Abstract
Purpose:
Retinitis pigmentosa (RP) is the most common form of hereditary retinal degeneration. Mutations of the PRCD gene are associated with RP in both dogs and humans. The human PRCD gene encodes for 54 amino acids (aa) of unknown function. The first 20 aa appear to encode for a signal peptide (SP), suggesting that PRCD is a secreted protein. We aimed to test this hypothesis.
Methods:
A sporadic RP female patient underwent complete ophthalmological examination and genetic testing for all genes associated with autosomal recessive RP. To study PRCD secretion, C-terminally myc-tagged PRCD was expressed in cultured cells. Cells and conditioned media were analyzed by Western blot and immunostaining. To characterize the secretory pathway of PRCD we studied the effect of various pharmacological agents, which interfere with transport of proteins through the ER and Golgi to the plasma membrane.
Results:
The patient was found to be compound heterozygote for two PRCD mutations: the previously reported p.R18X, and a novel variant, p.P25T. Her phenotype resembles that of previously reported patients with PRCD mutations. In transfected cells, PRCD was found in both cell extracts and conditioned cell media. Interestingly, a truncated PRCD protein lacking the first 20 aa was present only in cell extracts and not in media, confirming that PRCD is secreted extracellularly and that its secretion is mediated by its N-terminal SP. PRCD secretion was significantly inhibited by all tested pharmacological agents, confirming that it is secreted through the classic ER/Golgi-dependent secretory pathway. We tested the effect of two mutations on the PRCD protein, and found that p.C2Y, but not p.P25T, affects protein stability, and that neither mutation affects protein secretion.
Conclusions:
Our data suggest that PRCD functions as a secreted protein. These findings shed a new light on PRCD function and the etiology of RP.
Keywords: 688 retina •
695 retinal degenerations: cell biology •
696 retinal degenerations: hereditary