Inactivation of Chibby affects function of motile airway cilia

Chibby (Cby) is a conserved component of the Wnt–β-catenin pathway. Cby physically interacts with β-catenin to repress its activation of transcription. To elucidate the function of Cby in vertebrates, we generated Cby−/− mice and found that after 2–3 d of weight loss, the majority of mice die before or around weaning. All Cby−/− mice develop rhinitis and sinusitis. When challenged with Pseudomonas aeruginosa isolates, Cby−/− mice are unable to clear the bacteria from the nasal cavity. Notably, Cby−/− mice exhibit a complete absence of mucociliary transport caused by a marked paucity of motile cilia in the nasal epithelium. Moreover, ultrastructural experiments reveal impaired basal body docking to the apical surface of multiciliated cells. In support of these phenotypes, endogenous Cby protein is localized at the base of cilia. As the phenotypes of Cby−/− mice bear striking similarities to primary ciliary dyskinesia, Cby−/− mice may prove to be a useful model for this condition

Lymphoid Enhancer Factor-1 Links Two Hereditary Leukemia Syndromes through Core-binding Factor α Regulation of ELA2

Two hereditary human leukemia syndromes are severe congenital neutropenia (SCN), caused by mutations in the gene ELA2, encoding the protease neutrophil elastase, and familial platelet disorder with acute myelogenous leukemia (AML), caused by mutations in the gene AML1, encoding the transcription factor core-binding factor α (CBFα). In mice, CBFα regulates the expression of ELA2, suggesting a common link for both diseases. However, gene-targeted mouse models have failed to reproduce either human disease, thus prohibiting further in vivo studies in mice. Here we investigate CBFα regulation of the human ELA2 promoter, taking advantage of bone marrow obtained from patients with either illness. In particular, we have identified novel ELA2 promoter substitutions (-199 C to A) within a potential motif for lymphoid enhancer factor-1 (LEF-1), a transcriptional mediator of Wnt/β-catenin signaling, in SCN patients. The LEF-1 motif lies adjacent to a potential CBFα binding site that is in a different position in human compared with mouse ELA2. We find that LEF-1 and CBFα co-activate ELA2 expression. In vitro, the high mobility group domain of LEF-1 interacts with the runt DNA binding and proline-, serine-, threonine-rich activation domains of CBFα. ELA2 transcript levels are up-regulated in bone marrow of an SCN patient with the -199 C to A substitution. Conversely, a mutation of the CBFα activation domain, found in a patient with familial platelet disorder with AML, fails to stimulate the ELA2 promoter in vitro, and bone marrow correspondingly demonstrates reduced ELA2 transcript. Observations in these complementary patients indicate that LEF-1 cooperates with CBFα to activate ELA2 in vivo and also suggest the possibility that up-regulating promoter mutations can contribute to SCN. Two hereditary AML predisposition syndromes may therefore intersect via LEF-1, potentially linking them to more generalized cancer mechanisms

Loss of the ciliary protein Chibby1 in mice leads to exocrine pancreatic degeneration and pancreatitis

Primary cilia protrude from the apical surface of many cell types and act as a sensory organelle that regulates diverse biological processes ranging from chemo- and mechanosensation to signaling. Ciliary dysfunction is associated with a wide array of genetic disorders, known as ciliopathies. Polycystic lesions are commonly found in the kidney, liver, and pancreas of ciliopathy patients and mouse models. However, the pathogenesis of the pancreatic phenotype remains poorly understood. Chibby1 (Cby1), a small conserved coiled-coil protein, localizes to the ciliary base and plays a crucial role in ciliogenesis. Here, we report that Cby1-knockout (KO) mice develop severe exocrine pancreatic atrophy with dilated ducts during early postnatal development. A significant reduction in the number and length of cilia was observed in Cby1-KO pancreta. In the adult Cby1-KO pancreas, inflammatory cell infiltration and fibrosis were noticeable. Intriguingly, Cby1-KO acinar cells showed an accumulation of zymogen granules (ZGs) with altered polarity. Moreover, isolated acini from Cby1-KO pancreas exhibited defective ZG secretion in vitro. Collectively, our results suggest that, upon loss of Cby1, concomitant with ciliary defects, acinar cells accumulate ZGs due to defective exocytosis, leading to cell death and progressive exocrine pancreatic degeneration after birth

