Increased SK3 expression in DM1 lens cells leads to impaired growth through a greater calcium-induced fragility.
Although cataract is a characteristic feature of myotonic dystrophy type 1 (DM1), little is known of the underlying mechanisms. We generated four lens epithelial cell lines derived from DM1 cataracts and two from age-matched, non-DM cataracts. Small-pool PCR revealed typical large triplet repeat expansions in the DM1 cells. Furthermore, real-time PCR analysis showed reduced SIX5 expression and increased expression of the Ca(2+)-activated K(+) channel SK3 in the DM1 cells. These cells also exhibited longer population doubling times which did not arise through reduced proliferation, but rather increased cell death as shown by increased release of lactate dehydrogenase (LDH). Using (86)Rb(+) as a tracer for K(+), we found no difference in the resting K(+) influx or efflux kinetics. In all cases, the ouabain sensitive component of the influx contributed approximately 50% of the total. However, stimulating internal Ca(2+) by exposure to ionomycin not only caused greater stimulation of K(+) ((86)Rb) efflux in the DM1 cells but also induced a higher rate of cell death (LDH assay). Since both the hyper-stimulation of K(+) efflux and cell death were reduced by the highly specific SK inhibitor apamin, we suggest that increased expression of SK3 has a critical role in the increased Ca(2+)-induced fragility in DM1 cells. The present data, therefore, both help explain the lower epithelial cell density previously observed in DM1 cataracts and provide general insights into mechanisms underlying the fragility of other DM1-affected tissues.
Rhodes
Jeremy D
JD
School of Biological Sciences, University of East Anglia, Norwich, UK. j.rhodes@uea.ac.uk
Monckton
Darren G
DG
McAbney
John P
JP
Prescott
Alan R
AR
Duncan
George
G
D000328:Adult; D000368:Aged; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism*; D002386:Cataract / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D002460:Cell Line; D049109:Cell Proliferation; D002470:Cell Survival; D004847:Epithelial Cells / Q000378:metabolism* / Q000473:pathology; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D007908:Lens, Crystalline / Q000378:metabolism / Q000473:pathology; D008875:Middle Aged; D009223:Myotonic Dystrophy / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D016133:Polymerase Chain Reaction; D011188:Potassium / Q000378:metabolism; D051640:Small-Conductance Calcium-Activated Potassium Channels / Q000235:genetics* / Q000378:metabolism; D019680:Trinucleotide Repeat Expansion / Q000235:genetics
D018398:Homeodomain Proteins; C406328:KCNN3 protein, human; C106687:SIX5 protein, human; D051640:Small-Conductance Calcium-Activated Potassium Channels; D011188:Potassium; D002118:Calcium