Molecular diversity of the HLA-C gene identified in a Caucasian population

A DNA typing procedure, based on a two stage polymerase chain reaction-sequence-specific oligonucleotide probe (PCR-SSOP) typing strategy, has been developed and applied to DNA from 1000 healthy individuals from the Northern Ireland region. The two-stage procedure involves human leukocyte antigen (HLA-C) identification through the use of a medium resolution PCR-SSOP system, followed by four secondary group specific PCR-SSOP systems, to enable allele resolution. The PCR-SSOP systems were designed for the identification of HLA-Cw alleles with possible discrimination within exons 2 and 3 of the HLA-C gene, i.e., HLA-Cw*01-Cw*16. PCR-SSP tests were designed for the resolution of HLA-Cw*17 and -Cw*18 alleles. The systems can also be used independently of each other if selective allele resolution is required. HLA-Cw allele frequencies occuring within the Northern Ireland population have been compiled, along with estimations of HLA-B/Cw haplotype frequencies. (C) American Society for Histocompatibility and Immunogenetics, 2002. Published by Elsevier Science Inc.

Four independent gene phylogenies confirm the ancient relationships of Madagascar endemic species in the Papilio demoleus group (Lepidoptera, Papilionidae).

Madagascar is home to numerous endemic species and lineages, but the processes that have contributed to its endangered diversity are still poorly understood. Evidence is accumulating to demonstrate the importance of Tertiary dispersal across varying distances of oceanic barriers, supplementing vicariance relationships dating back to the Cretaceous, but these hypotheses remain tentative in the absence of well-supported phylogenies. In the Papilio demoleus group of swallowtail butterflies, three of the five recognized species are restricted to Madagascar, whereas the remaining two species range across the Afrotropical zone and southern Asia plus Australia. We reconstructed phylogenetic relationships for all species in the P. demoleus group, as well as 11 outgroup Papilio species, using 60 morphological characters and about 4 kb of nucleotide sequences from two mitochondrial (cytochrome oxidase I and II) and two nuclear (wg and EF-1a) genes. Of the three endemic Malagasy species, the two that are formally listed as endangered or at risk represented the most basal divergences in the group, while the more common third endemic was clearly related to African P. demodocus. The fifth species, P. demoleus, showed little differentiation across southern Asia, but showed divergence from its subspecies sthenelus in Australia. Dispersal-vicariance analysis using cladograms derived from morphology and three independent genes indicated a Malagasy diversification of lime swallowtails in the middle Miocene. Thus, diversification processes on the island of Madagascar may have contributed to the origin of common butterflies that now occur throughout much of the Old World tropical and subtemperate regions. An alternative hypothesis, that Madagascar is a refuge for ancient lineages resulting from successive colonizations from Africa, is less parsimonious and does not explain the relatively low continental diversity of the group.

The trace amine associated receptor (TAAR6) gene is not associated with schizophrenia in the Irish Case-Control Study of Schizophrenia (ICCSS) sample

To replicate previous association between TAAR6 and schizophrenia, including our own finding in the Irish Study of High Density Schizophrenia Families (ISHDSF) sample, we genotyped 12 single nucleotide polymorphisms (SNPs) in the Irish Case-Control Study of Schizophrenia (ICCSS) sample. Only rs9389020 provided nominal evidence for association (p

Clinical Utility of Microarray-Based Gene Expression Profiling in the Diagnosis and Subclassification of Leukemia: Report From the International Microarray Innovations in Leukemia Study Group

The Microarray Innovations in Leukemia study assessed the clinical utility of gene expression profiling as a single test to subtype leukemias into conventional categories of myeloid and lymphoid malignancies. METHODS: The investigation was performed in 11 laboratories across three continents and included 3,334 patients. An exploratory retrospective stage I study was designed for biomarker discovery and generated whole-genome expression profiles from 2,143 patients with leukemias and myelodysplastic syndromes. The gene expression profiling-based diagnostic accuracy was further validated in a prospective second study stage of an independent cohort of 1,191 patients. RESULTS: On the basis of 2,096 samples, the stage I study achieved 92.2% classification accuracy for all 18 distinct classes investigated (median specificity of 99.7%). In a second cohort of 1,152 prospectively collected patients, a classification scheme reached 95.6% median sensitivity and 99.8% median specificity for 14 standard subtypes of acute leukemia (eight acute lymphoblastic leukemia and six acute myeloid leukemia classes, n = 693). In 29 (57%) of 51 discrepant cases, the microarray results had outperformed routine diagnostic methods. CONCLUSION: Gene expression profiling is a robust technology for the diagnosis of hematologic malignancies with high accuracy. It may complement current diagnostic algorithms and could offer a reliable platform for patients who lack access to today's state-of-the-art diagnostic work-up. Our comprehensive gene expression data set will be submitted to the public domain to foster research focusing on the molecular understanding of leukemias

An international standardization programme towards the application of gene expression profiling in routine leukaemia diagnostics: the Microarray Innovations in LEukemia study prephase

Gene expression profiling has the potential to enhance current methods for the diagnosis of haematological malignancies. Here, we present data on 204 analyses from an international standardization programme that was conducted in 11 laboratories as a prephase to the Microarray Innovations in LEukemia (MILE) study. Each laboratory prepared two cell line samples, together with three replicate leukaemia patient lysates in two distinct stages: (i) a 5-d course of protocol training, and (ii) independent proficiency testing. Unsupervised, supervised, and r(2) correlation analyses demonstrated that microarray analysis can be performed with remarkably high intra-laboratory reproducibility and with comparable quality and reliability.

