Pathogenesis of retinitis pigmentosa associated with apoptosis-inducing mutations in carbonic anhydrase IV

Abstract

Missense mutations in the carbonic anhydrase IV (CA IV) gene have been identified in patients with an autosomal dominant form of retinitis pigmentosa (RP17). We used two transient expression systems to investigate the molecular mechanism by which the newly identified CA IV mutations, R69H and R219S, contribute to retinal pathogenesis. Although the R219S mutation drastically reduced the activity of the enzyme, the R69H mutation had a minimal effect, suggesting that loss of CA activity is not the molecular basis for their pathogenesis. Defective processing was apparent for both mutant proteins. Cell surface-labeling techniques showed that the R69H and R219S mutations both impaired the trafficking of CA IV to the cell surface, resulting in their abnormal intracellular retention. Expression of both CA IV mutants induced elevated levels of the endoplasmic reticulum (ER) stress markers, BiP and CHOP, and led to cell death by apoptosis. They also had a dominant-negative effect on the secretory function of the ER. These properties are similar to those of R14W CA IV, the signal sequence variant found in the original patients with RP17. These findings suggest that toxic gain of function involving ER stress-induced apoptosis is the common mechanism for pathogenesis of this autosomal-dominant disease. Apoptosis induced by the CA IV mutants could be prevented, at least partially, by treating the cells with dorzolamide, a CA inhibitor. Thus, the use of a CA inhibitor as a chemical chaperone to reduce ER stress may delay or prevent the onset of blindness in RP17.autosomal dominant ͉ chemical chaperone ͉ ER stress ͉ missense mutation ͉ Naϩ/bicarbonate co-transporter 1

Progressive renal injury from transgenic expression of human carbonic anhydrase IV folding mutants is enhanced by deficiency of p58 IPK

“…When expressed in Cos-7 or HEK-293 cells, the ER folding/maturation of the R219S hCAIV was drastically impaired such that its cell surface expression was only approximately 10% compared with WT. Its enzymatic activity was also significantly reduced (>90%) (10). In contrast, the R14W was a milder mutation, causing only 30% reduction in enzymatic activity and a similar reduction in the rate of maturation (6).…”

“…The R219S and R14W hCAIV were characterized as "folding mutants" based on previous cell culture studies (6,9,10). When expressed in Cos-7 or HEK-293 cells, the ER folding/maturation of the R219S hCAIV was drastically impaired such that its cell surface expression was only approximately 10% compared with WT.…”

“…Previously we demonstrated that in vitro expression of the folding mutants of hCAIV in HEK293 and COS-7 cells caused abnormal processing in the ER that ultimately led to ER stress-induced apoptosis (6,10). To examine whether renal tubular cells of the mice carrying the R219S or R14W hCAIV transgene undergo similar cell death, we performed TUNEL staining on the kidney sections of mice at various ages ( Fig.…”

“…1A). They also differ in their impacts on the enzymatic activity and folding, R14W being a less severe mutation than R219S (6,10). How these differences, observed in vitro, affect their proteotoxicity in vivo To whom correspondence should be addressed.…”

“…This treatment also rescued the cells from proteotoxicity. Based on these data, we proposed that toxic gain of function involving ER stress-induced apoptosis is the common mechanism for the pathogenesis of RP17 and that this blinding disease is potentially treatable with chemical chaperone therapy (6,9,10).…”