Characterization of gene expression in human trabecular meshwork using single-pass sequencing of 1060 clones.

PURPOSE

To study the gene expression profile of the human trabecular meshwork (HTM).

METHODS

A polymerase chain reaction (PCR)-amplified cDNA library was constructed using RNA from the TM of a 67-year-old normal, perfused human eye. A total of 1060 clones were randomly selected for sequencing of one end. These sequences were searched against nonredundant GenBank and dbEST databases for similarity comparison by using a FASTA file and the BLASTcl3 program. Relative expression patterns of those clones that matched other expressed sequence tags (ESTs) were determined using the National Center for Biotechnology Information (NCBI) Unique Human Gene Sequence Collection (UniGene) database.

RESULTS

Of the 1060 clones analyzed, 519 (48.9%) had sequences identical with known genes, 125 (11.8%) matched ESTs, and 189 (17.8%) did not match any database sequences. Of the remaining clones, 31 (3%) corresponded to mitochondrial transcripts and 196 (18.5%) to repetitive and noninformative sequences. It is notable that some of the genes highly represented in this library are not ubiquitously expressed in other tissues, which suggests a potentially important role in the HTM. As evidence for the presence of true novel genes in the library, one of the clones was fully sequenced. This clone comprised a complete open reading frame of 966 nucleotides, and its deduced amino acid sequence corresponded to a protein 33% similar to the MAS-related G-protein-coupled receptor.

CONCLUSIONS

The identification of the more highly expressed genes in HTM and the discovery of novel genes expressed in this tissue provides basic information for further research on the physiology of the TM and for the identification of glaucoma candidate genes.