Porcine S-antigen: cDNA sequence and expression in retina, ciliary epithelium and iris.
cDNA clones encoding S-antigen (arrestin) were isolated from the expression library constructed from porcine retina and sequenced. The 1490 base pair fragment contained a 1215 base pair open reading frame. From the nucleotide sequence, an amino acid sequence consisting of 405 residues was deduced and a molecular mass of 45,102 daltons was calculated. In order to determine whether the S-antigen mRNA transcript was expressed in anterior eye tissues, mRNA from ciliary non-pigmented epithelial cells and pigmented epithelial cells and iris was analysed by the reverse transcription polymerase chain reaction (PCR) using primers taken from sequences flanking the coding and non-coding regions of retinal S-antigen. Sequence analysis of the expected 611 base pairs in the 5' region and 672 base pairs in the 3' region of DNA fragments indicated that an identical mRNA for S-antigen was expressed in the anterior tissues examined. To investigate the in situ expression of S-antigen mRNA, 35S-labeled sense and antisense RNA probes were synthesized from the cDNA to label frozen sections of retina, ciliary body and iris and the treated sections were examined by autoradiography. The antisense probe labeled the layer between retinal pigmented epithelium and the outer nuclear layer of the retina, ciliary epithelium, and iris epithelium. From the results of sequencing PCR products and in situ hybridization, we concluded that, in porcine eye, the mRNA for S-antigen is expressed not only in the retina but also in the anterior tissues such as the ciliary epithelium and iris epithelium.
Singh
A K
AK
Department of Ophthalmology, Wayne State University School of Medicine, Detroit, MI 48201, USA.
Kumar
G
G
Shinohara
T
T
Shichi
H
H
D000818:Animals; D000941:Antigens / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics; D019393:Arrestin; D001324:Autoantigens / Q000032:analysis; D001483:Base Sequence; D002924:Ciliary Body / Q000276:immunology*; D018076:DNA, Complementary / Q000737:chemistry*; D005136:Eye Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics; D017403:In Situ Hybridization; D007498:Iris / Q000276:immunology*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000032:analysis; D012160:Retina / Q000276:immunology*; D013552:Swine
D000941:Antigens; D019393:Arrestin; D001324:Autoantigens; D018076:DNA, Complementary; D005136:Eye Proteins; D012333:RNA, Messenger