Low expression of alphaA-crystallins and rhodopsin kinase of photoreceptors in retinal dystrophy rat.
OBJECTIVE
The Royal College of Surgeons (RCS) rat has been extensively characterized as a model for inherited retinal dystrophy such as retinitis pigmentosa. In the present study, compositions of retinal proteins were compared between RCS (rdy-/-) and control (rdy+/+) rats during progression of the disease to understand the molecular pathologic course of the retinal degeneration.


METHODS
Protein mapping was performed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) or two-dimensional (2D)-PAGE using whole retinas or rod outer segments (ROS) obtained by a sucrose-density gradient centrifugation method from RCS or control rats at the age of 3 to 8 weeks.


RESULTS
2D-PAGE showed that retinal proteins of RCS rats were generally less abundant than those of the control animals and that the difference became more evident with aging. However, no significant difference was observed in the protein-mapping patterns in 2D-PAGE between RCS and control rats in any ages tested. Analysis by SDS-PAGE of ROS proteins and by western blot using antibodies against opsin, rhodopsin kinase (RK), recoverin, or arrestin demonstrated that a 20-kDa protein and RK were selectively less abundant in RCS than in control rats. Edman sequence analysis of the proteolytic peptides obtained by in-gel digestion of the corresponding protein band using endoproteinase Lys C identified the 20-kDa protein as alphaA-crystallin. Reverse transcription-polymerase chain reaction confirmed selective low levels of mRNA expressions of alphaA-crystallins and RK in RCS rats.


CONCLUSIONS
This study demonstrates that decreased expression of alphaA-crystallins and RK in RCS rats, may have significant roles in the development of retinal dystrophy.
Maeda
A
A
Department of Ophthalmology, Sapporo Medical University School of Medicine, Japan.
Ohguro
H
H
Maeda
T
T
Nakagawa
T
T
Kuroki
Y
Y
D000595:Amino Acid Sequence; D000818:Animals; D019393:Arrestin / Q000378:metabolism; D015153:Blotting, Western; D002135:Calcium-Binding Proteins / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D003459:Crystallins / Q000235:genetics / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional; D004591:Electrophoresis, Polyacrylamide Gel; D005136:Eye Proteins; D051606:G-Protein-Coupled Receptor Kinase 1; D015870:Gene Expression; D051597:Hippocalcin; D008074:Lipoproteins; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins; D010449:Peptide Mapping; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D011922:Rats, Mutant Strains; D051603:Recoverin; D012162:Retinal Degeneration / Q000235:genetics / Q000378:metabolism*; D012374:Rod Cell Outer Segment / Q000378:metabolism*; D017299:Rod Opsins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
D019393:Arrestin; D002135:Calcium-Binding Proteins; D003459:Crystallins; D005136:Eye Proteins; D008074:Lipoproteins; D009419:Nerve Tissue Proteins; D012333:RNA, Messenger; C496733:Rcvrn protein, rat; D017299:Rod Opsins; D051603:Recoverin; D051597:Hippocalcin; D011494:Protein Kinases; D051606:G-Protein-Coupled Receptor Kinase 1; C496765:Grk1 protein, rat