RPE65 is an abundant protein in the retinal pigment epithelium. Mutations in RPE65 are associated with inherited retinal dystrophies. Although it is known that RPE65 is critical for regeneration of 11-cis retinol in the visual cycle, the function of RPE65 is elusive. Here we show that recombinant RPE65, when expressed in QBI-293A and COS-1 cells, has robust enzymatic activity of the previous unidentified isomerohydrolase, an enzyme converting all-trans retinyl ester to 11-cis retinol in the visual cycle. The initial rate for the reaction is 2.9 pmol͞min per mg of RPE65 expressed in 293A cells. The isomerohydrolase activity of RPE65 requires coexpression of lecithin retinol acyltransferase in the same cell to provide its substrate. This enzymatic activity is linearly dependent on the expression levels of RPE65. This study demonstrates that RPE65 is the long-sought isomerohydrolase and fills a major gap in our understanding of the visual cycle. Identification of the function of RPE65 will contribute to the understanding of the pathogenesis for retinal dystrophies associated with RPE65 mutations.isomerase ͉ LRAT ͉ retinal dystrophy ͉ retinyl ester ͉ 11-cis retinal I n vertebrates, vision is initiated in rod and cone photoreceptors.The photosensitive entities in these cells are the visual pigments which consist of an apoprotein, opsin, and a chromophore, 11-cis retinal, which is attached to the opsin by a Schiff's base bond (1). Upon absorption of light by the pigments, 11-cis retinal is isomerized to all-trans retinal, which leads to the conformational changes of opsin and subsequently activates G protein transducin, initiating vision (1, 2). Efficient regeneration of 11-cis retinal, referred to as the visual retinoid cycle (see Fig. 1) is critical for the regeneration of the visual pigments (for reviews, see refs. 3-5) and maintenance of visual function. Disruption of the visual cycle by mutation or dysfunction of one of the numerous enzymes involved in this process leads to a number of blinding disorders (6).The enzyme in the visual cycle that has eluded identification is the isomerohydrolase, which is responsible for the isomerization and hydrolysis of all-trans retinyl ester to 11-cis retinol. Rando and colleagues (7) first proposed that all-trans retinol is first acylated to retinyl esters by lecithin retinol acyltransferase (LRAT). The generated all-trans retinyl ester is then directly isomerized and hydrolized into 11-cis retinol in the retinal pigment epithelium (RPE). This hydrolysis-isomerization process has been proposed to be catalyzed by a single enzyme, referred to as isomerohydrolase, which is associated with the membrane in RPE microsomes (8).Although isomerohydrolase activity had been demonstrated in the RPE almost 20 years ago, identification of the enzyme catalyzing this reaction has been difficult, because this activity is associated with the membrane and is abolished by solubilization in all of the detergents investigated (9). As a result, the isomerohydrolase has not been identified despite in...
We associated environmental and genetic factors with base substitution patterns of somatic mutations and provide a registry of genes and pathways that are disrupted in ESCCs. These findings might be used to design specific treatments for patients with esophageal squamous cancers.
Background:Functional microRNAs (miRNAs) in exosomes have been recognised as potential stable biomarkers in cancers. The aim of this study is to identify specific miRNAs in exosome as serum biomarkers for the early detection of recurrence in human colorectal cancer (CRC).Methods:Serum samples were sequentially obtained from six patients with and without recurrent CRC. The miRNAs were purified from exosomes, and miRNA microarray analysis was performed. The miRNA expression profiles and copy number aberrations were explored using microarray and array CGH analyses in 124 CRC tissues. Then, we validated exosomal miRNAs in 2 serum sample sets (90 and 209 CRC patients) by quantitative real-time RT–PCR.Results:Exosomal miR-17-92a cluster expression level in serum was correlated with the recurrence of CRC. Exosomal miR-19a expression levels in serum were significantly increased in patients with CRC as compared with healthy individuals with gene amplification. The CRC patients with high exosomal miR-19a expression showed poorer prognoses than the low expression group (P<0.001).Conclusions:Abundant expression of exosomal miR-19a in serum was identified as a prognostic biomarker for recurrence in CRC patients.
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