Abstract
The ability of the human insulin receptor promoter to direct expression of a linked chloramphenicol acetyltransferase gene was assessed in transient transfections into HepG2 and Hela cells. A 5'-deletional analysis of the promoter showed that regions between -646 and -489 were important for the activity of the proximal promoter. In addition, a possible negative regulatory element was identified between -1311 and -877 and a positive element between -1823 and -1311. DNase I footprint and gel retardation analysis showed that multiple factors bind to the human insulin receptor promoter. In particular, DNase I protection patterns were observed over the Sp1 sites at -620 to -599 and -438 to -392, a TC box at -533, four homopyrimidine/homopurine sites clustered around -1150, and a site at -1420 that contains the motif TGGCCC which has been shown to bind the liver-specific transcription factor LF-A1.
Highlights
The ability of the human insulin receptor promoter utilized by the gene are still unclear; no two groups agree on to direct expression of a linked chloramphenicol ace- the exact sites
As reported by specific receptor situated on the plasmamembrane [1, 2]. others, we find that a region containing the cluster of four This receptor belongs to thefamily of tyrosine kinases which S p l sites is important for full promoter activity in addition includes the receptors for epidermal growth factor and plate- to a TC box at -533 that has been described previously (7, let-derived growth factor [3].The insulin receptor is expressed 14)
A clone conlevels of IR expression, but thedifferences in mRNA half-life taining 1.8 kb of the human insulin receptor promoter cloned alone are notsufficient to account for the levels of IR seen in upstream of the coding sequence for the chloramphenicol acetyltransdifferent cells [5]
Summary
The vector pBLCAT8+ was digested with BglII, filled in with T4 DNA the transcription factor Spl. The transcriptional start sites polymerase, redigested with BamHI to excise the thymidine kinase promoter, and treatedwith calf intestinal phosphatase [16]. The vector pBLCAT8+ was digested with HindIII, filled in with T4 DNA polymerase, redigested with SalI, and treated with calf intestinal phosphatase. PhIRP1 was digested with HindIII, filled in with T4 DNA polymerase, redigested with XhoI, and the 300-bp fragment gel purified. The vector pBLCAT8+ was digested with BamHI, filled in with T4 DNA polymerase, redigested with SalI, and treated with calf intestinal phosphatase. Vector pBLCAT8+ was digested with BamHI, filled in with T4 DNA polymerase, redigested with HindIII, and treated with calf intestinal phosphatase. A G+A sequence ladder was generated by performing Maxam-Gilbert sequencing on each probe [15]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
