PRISM and Molecular Biology

Mutations in LRRK2 were first reported as causing Parkinson's disease (PD) in late 2004. Since then, approximately a dozen LRRK2 substitutions have been identified that are believed to be pathogenic mutations. The substitution of adenine for guanine at nucleotide 4541 (4541G>A) in LRRK2 was recently reported. This substitution resulted in the replacement of an arginine at position 1514 with a glutamine (R1514Q). Although this substitution was not found in a large cohort of controls, its pathogenicity could not be verified. We have now genotyped the R1514Q substitution in a sample of 954 PD patients from 429 multiplex PD families. This substitution was identified in 1.8% of the PD patients; however, the majority of the PD sibships segregating this substitution were discordant for this putative mutation. In addition, the R1514Q substitution was detected in 1.4% of neurologically evaluated, control individuals. These data suggest that the R1514Q variant is not a pathogenic LRRK2 mutation. We believe it is imperative that the causative nature of any newly identified genetic variant be determined before it is included in any panel for diagnostic testing.

Objective. To characterise and investigate the functional consequences of a novel TNFRSF1A splice site mutation causing TRAPS in a 16 year old male patient and his mother. Methods. Mutational DNA screening was performed in the patient and his mother. Western blotting was used to analyse protein expression levels of TNFR1. A multiplex bead immunoassay was used to quantify serum levels of range of cytokines, and an ELISA-based transcription factor assay to measure NF-κB transactivation. Serum levels of soluble TNFR1 (sTNFR1) were measured by ELISA and FACS analysis used to measure monocyte TNFR1 cell surface expression. Results. A novel mutation, c.472+1G>A (C158delinsYERSSPEAKPSPHPRG), involving a splice site in intron 4 of TNFRSF1A, was found in both the proband and affected mother leading to a 45 nucleotide insertion of intronic DNA into the mRNA, resulting in an in-frame insertion of 15 amino acids in the mature TNFR1 protein and a deletion of a cysteine residue C129(158) in cysteine rich domain (CRD) 3. The patients had reduced serum sTNFR1 and surface expression levels of TNFR1, with marked increases in both pro-and antiinflammatory cytokine. Their peripheral blood mononuclear cells (PBMC) had increased basal NF-κB activation compared with healthy controls and also had increased p50 nuclear expression following TNF stimulation compared with PBMC from healthy controls, as well as T50M (T79M) and C88R (C117R) TRAPS patients and RA patients. Conclusion. A novel, TRAPS causing, TNFRSF1A splice site mutation is associated with decreased sTNFR1 levels, cell surface and whole cell extract expression, and increased NF-κB transcription factor activation.

The MMR (DNA mismatch repair) system helps to maintain the integrity of the genome. This involves eliminating base-base mismatches and insertion/deletion loops, which can lead to microsatellite instability, as seen in tumour cells. Hereditary non-polyposis colon cancer is the result of dominant mutations in MMR genes, such as MLH1, MSH2 and MSH6. More recently there have been case reports of biallelic mutations in the MMR genes MLH1, MSH2 and PMS2. These result in a distinct autosomal recessive cancer predisposition syndrome. The syndrome is characterized by childhood haematological malignancies, brain tumours and the presence of café au lait patches. Second primaries occur frequently in this condition, and survival into adulthood is rare.

Until recently, the PMS2 DNA mismatch repair gene has only rarely been implicated as a cancer susceptibility locus. New studies have shown, however, that earlier analyses of this gene have had technical limitations and also that the genetic behavior of mutant PMS2 alleles is unusual, in that, unlike MLH1 or MSH2 mutations, PMS2 mutations show low heterozygote penetrance. As a result, a dominantly inherited cancer predisposition has not been a feature reported in families with PMS2 mutations. Such families have instead been ascertained through childhood-onset cancers in homozygotes or through apparently sporadic colorectal cancer in heterozygotes. We present further information on the phenotype associated with homozygous PMS2 defi ciency in 13 patients from six families of Pakistani origin living in the United Kingdom. This syndrome is characterized by café-au-lait skin pigmentation and a characteristic tumor spectrum, including leukemias, lymphomas, cere bral malignancies (such as supratentorial primitive neuroectodermal tumors, astrocytomas, and glioblastomas), and colorectal neoplasia with an onset in early adult life. We present evidence for a founder effect in fi ve families, all of which carried the same R802 → X mutation (i.e., arginine-802 to stop) in PMS2. This cancer syndrome can be mistaken for neurofibromatosis type 1, with important management implications including the risk of the disorder occurring in siblings and the likelihood of tumor development in affected individuals. [J Natl Cancer Inst 2006;98:358 -61]

