The evolutionarily conserved polarity proteins PAR-3, atypical protein kinase C (aPKC) and PAR-6 critically regulate the apical membrane development required for epithelial organ development. However, the molecular mechanisms underlying their roles remain to be clarified. We demonstrate that PAR-3 knockdown in MDCK cells retards apical protein delivery to the plasma membrane, and eventually leads to mislocalized apical domain formation at intercellular regions in both twodimensional and three-dimensional culture systems. The defects in PAR-3 knockdown cells are efficiently rescued by wild-type PAR-3, but not by a point mutant (S827/829A) that lacks the ability to interact with aPKC, indicating that formation of the PAR-3-aPKC-PAR-6 complex is essential for apical membrane development. This is in sharp contrast with tight junction maturation, which does not necessarily depend on the aPKC-PAR-3 interaction, and indicates that the two fundamental processes essential for epithelial polarity are differentially regulated by these polarity proteins. Importantly, highly depolarized cells accumulate aPKC and PAR-6, but not PAR-3, on apical protein-containing vacuoles, which become targeted to PAR-3-positive primordial cell-cell contact sites during the initial stage of the repolarization process. Therefore, formation of the PAR-3-aPKC-PAR-6 complex might be required for targeting of not only the aPKC-PAR-6 complex but also of apical protein carrier vesicles to primordial junction structures. Supplementary material available online at

Epithelial cells possess apical-basolateral polarity and form tight junctions (TJs) at the apical-lateral border, separating apical and basolateral membrane domains. The PAR3-aPKC-PAR6 complex plays a central role in TJ formation and apical domain development during tissue morphogenesis. Inactivation and overactivation of aPKC kinase activity disrupts membrane polarity. The mechanism that suppresses active aPKC is unknown. KIBRA, an upstream regulator of the Hippo pathway, regulates tissue size in Drosophila and can bind to aPKC. However, the relationship between KIBRA and the PAR3-aPKC-PAR6 complex remains unknown. We report that KIBRA binds to the PAR3-aPKC-PAR6 complex and localizes at TJs and apical domains in epithelial tissues and cells. The knockdown of KIBRA causes expansion of the apical domain in MDCK three-dimensional cysts and suppresses the formation of apical-containing vacuoles through enhanced de novo apical exocytosis. These phenotypes are restored by inhibition of aPKC. In addition, KIBRA directly inhibits the kinase activity of aPKC in vitro. These results strongly support the notion that KIBRA regulates epithelial cell polarity by suppressing apical exocytosis through direct inhibition of aPKC kinase activity in the PAR3-aPKC-PAR6 complex.

Epithelial apicobasal polarity has fundamental roles in epithelial physiology and morphogenesis. The PAR complex, comprising PAR-3, PAR-6 and aPKC, is involved in determining cell polarity in various biological contexts, including in epithelial cells. However, it is not fully understood how the PAR complex induces apicobasal polarity. In this study, we show that PAR-3 regulates the protein expression of Girdin (GIV/CCDC88A), a guanine nucleotide exchange factor (GEF) for heterotrimeric Gαi subunits, at the transcriptional level by cooperating with the AP-2 transcription factor. In addition, we confirmed that PAR-3 physically interacts with Girdin, and show that Girdin, together with the Gαi3, controls tight junction formation, apical domain development and actin organization downstream of PAR-3. Taken together, our findings suggest that transcriptional upregulation of Girdin and Girdin–Gαi3 signaling play crucial roles in regulating epithelial apicobasal polarity via the PAR complex.

Fusarium oxysporum f. sp. cepae causes Fusarium basal rot in onion (common onion) and Fusarium wilt in Welsh onion. Although these diseases have been detected in various areas in Japan, knowledge about the genetic and pathogenic variability of F. oxysporum f. sp. cepae is very limited. In this study, F. oxysporum f. sp. cepae was isolated from onion and Welsh onion grown in 12 locations in Japan, and a total of 55 F. oxysporum f. sp. cepae isolates (27 from onion and 28 from Welsh onion) were characterized based on their rDNA intergenic spacer (IGS) and translation elongation factor-1α (EF-1α) nucleotide sequences, vegetative compatibility groups (VCGs), and the presence of the SIX (secreted in xylem) homologs. Phylogenetic analysis of IGS sequences showed that these isolates were grouped into eight clades (A to H), and 20 onion isolates belonging to clade H were monophyletic and assigned to the same VCG. All the IGS-clade H isolates possessed homologs of SIX3, SIX5, and SIX7. The SIX3 homolog was located on a 4 Mb-sized chromosome in the IGS-clade H isolates. Pathogenicity tests using onion seedlings showed that all the isolates with high virulence were in the IGS-clade H. These results suggest that F. oxysporum f. sp. cepae isolates belonging to the IGS-clade H are genetically and pathogenically different from those belonging to the other IGS clades.

Mediator is a coregulatory complex that regulates transcription of Pol II-dependent genes. Previously, we showed that human Mediator subunit MED26 plays a role in the recruitment of Super Elongation Complex (SEC) or Little Elongation Complex (LEC) to regulate the expression of certain genes. MED26 plays a role in recruiting SEC to protein-coding genes including c-myc and LEC to small nuclear RNA (snRNA) genes. However, how MED26 engages SEC or LEC to regulate distinct genes is unclear. Here, we provide evidence that MED26 recruits LEC to modulate transcription termination of non-polyadenylated transcripts including snRNAs and mRNAs encoding replication-dependent histone (RDH) at Cajal bodies. Our findings indicate that LEC recruited by MED26 promotes efficient transcription termination by Pol II through interaction with CBC-ARS2 and NELF/DSIF, and promotes 3′ end processing by enhancing recruitment of Integrator or Heat Labile Factor to snRNA or RDH genes, respectively.