Related Experiment Videos
Expression of the ETS transcription factor ELF3 in the retinal pigment epithelium
Andrew Ian Jobling1, Zhiping Fang, Daniela Koleski
1National Vision Research Institute of Australia, Carlton, Victoria, Australia. a.jobling@optometry@optometry.unimelb.edu.au
Purpose:
The ETS family of transcription factors regulate several critical cellular functions. They have also been implicated in invertebrate ocular development. This work was undertaken to determine whether epithelium-specific ETS transcription factors are expressed in the retinal pigment epithelium and to investigate the possible role of these factors in retinal diseases such as age-related macular degeneration.
Methods:
The expression of the epithelial ETS transcription factors ELF5, ESE3, and ELF3 was assessed by RT-PCR in the human RPE cell lines D407 and hTERT-RPE1. The full-length coding sequence of rat Elf3 was isolated with 3' rapid amplification of cDNA ends (RACE) and degenerative primers, and its expression was determined in various rat tissues, by RT-PCR and real-time PCR. A polyclonal ELF3 antibody produced from a C-terminal peptide was used to observe the distribution of the transcription factor within the retina. To assess the possible ELF3 regulation of the TIMP3 promoter, transient transfection assays were performed. Promoter activity was determined with a firefly luciferase reporter gene construct.
Results:
The epithelium-specific ETS transcription factor ELF3 was expressed in the D407 and hTERT-RPE1 cell lines. Neither ESE3 nor ELF5 was detected in the RPE. The cloning of rat Elf3 produced two splice variants, designated Elf3a (1786 bp) and Elf3b (1855 bp). The larger form, Elf3b, contained a 69-bp insert in the coding sequence, which showed high homology to a similar insert previously identified in murine Elf3. Both splice variants were expressed in rat lung, kidney, liver, and retina, but were absent in heart tissue. Real-time PCR analysis showed the retina to contain high levels of Elf3, which was subsequently localized to the RPE. Elf3 upregulated the TIMP3 promoter, with Elf3a and -3b inducing an approximate sixfold increase in activity.
Conclusions:
The ELF3 transcription factor is highly expressed in the RPE and can regulate important ocular genes, such as TIMP3, in vitro. The specific expression of ELF3 in the RPE may reflect an important role for this transcription factor in retinal function. Furthermore, its regulation of TIMP3 may have implications for degenerative retinal diseases, such as age-related macular degeneration.