Abstract：
PURPOSE:To determine whether antisense oligonucleotides (AODNs) targeted against CRX, a photoreceptor-specific trans-acting factor, suppress CRX expression and interphotoreceptor retinoid binding protein (IRBP) promoter activity. METHODS:Cultures of human retinoblastoma cells were transfected with chloramphenicol acetyltransferase (CAT) reporter plasmids containing a mouse IRBP promoter and AODNs directed against CRX. RT-PCR using primers specific to CRX, OTX2, GAPDH, or RNase H was conducted on total RNA isolated from retinoblastoma cells at various times following transfection with AODNs. RESULTS:Transfection of retinoblastoma cells with IRBP promoter CAT constructs alone produced high activity. Co-transfection with AODNs suppressed IRBP promoter activity in a concentration-dependent manner, with half-maximal effect produced at about 2 nM AODN concentration. Transfection with CAT constructs containing an SV40 promoter produced high activity that was unaffected by co-transfection with AODNs. RT-PCR products were obtained for all target sequences. CRX RT-PCR product from cells transfected with AODNs was greatly diminished following transfection with an AODN whereas control transcripts, including that of OTX2, were relatively unaffected. CONCLUSIONS:The CRX-specific AODNs specifically and potently suppressed CRX expression and IRBP promoter activity, as measured by RT-PCR and transient transfection assays, respectively. Little or no effect was seen on controls. These data suggest that endogenous CRX is required for IRBP promoter activity in retinoblastoma cells.
journal_name
Brain Resjournal_title
Brain researchauthors
Boatright JH,Borst DE,Stodulkova E,Nickerson JMdoi
10.1016/s0006-8993(01)02884-0subject
Has Abstractpub_date
2001-10-19 00:00:00pages
136-42issue
1-2eissn
0006-8993issn
1872-6240pii
S0006-8993(01)02884-0journal_volume
916pub_type
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