Alternative splicing of the APC gene in the neural retina and retinal pigment epithelium.
OBJECTIVE
Hypertrophy and hyperplasia of the retinal pigment epithelium (RPE) is associated with an inherited predisposition to human familial adenomatous polyposis coli, suggesting that expression of the adenomatous polyposis coli (APC) tumor suppressor may regulate RPE proliferation/differentiation. Distinctive APC isoforms exist in different cell types due to alternative splicing of the APC transcripts. We hypothesize that differences in expression patterns of APC protein isoforms are critical to RPE proliferation/differentiation.


METHODS
To investigate these relationships, APC gene expression was characterized in the retinas and RPE from fetal and adult human and mouse, and in the epiretinal membranes (ERM) from 5 patients with proliferative vitreoretinopathy (PVR). Expression patterns of alternative splice-forms of APC transcripts were evaluated by comparative quantitative RT-PCR. Exon 1 of APC encodes a heptad repeat that confers the ability of APC to homodimerize. APC protein isoforms containing or lacking this heptad were characterized by western blot analysis and immunohistochemistry.


RESULTS
Comparative quantitative RT-PCR demonstrated a predominant exon 1 containing, conventional APC splice-form in the early developing fetal RPE and retina, and in all the tested ERM samples from patients with PVR. This method also demonstrated an increased level of exon 1 lacking APC splice-form in the mature RPE and retina. Western blot analysis and immunofluorescence microscopy demonstrated the conventional APC only in the RPE, and the APC isoform without the first heptad repeat in both the retina and RPE. Immunofluorescence microscopy also demonstrated only the conventional APC in the ERM samples tested.


CONCLUSIONS
These results suggest that alternative splicing of APC leads to differential APC expression with potentially unique functions. APC isoform without the first heptad repeat may play a role in cell cycle cessation in the adult retina and RPE, and the down regulation of this APC isoform may contribute to the potential of RPE to migrate and proliferate.
Liou
Gregory I
GI
Department of Ophthalmology, Medical College of Georgia, Augusta, GA 30912, USA. giliou@mail.mcg.edu
Samuel
Sara
S
Matragoon
Suraporn
S
Goss
Kathleen H
KH
Santoro
Irma
I
Groden
Joanna
J
Hunt
Richard C
RC
Wang
Fei
F
Miller
Sheldon S
SS
Caldwell
Ruth B
RB
Rustgi
Anil K
AK
Singh
Harinderjit
H
Marcus
Dennis M
DM
D025601:Adenomatous Polyposis Coli Protein / Q000378:metabolism; D000368:Aged; D000369:Aged, 80 and over; D017398:Alternative Splicing; D000818:Animals; D015153:Blotting, Western; D002455:Cell Division; D002465:Cell Movement; D015536:Down-Regulation; D019773:Epiretinal Membrane / Q000378:metabolism; D005260:Female; D005786:Gene Expression Regulation / Q000502:physiology*; D017491:Genes, APC; D006801:Humans; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D008856:Microscopy, Fluorescence; D008875:Middle Aged; D010857:Pigment Epithelium of Eye / Q000378:metabolism*; D020033:Protein Isoforms / Q000235:genetics; D012160:Retina / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D025601:Adenomatous Polyposis Coli Protein; D020033:Protein Isoforms