Mutations in PNKP Cause Recessive Ataxia with Oculomotor Apraxia Type 4

Hereditary autosomal-recessive cerebellar ataxias are a genetically and clinically heterogeneous group of disorders. We used homozygosity mapping and exome sequencing to study a cohort of nine Portuguese families who were identified during a nationwide, population-based, systematic survey as displaying a consistent phenotype of recessive ataxia with oculomotor apraxia (AOA). The integration of data from these analyses led to the identification of the same homozygous PNKP (polynucleotide kinase 3′-phosphatase) mutation, c.1123G>T (p.Gly375Trp), in three of the studied families. When analyzing this particular gene in the exome sequencing data from the remaining cohort, we identified homozygous or compound-heterozygous mutations in five other families. PNKP is a dual-function enzyme with a key role in different pathways of DNA-damage repair. Mutations in this gene have previously been associated with an autosomal-recessive syndrome characterized by microcephaly; early-onset, intractable seizures; and developmental delay (MCSZ). The finding of PNKP mutations associated with recessive AOA extends the phenotype associated with this gene and identifies a fourth locus that causes AOA. These data confirm that MCSZ and some forms of ataxia share etiological features, most likely reflecting the role of PNKP in DNA-repair mechanisms.     

Influence of growth environment on the phosphoenolpyruvate: Glucose phosphotransferase activities of Escherichia coli and Klebsiella aerogenes: A comparative study

A consistent difference was found between glucose-limited cultures of Escherichia coli and Klebsiella aerogenes strains in the manner which their apparent cellular content of glucose: phosphoenolpyruvate phosphotransferase (glucose-PTS) varied with growth rate. With the former strains, activity increased as a function of growth rate; in the latter it decreased. However, under glucose-sufficient conditions (potassium-or ammonia-limitation) both species behaved similarly; the glucose-PTS activity was lower and bore no obvious relationship to the rate of glucose consumption expressed by the growing culture. These results are discussed in relation to the role of glucose as a regulator of glucose-PTS synthesis, and to the likely contribution which the glucose-PTS makes to the overall rate of glucose uptake, particularly by cells growing in glucose-sufficient environments.Abbreviation Glucose-PTS phosphoenolpyruvate phosphotransferase From May to November 1978 on study leave in the University of Amsterdam     

A bidirectional antagonism between aPKC and Yurt regulates epithelial cell polarity

During epithelial cell polarization, Yurt (Yrt) is initially confined to the lateral membrane and supports the stability of this membrane domain by repressing the Crumbs-containing apical machinery. At late stages of embryogenesis, the apical recruitment of Yrt restricts the size of the apical membrane. However, the molecular basis sustaining the spatiotemporal dynamics of Yrt remains undefined. In this paper, we report that atypical protein kinase C (aPKC) phosphorylates Yrt to prevent its premature apical localization. A nonphosphorylatable version of Yrt dominantly dismantles the apical domain, showing that its aPKC-mediated exclusion is crucial for epithelial cell polarity. In return, Yrt counteracts aPKC functions to prevent apicalization of the plasma membrane. The ability of Yrt to bind and restrain aPKC signaling is central for its role in polarity, as removal of the aPKC binding site neutralizes Yrt activity. Thus, Yrt and aPKC are involved in a reciprocal antagonistic regulatory loop that contributes to segregation of distinct and mutually exclusive membrane domains in epithelial cells.     

Differential expression, function and response to inflammatory stimuli of 11β-hydroxysteroid dehydrogenase type 1 in human fibroblasts: a mechanism for tissue-specific regulation of inflammation

Stromal cells such as fibroblasts play an important role in defining tissue-specific responses during the resolution of inflammation. We hypothesized that this involves tissue-specific regulation of glucocorticoids, mediated via differential regulation of the enzyme 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1). Expression, activity and function of 11β-HSD1 was assessed in matched fibroblasts derived from various tissues (synovium, bone marrow and skin) obtained from patients with rheumatoid arthritis or osteoarthritis. 11β-HSD1 was expressed in fibroblasts from all tissues but mRNA levels and enzyme activity were higher in synovial fibroblasts (2-fold and 13-fold higher mRNA levels in dermal and synovial fibroblasts, respectively, relative to bone marrow). Expression and activity of the enzyme increased in all fibroblasts following treatment with tumour necrosis factor-α or IL-1β (bone marrow: 8-fold and 37-fold, respectively, compared to vehicle; dermal fibroblasts: 4-fold and 14-fold; synovial fibroblasts: 7-fold and 31-fold; all P < 0.01 compared with vehicle). Treatment with IL-4 or interferon-γ was without effect, and there was no difference in 11β-HSD1 expression between fibroblasts (from any site) obtained from patients with rheumatoid arthritis or osteoarthritis. In the presence of 100 nmol/l cortisone, IL-6 production – a characteristic feature of synovial derived fibroblasts – was significantly reduced in synovial but not dermal or bone marrow fibroblasts. This was prevented by co-treatment with an 11β-HSD inhibitor, emphasizing the potential for autocrine activation of glucocorticoids in synovial fibroblasts. These data indicate that differences in fibroblast-derived glucocorticoid production (via the enzyme 11β-HSD1) between cells from distinct anatomical locations may play a key role in the predeliction of certain tissues to develop persistent inflammation.     

