Deciphering the Mechanisms of WT1 Glomerulopathy

Wilms' tumour 1 (WT1) is a transcription factor encoding a zinc finger protein that controls podocyte differentiation and is highly expressed in mature podocytes. WT1 mutations can lead to renal failure due to glomerular scarring, the underlying mechanisms, of which, are poorly understood. This project explored the mechanisms of glomerulosclerosis by using a tamoxifen-inducible Cre-LoxP system to delete Wt1 in adult mice. Following the fourth day post-induction with Tamoxifen, podocyte apoptosis was evident and increased as the disease progressed, highlighting Wt1’s key role in mature podocyte survival. At disease onset, increased podocyte Notch1 transcript and its downstream targets, including Nrarp and Hey2 were observed. Decreased expression of podocyte FoxC2 transcript at the same time-point was noted, thereby supporting previous findings in lower vertebrates for a transcriptional relationship between Wt1/FoxC2/Notch in podocyte function. Podocyte Notch1 and Hes1 protein expression was observed in mutant mouse glomeruli at the onset of glomerulosclerosis. Induced podocyte Hes1 expression was associated with an upregulation of Snai1 and Slug transcripts, genes associated with epithelial to mesenchymal transition (EMT), thus proposing a role for Hes1 in mediating podocyte EMT. Moreover, early pharmacological inhibition of Notch, with gamma secretase inhibitors, ameliorated glomerulosclerosis and albuminuria. This data provides evidence that Wt1 deletion modulates podocyte Notch signalling in mature podocytes, leading to early events in WT1-related glomerulosclerosis

Inhibition of VCP preserves retinal structure and function in autosomal dominant retinal degeneration

Due to continuously high production rates of rhodopsin (RHO) and high metabolic activity, photoreceptor neurons are especially vulnerable to defects in proteostasis. A proline to histidine substitution at position 23 (P23H) leads to production of structurally misfolded RHO, causing the most common form of autosomal dominant Retinitis Pigmentosa (adRP) in North America. The AAA-ATPase valosin-containing protein (VCP) extracts misfolded proteins from the ER membrane for cytosolic degradation. Here, we provide the first evidence that inhibition of VCP activity rescues degenerating P23H rod cells and improves their functional properties in P23H transgenic rat and P23H knock-in mouse retinae, both in vitro and in vivo. This improvement correlates with the restoration of the physiological RHO localization to rod outer segments (OS) and properly-assembled OS disks. As a single intravitreal injection suffices to deliver a long-lasting benefit in vivo, we suggest VCP inhibition as a potential therapeutic strategy for adRP patients carrying mutations in the RHO gene

The kinetochore protein, CENPF, is mutated in human ciliopathy and microcephaly phenotypes

Background: Mutations in microtubule-regulating genes are associated with disorders of neuronal migration and microcephaly. Regulation of centriole length has been shown to underlie the pathogenesis of certain ciliopathy phenotypes. Using a next-generation sequencing approach, we identified mutations in a novel centriolar disease gene in a kindred with an embryonic lethal ciliopathy phenotype and in a patient with primary microcephaly. Methods and results Whole exome sequencing data from a non-consanguineous Caucasian kindred exhibiting mid-gestation lethality and ciliopathic malformations revealed two novel non-synonymous variants in CENPF, a microtubule-regulating gene. All four affected fetuses showed segregation for two mutated alleles [IVS5-2A>C, predicted to abolish the consensus splice-acceptor site from exon 6; c.1744G>T, p.E582X]. In a second unrelated patient exhibiting microcephaly, we identified two CENPF mutations [c.1744G>T, p.E582X; c.8692 C>T, p.R2898X] by whole exome sequencing. We found that CENP-F colocalised with Ninein at the subdistal appendages of the mother centriole in mouse inner medullary collecting duct cells. Intraflagellar transport protein-88 (IFT-88) colocalised with CENP-F along the ciliary axonemes of renal epithelial cells in age-matched control human fetuses but did not in truncated cilia of mutant CENPF kidneys. Pairwise co-immunoprecipitation assays of mitotic and serum-starved HEKT293 cells confirmed that IFT88 precipitates with endogenous CENP-F. Conclusions: Our data identify CENPF as a new centriolar disease gene implicated in severe human ciliopathy and microcephaly related phenotypes. CENP-F has a novel putative function in ciliogenesis and cortical neurogenesis