No abstract

The surface expression and channel activation of transient receptor potential canonical 6 (TRPC6) were regulated by tyrosine phosphorylation and resultant binding with stimulatory PLC-γ1 and inhibitory nephrin. Disease-causing mutations made the TRPC6s insensitive to nephrin suppression, suggesting that the cell-type–specific regulation of TRPC6 might be involved in the pathogenesis.

Leber congenital amaurosis (LCA) is a hereditary early-onset retinal dystrophy that is accompanied by severe macular degeneration. In this study, novel compound heterozygous mutations were identified as LCA-causative in chaperonin-containing TCP-1, subunit 2 (CCT2), a gene that encodes the molecular chaperone protein, CCTβ. The zebrafish mutants of CCTβ are known to exhibit the eye phenotype while its mutation and association with human disease have been unknown. The CCT proteins (CCT α-θ) forms ring complex for its chaperon function. The LCA mutants of CCTβ, T400P and R516H, are biochemically instable and the affinity for the adjacent subunit, CCTγ, was affected distinctly in both mutants. The patient-derived induced pluripotent stem cells (iPSCs), carrying these CCTβ mutants, were less proliferative than the control iPSCs. Decreased proliferation under Cct2 knockdown in 661W cells was significantly rescued by wild-type CCTβ expression. However, the expression of T400P and R516H didn’t exhibit the significant effect. In mouse retina, both CCTβ and CCTγ are expressed in the retinal ganglion cells and connecting cilium of photoreceptor cells. The Cct2 knockdown decreased its major client protein, transducing β1 (Gβ1). Here we report the novel LCA mutations in CCTβ and the impact of chaperon disability by these mutations in cellular biology.

A GGGGCC hexanucleotide repeat expansion in the C9ORF72 gene has been identified as the most common genetic cause of amyotrophic lateral sclerosis and frontotemporal dementia. The repeat expansion undergoes unconventional translation to produce five dipeptide repeat proteins (DPRs). Although DPRs are thought to be neurotoxic, the molecular mechanism underlying the DPR-caused neurotoxicity has not been fully elucidated. The current study shows that poly-proline-arginine (poly-PR), the most toxic DPR in vitro, binds to and up-regulates nuclear paraspeckle assembly transcript 1 (NEAT1) that plays an essential role as a scaffold non-coding RNA during the paraspeckle formation. The CRISPR-assisted up-regulation of endogenous NEAT1 causes neurotoxicity. We also show that the poly-PR modulates the function of several paraspeckle-localizing heterogeneous nuclear ribonucleoproteins. Furthermore, dysregulated expression of TAR DNA-binding protein 43 (TDP-43) up-regulates NEAT1 expression and induces neurotoxicity. These results suggest that the increase in the paraspeckle formation may be involved in the poly-PR- and TDP-43-mediated neurotoxicity.

A GGGGCC repeat expansion in the C9ORF72 gene has been identified as the most common genetic cause of amyotrophic lateral sclerosis and frontotemporal dementia. The repeat expansion undergoes unconventional translation to produce dipeptide repeat (DPR) proteins. Although it has been reported that DPR proteins cause neurotoxicity, the underlying mechanism has not been fully elucidated. In this study, we have first confirmed that proline–arginine repeat protein (poly-PR) reduces levels of ribosomal RNA and causes neurotoxicity and found that the poly-PR-induced neurotoxicity is repressed by the acceleration of ribosomal RNA synthesis. These results suggest that the poly-PR-induced inhibition of ribosome biogenesis contributes to the poly-PR-induced neurotoxicity. We have further identified DEAD-box RNA helicases as poly-PR-binding proteins, the functions of which are inhibited by poly-PR. The enforced reduction in the expression of DEAD-box RNA helicases causes impairment of ribosome biogenesis and neuronal cell death. These results together suggest that poly-PR causes neurotoxicity by inhibiting the DEAD-box RNA helicase-mediated ribosome biogenesis.