q000494:pharmacology; D051379:Mice; D008969:Molecular Sequence Data; D010006:Osteoblasts / Q000378:metabolism; D053402:PHEX Phosphate Regulating Neutral Endopeptidase; D011401:Promoter Regions, Genetic / Q000187:drug effects / Q000235:genetics* / Q000502:physiology*; D011506:Proteins / Q000235:genetics*; D012333:RNA, Messenger / Q000378:metabolism; D055758:SOX9 Transcription Factor; D014157:Transcription Factors / Q000494:pharmacology; D014158:Transcription, Genetic / Q000502:physiology; D015854:Up-Regulation
D020121:5' Untranslated Regions; D006609:High Mobility Group Proteins; D011506:Proteins; D012333:RNA, Messenger; D055758:SOX9 Transcription Factor; C527933:Sox9 protein, mouse; D014157:Transcription Factors; D053402:PHEX Phosphate Regulating Neutral Endopeptidase; C506232:Phex protein, mouse