Complex interactions between genes controlling trafficking in primary cilia

Cilia-associated human genetic disorders are striking in the diversity of their abnormalities and their complex inheritance. Inactivation of the retrograde ciliary motor by mutations in DYNC2H1 causes skeletal dysplasias that have strongly variable expressivity. Here we define previously unknown genetic relationships between Dync2h1 and other genes required for ciliary trafficking. Mutations in mouse Dync2h1 disrupt cilia structure, block Sonic hedgehog signaling and cause midgestation lethality. Heterozygosity for Ift172, a gene required for anterograde ciliary trafficking, suppresses cilia phenotypes, Sonic hedgehog signaling defects and early lethality of Dync2h1 homozygotes. Ift122, like Dync2h1, is required for retrograde ciliary trafficking, but reduction of Ift122 gene dosage also suppresses the Dync2h1 phenotype. These genetic interactions illustrate the cell biology underlying ciliopathies and argue that mutations in intraflagellar transport genes cause their phenotypes because of their roles in cilia architecture rather than direct roles in signaling.     

The coiled-coil domain containing protein CCDC40 is essential for motile cilia function and left-right axis formation

Primary ciliary dyskinesia (PCD) is a genetically heterogeneous autosomal recessive disorder characterized by recurrent infections of the respiratory tract associated with the abnormal function of motile cilia. Approximately half of individuals with PCD also have alterations in the left-right organization of their internal organ positioning, including situs inversus and situs ambiguous (Kartagener's syndrome). Here, we identify an uncharacterized coiled-coil domain containing a protein, CCDC40, essential for correct left-right patterning in mouse, zebrafish and human. In mouse and zebrafish, Ccdc40 is expressed in tissues that contain motile cilia, and mutations in Ccdc40 result in cilia with reduced ranges of motility. We further show that CCDC40 mutations in humans result in a variant of PCD characterized by misplacement of the central pair of microtubules and defective assembly of inner dynein arms and dynein regulatory complexes. CCDC40 localizes to motile cilia and the apical cytoplasm and is required for axonemal recruitment of CCDC39, disruption of which underlies a similar variant of PCD.     

A genetic interaction network of five genes for human polycystic kidney and liver diseases defines polycystin-1 as the central determinant of cyst formation

Autosomal dominant polycystic liver disease results from mutations in PRKCSH or SEC63. The respective gene products, glucosidase IIβ and SEC63p, function in protein translocation and quality control pathways in the endoplasmic reticulum. Here we show that glucosidase IIβ and Sec63p are required in mice for adequate expression of a functional complex of the polycystic kidney disease gene products, polycystin-1 and polycystin-2. We find that polycystin-1 is the rate-limiting component of this complex and that there is a dose-response relationship between cystic dilation and levels of functional polycystin-1 following mutation of Prkcsh or Sec63. Reduced expression of polycystin-1 also serves to sensitize the kidney to cyst formation resulting from mutations in Pkhd1, the recessive polycystic kidney disease gene. Finally, we show that proteasome inhibition increases steady-state levels of polycystin-1 in cells lacking glucosidase IIβ and that treatment with a proteasome inhibitor reduces cystic disease in orthologous gene models of human autosomal dominant polycystic liver disease.     

Genome-wide association study reveals three susceptibility loci for common migraine in the general population

Migraine is a common, heterogeneous and heritable neurological disorder. Its pathophysiology is incompletely understood, and its genetic influences at the population level are unknown. In a population-based genome-wide analysis including 5,122 migraineurs and 18,108 non-migraineurs, rs2651899 (1p36.32, PRDM16), rs10166942 (2q37.1, TRPM8) and rs11172113 (12q13.3, LRP1) were among the top seven associations (P < 5 × 10(-6)) with migraine. These SNPs were significant in a meta-analysis among three replication cohorts and met genome-wide significance in a meta-analysis combining the discovery and replication cohorts (rs2651899, odds ratio (OR) = 1.11, P = 3.8 × 10(-9); rs10166942, OR = 0.85, P = 5.5 × 10(-12); and rs11172113, OR = 0.90, P = 4.3 × 10(-9)). The associations at rs2651899 and rs10166942 were specific for migraine compared with non-migraine headache. None of the three SNP associations was preferential for migraine with aura or without aura, nor were any associations specific for migraine features. TRPM8 has been the focus of neuropathic pain models, whereas LRP1 modulates neuronal glutamate signaling, plausibly linking both genes to migraine pathophysiology.     

Disruption of mouse Slx4, a regulator of structure-specific nucleases, phenocopies Fanconi anemia

The evolutionarily conserved SLX4 protein, a key regulator of nucleases, is critical for DNA damage response. SLX4 nuclease complexes mediate repair during replication and can also resolve Holliday junctions formed during homologous recombination. Here we describe the phenotype of the Btbd12 knockout mouse, the mouse ortholog of SLX4, which recapitulates many key features of the human genetic illness Fanconi anemia. Btbd12-deficient animals are born at sub-Mendelian ratios, have greatly reduced fertility, are developmentally compromised and are prone to blood cytopenias. Btbd12(-/-) cells prematurely senesce, spontaneously accumulate damaged chromosomes and are particularly sensitive to DNA crosslinking agents. Genetic complementation reveals a crucial requirement for Btbd12 (also known as Slx4) to interact with the structure-specific endonuclease Xpf-Ercc1 to promote crosslink repair. The Btbd12 knockout mouse therefore establishes a disease model for Fanconi anemia and genetically links a regulator of nuclease incision complexes to the Fanconi anemia DNA crosslink repair pathway.     

