Alternative splicing of the APC gene in the neural retina andretinal pigment epithelium

[摘要] Purpose: Hypertrophy and hyperplasia of the retinal pigmentepithelium (RPE) is associated with an inherited predisposition to humanfamilial adenomatous polyposis coli, suggesting that expression of theadenomatous polyposis coli (APC) tumor suppressor may regulate RPEproliferation/differentiation. Distinctive APC isoforms exist indifferent cell types due to alternative splicing of the APCtranscripts. We hypothesize that differences in expression patterns ofAPC protein isoforms are critical to RPE proliferation/differentiation.Methods: To investigate these relationships, APC gene expressionwas characterized in the retinas and RPE from fetal and adult human andmouse, and in the epiretinal membranes (ERM) from 5 patients withproliferative vitreoretinopathy (PVR). Expression patterns ofalternative splice-forms of APC transcripts were evaluated bycomparative quantitative RT-PCR. Exon 1 of APC encodes a heptad repeatthat confers the ability of APC to homodimerize. APC protein isoformscontaining or lacking this heptad were characterized by western blotanalysis and immunohistochemistry.Results: Comparative quantitative RT-PCR demonstrated a predominantexon 1 containing, conventional APC splice-form in the earlydeveloping fetal RPE and retina, and in all the tested ERM samples frompatients with PVR. This method also demonstrated an increased level ofexon 1 lacking APC splice-form in the mature RPE and retina. Westernblot analysis and immunofluorescence microscopy demonstrated theconventional APC only in the RPE, and the APC isoform without the firstheptad repeat in both the retina and RPE. Immunofluorescence microscopyalso demonstrated only the conventional APC in the ERM samples tested.Conclusions: These results suggest that alternative splicing ofAPC leads to differential APC expression with potentially uniquefunctions. APC isoform without the first heptad repeat may play a rolein cell cycle cessation in the adult retina and RPE, and the downregulation of this APC isoform may contribute to the potential of RPE tomigrate and proliferate.

[学科分类] 生物化学/生物物理