BDDrFVIII is a B-domain deleted recombinant factor VIII (rFVIII) product for haemophilia A. Manufacture uniquely includes purification chromatography by synthetic-affinity ligand rather than murine-based monoclonal antibody, as well as an albumin-free cell culture process. BDDrFVIII was studied in 204 patients, including 62 subjects <16 years old, in two studies. A doubleblind, randomized, pharmacokinetic (PK) crossover study, utilizing a central laboratory assay (one-stage (OS)) for both drug potency assignment and plasma FVIII-activity measurements, demonstrated that BDDrFVIII was PK-equivalent to a fulllength rFVIII. Favourable efficacy and safety were observed: during defined routine prophylaxis in a patient population significant for preexisting target joints, nearly half (45.7%) of patients had no bleeding, and a low-annualized bleed rate (ABR) was achieved (median 1.9); 92.5% of haemorrhages (n = 187) required £2 infusions. Three subjects (1.5%, across both studies) developed de novo inhibitors (low-titre, transient), and the primary safety endpoint, based on a prospective Bayesian analysis, demonstrated the absence of neoantigenicity for BDDrFVIII. The PK-equivalence, based on central testing to align test and reference articles, and the novel Bayesian analysis of inhibitor safety in these investigations reflect robust experimental designs with relevance to future studies. This extensive dataset demonstrates the safety and efficacy of BDDrFVIII for haemophilia A.

Fossil foraminiferal faunas were studied in four, short, late Holocene cores (two localities) from the low tidal, estuarine fringes of the Waitemata Harbour, which is surrounded by New Zealand's largest city, Auckland. All cores record similar major changes in their fossil content since the arrival of humans (ca. 1300 AD), with faunal changes continuing through to the 1970s. Molluscs have disappeared from all cores, and the foraminiferal faunas have switched from dominantly calcareous (Ammonia association) to dominantly agglutinated (Textularia -Schlerochorella, Miliammina -Haplophragmoides associations). A twostep change is evident, with an intermediate mixed calcareous-agglutinated faunal zone (1950s -1970s), characterised by peak abundances (13 -20%) of Elphidium gunteri and Elphidium excavatum s.l.

The use of next-generation sequencing technologies to produce genomic copy number data has recently been described. Most approaches, however, reply on optimal starting DNA, and are therefore unsuitable for the analysis of formalinfixed paraffin-embedded (FFPE) samples, which largely precludes the analysis of many tumour series. We have sought to challenge the limits of this technique with regards to quality and quantity of starting material and the depth of sequencing required. We confirm that the technique can be used to interrogate DNA from cell lines, fresh frozen material and FFPE samples to assess copy number variation. We show that as little as 5 ng of DNA is needed to generate a copy number karyogram, and follow this up with data from a series of FFPE biopsies and surgical samples. We have used various levels of sample multiplexing to demonstrate the adjustable resolution of the methodology, depending on the number of samples and available resources. We also demonstrate reproducibility by use of replicate samples and comparison with microarray-based comparative genomic hybridization (aCGH) and digital PCR. This technique can be valuable in both the analysis of routine diagnostic samples and in examining large repositories of fixed archival material.

During the Late Pliocene-Middle Pleistocene, 56 species and 15 genera of elongate, cylindrical benthic foraminifera disappeared from the deep sea in the South China Sea (ODP Sites 1143 and 1146) as part of the last global extinction in the deep sea. This extinction group (Ext. Gp) exhibited a pulsed decline in abundance and species richness mostly during glacials, and often associated with periods of expansion of polar ice that resulted from increased cooling of the Earth's climate since $2.5 Ma, particularly during the Mid-Pleistocene Climate Transition (MPT, 1.2-0.6 Ma). We infer that the Ext. Gp decline in abundance and disappearance was a result of the increased glacial cooling and consequent increased ventilation of the deep-sea water masses. The detailed record of withdrawal of the Ext. Gp differs between the two sites, with far more disappearances occurring prior to the MPT in the deeper Site 1143 (2772 m) than in the shallower Site 1146 (2092 m). The Late Pliocene and Early Pleistocene declines in deeper parts of the South China Sea (Site 1143) may have resulted from enhanced glacial production of deep, southern-sourced water passing over the sill into the basin from the North-west Pacific. During the MPT however, the Ext. Gp declines and disappearances were of similar timing and magnitude in both sites, implying that both were influenced by the same deep-water mass during glacials at this time. This could have been North Pacific Deep Water, which many workers infer was formed in the northern Pacific during the last glacial, and may have begun forming during MPT glacials, in association with the progressively enhanced cooling of the Northern Hemisphere. r