Bacillus thuringiensis and control of plant pests

Summary The feasibility ofB. thuringiensis as an economic insect control agent is dependent upon various factors. Consideration of three situations. whereB. thuringiensis could be used illustrates the interaction of these factors, and their contribution to determining the success or failure of the organism as an insecticide. Apple crops suffer from attack by a complex of insect pests, many of which are very susceptible to theB. thuringiensis toxin. However chemical control methods are preferred by growers in order to satisfy the consumers' demand for top quality blemish-free fruit. By contrast cotton growers are beset with problems of pest resistance to chemical insecticides. This problem is tackled by using pest management strategies which orchestrate all possible methods of control. Such a situation is ideally suited toB. thuringiensis products, but as yet they have proven to be of inadequate efficacy. The softwood forest industry is an example whereB. thuringiensis is both needed and effective. The pest complex is relatively simple, and there is public concern about the health risks of chemical sprays. Recent developments of high performance formulations means thatB. thuringiensis is an economic, cost-effective, biological alternative to chemical control for forestry pests.

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Resumen La posibilidad de utilizarB. thuringiensis para el control de insectos de una forma económica depende de varios factores. La consideración de tres situaciones distintas en las queB. thuringiensis puede ser utilizado ilustra la interacción de estos factores y su contribución al éxito o fracaso del organismo como insecticida. El cultivo de manzanas sufre el ataque de una serie de plagas de insectos, muchas de las cuales son susceptibles a la toxina deB. thuringiensis. Sin embargo, los agricultores prefieren utilizar métodos de control químico a fin de satisfacer la demanda del consumidor por un fruto de primera calidad sin marcas ni señales. Por el contrario, los cultivadores de algodon estan sensibilizados a los problemas de la aparición de plagas resistentes a los insecticidas químicos. Este problema se ha abordado utilizando sistemas de control integrado que aglutinan todos los posibles métodos de control. Esta situación parece la ideal para el uso de productos a base deB. thuringiensis, pero hasta ahora estos han resultado ser poco eficaces. La industria de la madera blanda constituye en ejemplo en el cualB. thuringiensis es a la vez necesario y efectivo. Las plagas en cuestión forman un sistema relativamente simple y los efectos de los sprays químicos sobre la salud pública son causa de preocupación. Los desarrollos recientes en formulaciones de elevada eficacia convierten aB. thuringiensis en una alternativa biológica económicamente factible frente al control químico de plagas forestales.

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Résumé La possibilité d'utiliser économiquementB. thuringiensis dans la lutte contre les insectes nuisibles dépend de facteurs divers. L'examen de trois situations dans lesquellesB. thuringiensis peut être utilisé illustre l'interaction de ces facteurs et leur contribution dans le succès ou l'échec de cet organisme comme insecticide. Les récoltes de pommes sont menacées par les attaques d'un mélange complexe d'insectes, dont beaucoup sont sensibles à la toxine deB. thuringiensis. Cependant, les horticulteurs préfèrent les méthodes chimiques à cause de l'exigence des consommateurs pour des fruits de haute qualité et sans aucune tache. Par contre, les cultivateurs de coton sont obsédés par les problèmes de résistance aux insecticides chimiques. Ce problème est affronté par des stratégies faisant appel à tous les moyens de lutte possibles. C'est une situation convenant bien aux produits qui contiennentB. thuringiensis, mais jusqu'ici ces produits se sont montrés inefficaces. L'industrie du bois tendre forestier est toutefois un exemple de cas oùB. thuringiensis est à la fois nécessaire et efficace. En effet, le complexe des insectes agressifs est relativement simple et il existe en outre une prévention du public contre les risques sanitaires des pulvérisations de produits chimiques. La mise au point récente de formulations à haute performance démontre queB. thuringiensis est, pour la lutte contre les insectes nuisibles des forêts, une alternative économique et bon-marché.

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Invited paper presented at the VII International Conference on the Global Impacts of Applied Microbiology, Helsiki, 12–16 August 1985. Session 8     

Estimation of pairwise relatedness between individuals and characterization of isolation-by-distance processes using dominant genetic markers

A new estimator of the pairwise relatedness coefficient between individuals adapted to dominant genetic markers is developed. This estimator does not assume genotypes to be in Hardy-Weinberg proportions but requires a knowledge of the departure from these proportions (i.e. the inbreeding coefficient). Simulations show that the estimator provides accurate estimates, except for some particular types of individual pairs such as full-sibs, and performs better than a previously developed estimator. When comparing marker-based relatedness estimates with pedigree expectations, a new approach to account for the change of the reference population is developed and shown to perform satisfactorily. Simulations also illustrate that this new relatedness estimator can be used to characterize isolation by distance within populations, leading to essentially unbiased estimates of the neighbourhood size. In this context, the estimator appears fairly robust to moderate errors made on the assumed inbreeding coefficient. The analysis of real data sets suggests that dominant markers (random amplified polymorphic DNA, amplified fragment length polymorphism) may be as valuable as co-dominant markers (microsatellites) in studying microgeographic isolation-by-distance processes. It is argued that the estimators developed should find major applications, notably for conservation biology.     

