Peroxisome biogenesis disorders (PBDs)

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In conjunction with our collaborators and postdoctoral support by the Global Foundation for Peroxisome Disorders (GFPD) and the Wynne Mateffy Foundation the ADST laboratory of NCATS has used a high content automated microscopy-based imaging assay to conduct the screening of a 20K gene siRNA library targeting the human whole genome. Active genes whose knocking-down promoted peroxisome assembly were further evaluated in follow-up testing by immunofluorescence (IF) staining with antibodies to peroxisomal membrane protein PMP70 leading to the identification of several novel candidate genes. Gene ontology analysis revealed that several active genes were closely associated with the protein deneddylation process which plays critical roles in regulating cullin-RING ubiquitin ligase (CRL) activity. Knocking-down these genes in patient fibroblasts largely increased the number of peroxisomes with improved function as determined by catalase and PMP70 co-staining. In addition, the processing percentage of peroxisomal AGPS proteins was elevated as judged from Western blotting analysis. Intriguingly, PEX6, which forms a hexamer with PEX1 and facilitates peroxisomal import, was shown to be upregulated upon the siRNA transfection; whereas the other PEX proteins remain unchanged.