Transcriptional Regulation and Targeting of NF1 Gene Expression
Abstract
An essential requirement in understanding how genes contribute to genetic disease is the thorough knowledge of the mechanisms controlling gene expression at the level of transcription. In this project we are identifying and functionally characterizing the NF1 regulatory region, and evaluating the molecular mechanisms that regulate NF1 transcription during normal cellular differentiation and the events leading to NF1 -related neoplasia. Overlapping regions of the NFl proximal promoter have been cloned using a series of DNA fragments created by PCR and characterized for use in the luciferase assays to isolate specific binding motifs near the NF1 transcription start site. Luciferase reporter assays have identified a 425 bp region displaying activities 50 to 80 higher than baseline reporter levels. Mutations at putative CRE and SP1 binding sites immediately 5' to the transcription start site have dramatic effects that lead to a 70-90% decrease in reporter activity. As well, these assays have revealed a putative repressor region within the NFl promoter region corresponding to CCCTC rich sequences between the transcription and translation start sites. Preliminary gelshift assays confirm binding of SP1 and CRE to their putative recognition sequences and provide the first evidence identifying functional sites likely involved in regulating NF1 transcription.
Document Details
- Document Type
- Technical Report
- Publication Date
- Oct 01, 2001
- Accession Number
- ADA407208
Entities
People
- David I. Rodenhiser