Systematic documentation and analysis of human genetic variation in hemoglobinopathies using the microattribution approach

We developed a series of interrelated locus-specific databases to store all published and unpublished genetic variation related to hemoglobinopathies and thalassemia and implemented microattribution to encourage submission of unpublished observations of genetic variation to these public repositories. A total of 1,941 unique genetic variants in 37 genes, encoding globins and other erythroid proteins, are currently documented in these databases, with reciprocal attribution of microcitations to data contributors. Our project provides the first example of implementing microattribution to incentivise submission of all known genetic variation in a defined system. It has demonstrably increased the reporting of human variants, leading to a comprehensive online resource for systematically describing human genetic variation in the globin genes and other genes contributing to hemoglobinopathies and thalassemias. The principles established here will serve as a model for other systems and for the analysis of other common and/or complex human genetic diseases.     

On glioblastoma and the search for a cure: where do we stand?

Although brain tumours have been documented and recorded since the nineteenth century, 2016 marked 90 years since Percival Bailey and Harvey Cushing coined the term “glioblastoma multiforme”. Since that time, although extensive developments in diagnosis and treatment have been made, relatively little improvement on prognosis has been achieved. The resilience of GBM thus makes treating this tumour one of the biggest challenges currently faced by neuro-oncology. Aggressive and robust development, coupled with difficulties of complete resection, drug delivery and therapeutic resistance to treatment are some of the main issues that this nemesis presents today. Current treatments are far from satisfactory with poor prognosis, and focus on palliative management rather than curative intervention. However, therapeutic research leading to developments in novel treatment stratagems show promise in combating this disease. Here we present a review on GBM, looking at the history and advances which have shaped neurosurgery over the last century that cumulate to the present day management of GBM, while also exploring future perspectives in treatment options that could lead to new treatments on the road to a cure.     

Germline-specific dgcr8 knockout in zebrafish using a BACK approach

Zebrafish is an important model to study developmental biology and human diseases. However, an effective approach to achieve spatial and temporal gene knockout in zebrafish has not been well established. In this study, we have developed a new approach, namely bacterial artificial chromosome-rescue-based knockout (BACK), to achieve conditional gene knockout in zebrafish using the Cre/loxP system. We have successfully deleted the DiGeorge syndrome critical region gene 8 (dgcr8) in zebrafish germ line and demonstrated that the maternal-zygotic dgcr8 (MZdgcr8) embryos exhibit MZdicer-like phenotypes with morphological defects which could be rescued by miR-430, indicating that canonical microRNAs play critical role in early development. Our findings establish that Cre/loxP-mediated tissue-specific gene knockout could be achieved using this BACK strategy and that canonical microRNAs play important roles in early embryonic development in zebrafish.     

Epigenetics: a link between addiction and social environment

The detrimental effects of drug abuse are apparently not limited to individuals but may also impact the vulnerability of their progenies to develop addictive behaviours. Epigenetic signatures, early life experience and environmental factors, converge to influence gene expression patterns in addiction phenotypes and consequently may serve as mediators of behavioural trait transmission between generations. The majority of studies investigating the role of epigenetics in addiction do not consider the influence of social interactions. This shortcoming in current experimental approaches necessitates developing social models that reflect the addictive behaviour in a free-living social environment. Furthermore, this review also reports on the advancement of interventions for drug addiction and takes into account the emerging roles of histone deacetylase (HDAC) inhibitors in the etiology of drug addiction and that HDAC may be a potential therapeutic target at nucleosomal level to improve treatment outcomes.     

Basal autophagy is involved in the degradation of the ERAD component EDEM1

Little is known about the fate of machinery proteins of the protein quality control and endoplasmic reticulum(ER)-associated degradation (ERAD). We investigated the degradation of the ERAD component EDEM1, which directs overexpressed misfolded glycoproteins to degradation. Endogenous EDEM1 was studied since EDEM1 overexpression not only resulted in inappropriate occurrence throughout the ER but also caused cytotoxic effects. Proteasome inhibitors had no effect on the clearance of endogenous EDEM1 in non-starved cells. However, EDEM1 could be detected by immunocytochemistry in autophagosomes and biochemically in LC3 immuno-purified autophagosomes. Furthermore, influencing the lysosome-autophagy pathway by vinblastine or pepstatin A/E64d and inhibiting autophagosome formation by 3-methyladenine or ATGs short interfering RNA knockdown stabilized EDEM1. Autophagic degradation involved removal of cytosolic Triton X-100-insoluble deglycosylated EDEM1, but not of EDEM1-containing ER cisternae. Our studies demonstrate that endogenous EDEM1 in cells not stressed by the expression of a transgenic misfolded protein reaches the cytosol and is degraded by basal autophagy. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. Received 15 January 2009; received after revision 16 February 2009; accepted 17 February 2009 V. Le Fourn, K. Gaplovska-Kysela: These authors equally contributed to this work.