Sediment core and trench data from a coastal lagoon on the West Coast of the South Island, New Zealand are used to investigate evidence for co-seismic subsidence and associated tsunami inundation. Physical data are used to document a salt marsh soil buried ∼ 80 cm below the modern sediment surface that is locally covered by a gravelly sand bed. The sediment record also contains geochemical and biological (diatom and foram) evidence for abrupt changes in salinity of lagoon waters that link to subsidence, tsunami flooding and to the open versus closed state of the lagoon tidal entrance. At the local scale, these relationships allow for separation of tsunami evidence from other agents of environmental change in the lagoon. We also propose a conceptual connection between these local changes and regional drivers of landscape development, most notably major earthquakes and resultant pulses in sediment supply to the coast.

EMBO THE EMBO JOURNAL THE EMBO JOURNAL SAMHD1 restricts lentiviruses in vivo J Rehwinkel et al 2455 & 2013 European Molecular Biology Organization The EMBO Journal VOL 32 | NO 18 | 2013 SAMHD1 restricts lentiviruses in vivo J Rehwinkel et al SAMHD1 restricts lentiviruses in vivo J Rehwinkel et al

Background: Differential methylation of the two alleles is a hallmark of imprinted genes. Correspondingly, loss of DNA methyltransferase function results in aberrant imprinting and abnormal post-fertilization development. In the mouse, mutations of the oocyte-specific isoform of the DNA methyltransferase Dnmt1 (Dnmt1o) and of the methyltransferase-like Dnmt3L gene result in specific failures of imprint establishment or maintenance, at multiple loci. We have previously shown in humans that an analogous inherited failure to establish imprinting at multiple loci in the female germline underlies a rare phenotype of recurrent hydatidiform mole.

Random Forest is a prediction technique based on growing trees on bootstrap samples of data, in conjunction with a random selection of explanatory variables to define the best split at each node. In the case of a quantitative outcome, the tree predictor takes on a numerical value. We applied Random Forest to the first replicate of the Genetic Analysis Workshop 13 simulated data set, with the sibling pairs as our units of analysis and identity by descent (IBD) at selected loci as our explanatory variables. With the knowledge of the true model, we performed two sets of analyses on three phenotypes: HDL, triglycerides, and glucose. The goal was to approach the mapping of complex traits from a multivariate perspective. The first set of analyses mimics a candidate gene approach with a high proportion of true genes among the predictors while the second set represents a genome scan analysis using microsatellite markers. Random Forest was able to identify a few of the major genes influencing the phenotypes, such as baseline HDL and triglycerides, but failed to identify the major genes regulating baseline glucose levels.

The Asian Economic Crisis presents perhaps a final opportunity for Australia's IT&T industry to make its presence felt on the world stage. As a centre of innovation and a desirable location for IT&T facilities, Australia has often been overlooked as its near neighbours to the north gain all the attention. This paper summarises the main obstacles Australia has to overcome in order to gain mind-share amongst the global IT&T supplier community. ᭧ Journal of Strategic Information Systems 8 (1999) 21-27 STRINF 1101 0963-8687/99/$ -see front matter ᭧

Background: LTC4 synthase is essential for the production of cysteinyl leukotrienes (Cys-LT), critical mediators in asthma. We have identified a novel promoter polymorphism at position -1072 (G/A) and a -444 (A/C) polymorphism has previously been reported. The role of these polymorphisms in the genetic susceptibility to asthma was examined. Methods: To test for genetic association with asthma phenotypes, 341 white families (two asthmatic siblings) and 184 non-asthmatic control subjects were genotyped. Genetic association was assessed using case control and transmission disequilibrium test (TDT) analyses. LTC4S promoter luciferase constructs and transiently transfected human HeLa and KU812F cells were generated to determine the functional role of these polymorphisms on basal transcription. Results: No associations were observed in case control analyses (-1072 A, q=0.09; -444 C, q=0.29); the TDT identified a borderline association between the -444 C allele and bronchial responsiveness to methacholine (p=0.065). Asthmatic children with the -444 C allele had a lower mean basal forced expiratory volume in 1 second (97.4 v 92.7% predicted, p=0.005). LTC4S promoter luciferase analyses provided no evidence for a functional role of either polymorphism in determining basal transcription. Conclusion: This study does not support a role for these polymorphisms in genetic susceptibility to asthma but provides evidence to suggest a role in determining lung function parameters.

1. 1. The green marine alga Enteromorpha intestinalis evolves dimethyl sulfide on removal from the water. The precursor of the sulfide is dimethyl-β-propiothetin which has been isolated from the alga as the platinichloride, the picrate and the styphnate. 2. 2. The presence of a ...