Web database of molecular genetic data from fish stocks

Much research at the national and international level has been devoted to the development of several genetic methods for use in the characterization of fish stocks. We have developed a database that collates data from population genetic studies of fish. This database is accessible to researchers and control authorities on the Internet and should serve as a repository for genetic information of commercially important fish species. The prototype database has been developed in such a way that the new information can easily be updated by researchers in the field, who can submit their own data. The site can be found at http://fishgen.jrc.it.     

Characterization of terminal NeuNAcalpha2-3Galbeta1-4GlcNAc sequence in lipooligosaccharides of Neisseria meningitidis

Group B and C Neisseria meningitidis are the major cause of meningococcal disease in the United States and in Europe. N . meningitidis lipooligosaccharide (LOS), a major surface antigen, can be divided into 12 immunotypes of which L1 through L8 were found among Group B and C organisms. Groups B and C but not Group A may sialylate their LOSs with N-acetylneuraminic acid (NeuNAc) at the nonreducing end because they synthesize CMP-NeuNAc. Using sialic acid-galactose binding lectins as probes in an ELISA format, six of the eight LOS immunotypes (L2, L3, L4, L5, L7, and L8) in Groups B and C bound specifically to Maackia amurensis leukoagglutinin (MAL), which recognizes NeuNAcalpha2- 3Galbeta1-4GlcNAc/Glc sequence, but not to Sambucus nigra agglutinin, which binds NeuNAcalpha2-6Gal sequence. The combination of SDS-PAGE and MAL-blot analyses revealed that these six LOSs contained only the NeuNAcalpha2-3Galbeta1-4GlcNAc trisaccharide sequence in their 4.1 kDa LOS components, which have a common terminal lacto-N-neotetraose (LNnT, Galbeta1-4GlcNAcbeta1-3Galbeta1-4Glc) structure when nonsialylated as shown by previous studies. The LOS-lectin binding was abolished when the LOSs were treated with Newcastle disease viral neuraminidase which cleaves alpha2-->3 linked sialic acid. Methylation analysis of a representative LOS (L2) confirmed that NeuNAc is 2-->3 linked to Gal. Thus, these LOSs structurally mimic certain glycolipids, i.e., paragloboside (LNnT-ceramide) and sialylparagloboside and some glycoproteins in having LNnT and N-acetyllactosamine sequences, respectively, with or without alpha2-->3 linked NeuNAc. The molecular mimicry of the LOSs may play a role in the pathogenesis of N.meningitidis by assisting the organism to evade host immune defenses in man.      

Glucose Metabolism Disorder Is Associated with Pulmonary Tuberculosis in Individuals with Respiratory Symptoms from Brazil

BackgroundDiabetes mellitus (DM) has been associated with increased risk for pulmonary tuberculosis (PTB) in endemic settings but it is unknown whether PTB risk is also increased by pre-DM. Here, we prospectively examined the association between glucose metabolism disorder (GMD) and PTB in patients with respiratory symptoms at a tuberculosis primary care reference center in Brazil.MethodsOral glucose tolerance test was performed and levels of fasting plasma glucose and glycohemoglobin (HbA1c) were measured in a cohort of 892 individuals presenting with respiratory symptoms of more than two weeks duration. Patients were also tested for PTB with sputum cultures. Prevalence of pre-DM and DM (based on HbA1c) was estimated and tested for association with incident PTB. Other TB risk factors including smoking history were analyzed.ResultsThe majority of the study population (63.1%) exhibited GMD based on HbA1c ≥5.7%. Patients with GMD had higher prevalence of PTB compared to normoglycemic patients. Individuals with DM exhibited increased frequency of TB-related symptoms and detection of acid-fast bacilli in sputum smears. Among patients with previous DM diagnosis, sustained hyperglycemia (HbA1c ≥7.0%) was associated with increased TB prevalence. Smoking history alone was not significantly associated with TB in our study population but the combination of smoking and HbA1c ≥7.0% was associated with 6 times higher odds for PTB.ConclusionsSustained hyperglycemia and pre-DM are independently associated with active PTB. This evidence raises the question whether improving glycemic control in diabetic TB patients would reduce the risk of TB transmission and simultaneously reduce the clinical burden of disease. A better understanding of mechanisms underlying these associations, especially those suggesting that pre-DM may be a factor driving susceptibility to TB is warranted.