Inherited retinal degenerations (IRDs) are important causes of blindness that are characterized by progressive dysfunction and death of rod and cone photoreceptor cells leading to vision loss. Mutations in genes that encode the RNA splicing factors PRPF3, PRPF4, PRPF6, PRPF8, PRPF31, SNRNP200 and RP9 are the second most common cause of dominant Retinitis Pigmentosa (RP).
In eukaryotes, removal of intronic sequences from pre-mRNA molecules is intrinsic to mRNA maturation and gene expression. This function is termed splicing and is carried out by the spliceosome, a dynamic, multi-megadalton ribonucleoprotein (RNP) complex. Splicing is performed in a series of concerted steps involving several sub complexes each composed of hundreds of proteins and several small nuclear RNPs (snRNPs). One of those snRNPs that plays an indispensable role is the U4/U6.U5 tri-snRNP. Interestingly, all of the splicing factors found to be involved with RP are either components of or interact with the U4/U6.U5 tri-snRNP. Given that splicing is a fundamental cellular process that occurs in all the cells of the body it remains a mystery why mutations in such central players cause non syndromic RP.

In our lab we investigate RNA Splicing Factors dependent RP in both mouse and humans. We have developed knock-in mouse models mimicking the human mutations found in PRPF3 and PRPF8, as well as a knock-out mouse model of PRPF31. Phenotypic characterization of these models suggests the RPE is the primary site of pathogenesis for each of these models. The RPE undergoes morphological changes with a loss of basal infoldings and vacuolization at two years of age. Phagocytosis assays of primary RPE cultures suggest a loss of functional activity in two week-old mice. We hypothesize that aberrant splicing is the cause of pathogenesis in the RPE. To test this hypothesis we employed next generation sequencing technology to establish an RNAseq transcriptome data base from RPE, retina, brain and muscle tissues of all three mouse models. Analysis of this data base will allow us to identify the aberrant splicing events that lead to disease and identify targets for genetic therapeutics.
In parallel to our work with the mouse models we are also expanding our analysis to patient derived and CRISPR/Cas9 genome edited human induced pluripotent stem (hiPS) cells. These cells are differentiated in vitro to form RPE or used in in-vitro organogenesis to form Optic Vesicles (OV) that mimic the function of the retina.