Inactivation of Chibby affects function of motile airway cilia

Chibby (Cby) is a conserved component of the Wnt–β-catenin pathway. Cby physically interacts with β-catenin to repress its activation of transcription. To elucidate the function of Cby in vertebrates, we generated Cby−/− mice and found that after 2–3 d of weight loss, the majority of mice die before or around weaning. All Cby−/− mice develop rhinitis and sinusitis. When challenged with Pseudomonas aeruginosa isolates, Cby−/− mice are unable to clear the bacteria from the nasal cavity. Notably, Cby−/− mice exhibit a complete absence of mucociliary transport caused by a marked paucity of motile cilia in the nasal epithelium. Moreover, ultrastructural experiments reveal impaired basal body docking to the apical surface of multiciliated cells. In support of these phenotypes, endogenous Cby protein is localized at the base of cilia. As the phenotypes of Cby−/− mice bear striking similarities to primary ciliary dyskinesia, Cby−/− mice may prove to be a useful model for this condition

TAp73 is a central transcriptional regulator of airway multiciliogenesis.

Motile multiciliated cells (MCCs) have critical roles in respiratory health and disease and are essential for cleaning inhaled pollutants and pathogens from airways. Despite their significance for human disease, the transcriptional control that governs multiciliogenesis remains poorly understood. Here we identify TP73, a p53 homolog, as governing the program for airway multiciliogenesis. Mice with TP73 deficiency suffer from chronic respiratory tract infections due to profound defects in ciliogenesis and complete loss of mucociliary clearance. Organotypic airway cultures pinpoint TAp73 as necessary and sufficient for basal body docking, axonemal extension, and motility during the differentiation of MCC progenitors. Mechanistically, cross-species genomic analyses and complete ciliary rescue of knockout MCCs identify TAp73 as the conserved central transcriptional integrator of multiciliogenesis. TAp73 directly activates the key regulators FoxJ1, Rfx2, Rfx3, and miR34bc plus nearly 50 structural and functional ciliary genes, some of which are associated with human ciliopathies. Our results position TAp73 as a novel central regulator of MCC differentiation

P-value based visualization of codon usage data

Two important and not yet solved problems in bacterial genome research are the identification of horizontally transferred genes and the prediction of gene expression levels. Both problems can be addressed by multivariate analysis of codon usage data. In particular dimensionality reduction methods for visualization of multivariate data have shown to be effective tools for codon usage analysis. We here propose a multidimensional scaling approach using a novel similarity measure for codon usage tables. Our probabilistic similarity measure is based on P-values derived from the well-known chi-square test for comparison of two distributions. Experimental results on four microbial genomes indicate that the new method is well-suited for the analysis of horizontal gene transfer and translational selection. As compared with the widely-used correspondence analysis, our method did not suffer from outlier sensitivity and showed a better clustering of putative alien genes in most cases

Pygo2 expands mammary progenitor cells by facilitating histone H3 K4 methylation

Recent studies have unequivocally identified multipotent stem/progenitor cells in mammary glands, offering a tractable model system to unravel genetic and epigenetic regulation of epithelial stem/progenitor cell development and homeostasis. In this study, we show that Pygo2, a member of an evolutionarily conserved family of plant homeo domain–containing proteins, is expressed in embryonic and postnatal mammary progenitor cells. Pygo2 deficiency, which is achieved by complete or epithelia-specific gene ablation in mice, results in defective mammary morphogenesis and regeneration accompanied by severely compromised expansive self-renewal of epithelial progenitor cells. Pygo2 converges with Wnt/β-catenin signaling on progenitor cell regulation and cell cycle gene expression, and loss of epithelial Pygo2 completely rescues β-catenin–induced mammary outgrowth. We further describe a novel molecular function of Pygo2 that is required for mammary progenitor cell expansion, which is to facilitate K4 trimethylation of histone H3, both globally and at Wnt/β-catenin target loci, via direct binding to K4-methyl histone H3 and recruiting histone H3 K4 methyltransferase complexes

