永岡 唯宏

Although the core constituents of the Wnt/planar cell polarity (PCP) signaling have been extensively studied, their downstream molecules and protein-protein interactions have not yet been fully elucidated. Here, we show genetic and molecular evidence that the PCP factor, Vangl2, functionally interacts with the cell-cell adhesion molecule, N-cadherin (also known as Cdh2), for typical PCP-dependent neural development. Vangl2 and N-cadherin physically interact in the neural plates undergoing convergent extension. Unlike monogenic heterozygotes, digenic heterozygous mice with Vangl2 and Cdh2 mutants exhibited defects in neural tube closure and cochlear hair cell orientation. Despite this genetic interaction, neuroepithelial cells derived from the digenic heterozygotes did not show additive changes from the monogenic heterozygotes of Vangl2 in the RhoA-ROCK-Mypt1 and c-Jun N-terminal kinase (JNK)-Jun pathways of Wnt/PCP signaling. Thus, cooperation between Vangl2 and N-cadherin is at least partly via direct molecular interaction; it is essential for the planar polarized development of neural tissues but not significantly associated with RhoA or JNK pathways.

Ribosomal RNA (rRNA), the most abundant non-coding RNA species, is a major component of the ribosome. Impaired ribosome biogenesis causes the dysfunction of protein synthesis and diseases called "ribosomopathies," including genetic disorders with cancer risk. However, the potential role of rRNA gene (rDNA) alterations in cancer is unknown. We investigated germline and somatic single-nucleotide variants (SNVs) in the rDNA promoter region (positions -248 to +100, relative to the transcription start site) in 82 lung adenocarcinomas (LUAC). Twenty-nine tumors (35.4%) carried germline SNVs, and eight tumors (9.8%) harbored somatic SNVs. Interestingly, the presence of germline SNVs between positions +1 and +100 (n = 12; 14.6%) was associated with significantly shorter recurrence-free survival (RFS) and overall survival (OS) by univariate analysis (p < 0.05, respectively), and was an independent prognostic factor for RFS and OS by multivariate analysis. LUAC cell line PC9, carrying rDNA promoter SNV at position +49, showed significantly higher ribosome biogenesis than H1650 cells without SNV. Upon nucleolar stress induced by actinomycin D, PC9 retained significantly higher ribosome biogenesis than H1650. These results highlight the possible functional role of SNVs at specific sites of the rDNA promoter region in ribosome biogenesis, the progression of LUAC, and their potential prognostic value.

The PET and LIM domain-containing protein, Prickle, plays a key role in planar cell polarity (PCP) in Drosophila. It has been reported that mutations in the PRICKLE2 gene, which encodes one of the human orthologues of Prickle, are associated with human diseases such as epilepsy and autism spectrum disorder. To develop preventive and therapeutic strategies for these intractable diseases, we studied the regulation of Prickle2 protein levels in transfected HEK293T cells. Prickle2 levels were negatively regulated by a physical interaction with another PCP protein, Van Gogh-like 2 (Vangl2). The Vangl2-mediated reduction in Prickle2 levels was, at least in part, relieved by proteasome inhibitors or by functional inhibition of the Cullin-1 E3 ubiquitin ligase. Furthermore, the expression of Vangl2 enhanced the polyubiquitination of Prickle2. This ubiquitination was partially blocked by co-expression of a ubiquitin mutant, which cannot be polymerised through their Lys48 residue to induce target proteins toward proteasomal degradation. Together, these results suggest that Prickle2 is polyubiquitinated by the Vangl2 interaction in a Cullin-1-dependent manner to limit its expression levels. This regulation may play a role in the local and temporal fine-tuning of Prickle protein levels during PCP signal-dependent cellular behaviours.

The excitatory postsynaptic region of the vertebrate hippocampus is usually compartmentalized into the postsynaptic density (PSD) and N-cadherin-rich domain, which is important for synaptic adhesion. However, the molecular mechanisms underlying the compartment formation are unknown. In the present report, we show that the planar cell polarity (PCP) protein Van Gogh-like 2 (Vangl2) plays a role in this regionalization. In cultured rat hippocampal neurons that were subjected to Vangl2 expression silencing, the formed clusters of PSD-95, one of the major scaffolding proteins in PSD, tended to overlap with those of N-cadherin. Further, in the dendrites of these neurons, the immunofluorescence of PSD-95 was to some extent diffused, without a significant change in the total signal. Because Vangl2 physically interacts with both PSD-95 and N-cadherin in vivo, these results suggest that a PCP-related direct molecular mechanism underlies the horizontal polarization of the postsynaptic regions. (c) 2015 Elsevier Ireland Ltd. All rights reserved.

Postsynaptic density-95/Discs large/Zonula occludens-1 (PDZ) domain-mediated protein interactions play pivotal roles in various molecular biological events, including protein localisation, assembly, and signal transduction. Although the vertebrate regulator of planar cell polarity Van Gogh-like 2 (Vangl2) was recently described as a postsynaptic molecule with a PDZ-binding motif, the role of its PDZ interaction at the synapse is unknown. In this report, we demonstrate that the PDZ interaction was dispensable for the normal cluster formation of Vangl2 and not absolutely required for the synapse-associated localisation of Vangl2 in cultured hippocampal neurons. We further showed that the synaptic localisation of Vangl2 was categorised into two types: overlapping co-localisation with postsynaptic density (PSD)-95 or highly correlated but complementary pattern of association with PSD-95. Only the former was significantly sensitive to deletion of the PDZ-binding motif. In addition, the PDZ interaction enhanced the protein interactions between PSD-95 and Prickle2, which is another planar cell polarity factor that is localised at the postsynaptic density. Taken together with our recent report that the density of PSD-95 clusters was reduced in Vangl2-silenced neurons, these results suggest that Vangl2 determines the complex formation and clustering of postsynaptic molecules for synaptogenesis in mammalian brains.