Differential display detects altered gene expression between cataractous and normal human lenses.
OBJECTIVE
To identify and analyze differentially genes expressed between lens epithelia dissected from age-related cataractous and noncataractous human lenses.


METHODS
RNAs from 50 pooled cataractous and 25 pooled noncataractous epithelia were compared by reverse transcription-polymerase chain reaction differential display (RT-PCR-DD). Two differentially displayed bands were chosen for further study. These were reamplified, cloned, and sequenced. Expression of these genes was further evaluated in pooled and individual epithelia by RT-PCR with gene-specific primers.


RESULTS
Significant differences in gene expression were detected between the cataractous and the noncataractous epithelia. Three mRNAs displayed higher levels and 12 mRNAs displayed lower levels of expression in the cataractous samples compared with that in the noncataractous samples. Of the mRNAs expressed at higher levels, one was identified as metallothionein IIa (METII). Of the mRNAs with decreased expression, one was identified as protein phosphatase 2A regulatory subunit (P2A-RS). Overexpression of METII and underexpression of P2A-RS were confirmed in pooled and individual epithelia.


CONCLUSIONS
These results provide evidence that age-related cataract is associated with alterations in the expression of multiple epithelial genes including METII and P2A-RS. METII is a detoxification protein induced by oxidative stress, and P2A-RS is a mitotic suppressor involved in cell-cycle control. These results implicate these proteins and their associated functions in the maintenance of lens transparency.
Kantorow
M
M
The Jules Stein Eye Institute, Los Angeles, California 90095-7065, USA.
Kays
T
T
Horwitz
J
J
Huang
Q
Q
Sun
J
J
Piatigorsky
J
J
Carper
D
D
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D001483:Base Sequence; D002386:Cataract / Q000235:genetics* / Q000378:metabolism; D003459:Crystallins / Q000235:genetics* / Q000378:metabolism; D017931:DNA Primers / Q000737:chemistry; D004587:Electrophoresis, Agar Gel; D004848:Epithelium / Q000378:metabolism; D005260:Female; D015870:Gene Expression; D006801:Humans; D007908:Lens, Crystalline / Q000378:metabolism*; D008297:Male; D008668:Metallothionein / Q000235:genetics* / Q000378:metabolism; D008875:Middle Aged; D008969:Molecular Sequence Data; D010749:Phosphoprotein Phosphatases / Q000235:genetics* / Q000378:metabolism; D054648:Protein Phosphatase 2; D012313:RNA / Q000302:isolation & purification; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment
D003459:Crystallins; D017931:DNA Primers; D012333:RNA, Messenger; D012313:RNA; D008668:Metallothionein; D010749:Phosphoprotein Phosphatases; D054648:Protein Phosphatase 2