Presenilin Controls CBP Levels in the Adult Drosophila Central Nervous System

Background: Dominant mutations in both human Presenilin (Psn) genes have been correlated with the formation of amyloid plaques and development of familial early-onset Alzheimer’s disease (AD). However, a definitive mechanism whereby plaque formation causes the pathology of familial and sporadic forms of AD has remained elusive. Recent discoveries of several substrates for Psn protease activity have sparked alternative hypotheses for the pathophysiology underlying AD. CBP (CREB-binding protein) is a haplo-insufficient transcriptional co-activator with histone acetly-transferase (HAT) activity that has been proposed to be a downstream target of Psn signaling. Individuals with altered CBP have cognitive deficits that have been linked to several neurological disorders. Methodology/Principal Findings: Using a transgenic RNA-interference strategy to selectively silence CBP, Psn, and Notch in adult Drosophila, we provide evidence for the first time that Psn is required for normal CBP levels and for maintaining specific global acetylations at lysine 8 of histone 4 (H4K8ac) in the central nervous system (CNS). In addition, flies conditionally compromised for the adult-expression of CBP display an altered geotaxis behavior that may reflect a neurological defect. Conclusions/Significance: Our data support a model in which Psn regulates CBP levels in the adult fly brain in a manner that is independent of Notch signaling. Although we do not understand the molecular mechanism underlying th

Wdpcp, a PCP Protein Required for Ciliogenesis, Regulates Directional Cell Migration and Cell Polarity by Direct Modulation of the Actin Cytoskeleton

Planar cell polarity (PCP) regulates cell alignment required for collective cell movement during embryonic development. This requires PCP/PCP effector proteins, some of which also play essential roles in ciliogenesis, highlighting the long-standing question of the role of the cilium in PCP. Wdpcp, a PCP effector, was recently shown to regulate both ciliogenesis and collective cell movement, but the underlying mechanism is unknown. Here we show Wdpcp can regulate PCP by direct modulation of the actin cytoskeleton. These studies were made possible by recovery of a Wdpcp mutant mouse model. Wdpcp-deficient mice exhibit phenotypes reminiscent of Bardet-Biedl/Meckel-Gruber ciliopathy syndromes, including cardiac outflow tract and cochlea defects associated with PCP perturbation. We observed Wdpcp is localized to the transition zone, and in Wdpcp-deficient cells, Sept2, Nphp1, and Mks1 were lost from the transition zone, indicating Wdpcp is required for recruitment of proteins essential for ciliogenesis. Wdpcp is also found in the cytoplasm, where it is localized in the actin cytoskeleton and in focal adhesions. Wdpcp interacts with Sept2 and is colocalized with Sept2 in actin filaments, but in Wdpcp-deficient cells, Sept2 was lost from the actin cytoskeleton, suggesting Wdpcp is required for Sept2 recruitment to actin filaments. Significantly, organization of the actin filaments and focal contacts were markedly changed in Wdpcp-deficient cells. This was associated with decreased membrane ruffling, failure to establish cell polarity, and loss of directional cell migration. These results suggest the PCP defects in Wdpcp mutants are not caused by loss of cilia, but by direct disruption of the actin cytoskeleton. Consistent with this, Wdpcp mutant cochlea has normal kinocilia and yet exhibits PCP defects. Together, these findings provide the first evidence, to our knowledge, that a PCP component required for ciliogenesis can directly modulate the actin cytoskeleton to regulate cell polarity and directional cell migration