PurposeWe characterize calpain-5 (CAPN5) expression in retinal and neuronal subcellular compartments.MethodsCAPN5 gene variants were classified using the exome variant server, and RNA-sequencing was used to compare expression of CAPN5 mRNA in the mouse and human retina and in retinoblastoma cells. Expression of CAPN5 protein was ascertained in humans and mice in silico, in mouse retina by immunohistochemistry, and in neuronal cancer cell lines and fractionated central nervous system tissue extracts by Western analysis with eight antibodies targeting different CAPN5 regions.ResultsMost CAPN5 genetic variation occurs outside its protease core; and searches of cancer and epilepsy/autism genetic databases found no variants similar to hyperactivating retinal disease alleles. The mouse retina expressed one transcript for CAPN5 plus those of nine other calpains, similar to the human retina. In Y79 retinoblastoma cells, the level of CAPN5 transcript was very low. Immunohistochemistry detected CAPN5 expression in the inner and outer nuclear layers and at synapses in the outer plexiform layer. Western analysis of fractionated retinal extracts confirmed CAPN5 synapse localization. Western blots of fractionated brain neuronal extracts revealed distinct subcellular patterns and the potential presence of autoproteolytic CAPN5 domains.ConclusionsCAPN5 is moderately expressed in the retina and, despite higher expression in other tissues, hyperactive disease mutants of CAPN5 only manifest as eye disease. At the cellular level, CAPN5 is expressed in several different functional compartments. CAPN5 localization at the photoreceptor synapse and with mitochondria explains the neural circuitry phenotype in human CAPN5 disease alleles.

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Early PVR is characterized by activation of T cells and mTOR signaling, whereas advanced PVR is characterized by a chronic monocyte response. PVR might be treated by rational repositioning of existing drugs that target mTOR and IL-6. Our analysis demonstrates that successful therapeutic intervention will be highly dependent on the specific therapeutic target and the stage of PVR. This study provides insights into cytokines that will serve as biomarkers and therapeutic targets. These biomarkers will help design clinical trials that intervene at appropriate times.