Association of polymorphisms in the hepatocyte growth factor gene promoter with keratoconus

Purpose. Keratoconus is a progressive disorder of the cornea that can lead to severe visual impairment or blindness. Although several genomic regions have been linked to rare familial forms of keratoconus, no genes have yet been definitively identified for common forms of the disease. Methods. Two genome-wide association scans were undertaken in parallel. The first used pooled DNA from an Australian cohort, followed by typing of top-ranked single-nucleotide polymorphisms (SNPs) in individual DNA samples. The second was conducted in individually genotyped patients, and controls from the USA. Tag SNPs around the hepatocyte growth factor (HGF) gene were typed in three additional replication cohorts. Serum levels of HGF protein in normal individuals were assessed with ELISA and correlated with genotype. Results. The only SNP observed to be associated in both the pooled discovery and primary replication cohort was rs1014091, located upstream of the HGF gene. The nearby SNP rs3735520 was found to be associated in the individually typed discovery cohort (P = 6.1 × 10 ). Genotyping of tag SNPs around HGF revealed association at rs3735520 and rs17501108/rs1014091 in four of the five cohorts. Meta-analysis of all five datasets together yielded suggestive P values for rs3735520 (P = 9.9 × 10 ) and rs17501108 (P = 9.9 × 10 ). In addition, SNP rs3735520 was found to be associated with serum HGF level in normal individuals (P = 0.036). Conclusions. Taken together, these results implicate genetic variation at the HGF locus with keratoconus susceptibility. © 2011 The Association for Research in Vision and Ophthalmology, Inc.

Schistosome I/Lamides - A new family of bioactive helminth neuropeptides

Here we report the identification of a new family of helminth neuropeptides with members in both nematodes and flatworms, and include preliminary cell biological and functional characterisation of one of the peptides from the trematode parasite of humans, Schistosoma mansoni. Bioinformatics and Rapid Amplification of cDNA Ends (RACE)-PCR were used to identify the completes. mansoni neuropeptide precursor gene Sm-npp-1, which encodes three pentapeptides bearing the motif (A/G)FVR(I/L).NH2. Similar peptides were identified in three other flatworm species and in 15 nematode species. Quantitative PCR (qPCR) and immunocytochemical (ICC) analyses showed that Sm-npp-1 is constitutively expressed in larval and adult worms. ICC and confocal microscopy were employed to localise one of the schistosome NPP-1 peptides (GFVRIamide) in adult worms and schistosomules; antibodies labelled a pair of neurones in the cerebral ganglia that extend posteriorly along the main nerve cords. GFVRIamide displayed no detectable co-localisation with FMRFamide-like peptides (FLPs), nor was it detectable in muscle innervation. Exogenously applied peptide had a significant inhibitory effect on the mobility of whole adult worm pairs at 10(-5) M (n = 9). Finally, we explored Sm-npp-1 function in schistosomules using RNA interference (RNAi); we successfully achieved specific knockdown of the Sm-npp-1 transcript (54.46 +/- 10.41% knockdown, n = 3), but did not detect any clear, aberrant mobility or morphological phenotypes. NPP-1-like peptides are a new family of helminth peptides with a cell-specific expression pattern distinct from FLPs and a modulatory effect on schistosome muscular activity. (C) 2011 Australian Society for Parasitology Inc. Published by Elsevier Ltd. All rights reserved.

Loss-of-function mutations in the cathepsin C gene result in periodontal disease and palmoplantar keratosis

Papillon-Lefevre syndrome, or keratosis palmoplantaris with periodontopathia (PLS, MIM 245000), is an autosomal recessive disorder that is mainly ascertained by dentists because of the severe periodontitis that afflicts patients(1,2). Both the deciduous and permanent dentitions are affected, resulting in premature tooth loss. Palmoplantar keratosis, varying from mild psoriasiform scaly skin to overt hyperkeratosis, typically develops within the first three years of life. Keratosis also affects other sites such as elbows and knees. Most PLS patients display both periodontitis and hyperkeratosis. some patients have only palmoplantar keratosis or periodontitis, and in rare individuals the periodontitis is mild and of late onset(3-6). The PLS locus has been mapped to chromosome 11q14-q21 (refs 7-9). Using homozygosity mapping in eight small consanguineous families, we have narrowed the candidate region to a 1.2-cM interval between D11S4082 and D11S931. The gene (CTSC) encoding the lysosomal protease cathepsin C (or dipeptidyl aminopeptidase I) lies within this interval. We defined the genomic structure of CTSC and found mutations in all eight families. In two of these families we used a functional assay to demonstrate an almost total loss of cathepsin C activity in PLS patients and reduced activity in obligate carriers.

Characterization of IS2112, a new insertion sequence from <i>Rhodococcusi>, and its relationship with mobile elements belonging to the IS110 family

A new insertion sequence (IS2112) was identified in the genome of the 1-haloalkane-utilizing bacterium Rhodococcus rhodochrous NCIMB 13064. The insertion element is 1415 bp long, does not contain terminal inverted repeats, and is not flanked by directly repeated sequences. IS2112 belongs to the IS110 family of transposable elements, and forms a separate subfamily, along with IS116, Two copies of IS2112 were found in R, rhodochrous NCIMB 13064 and one, two or three copies of a similar sequence were detected in five other 1-haloalkane-degrading Rhodococcus strains. There were no sequences homologous to IS2112 found in the l-haloalkane-degrading 'Pseudomonas pavonaceae' 170 and Rhodococcus sp, HA1 or in several Rhodococcus strains which do not utilize haloalkanes, IS2112 was originally found in plasmid pRTL1 of R. rhodochrous NCIMB 13064 which harbours genes encoding utilization of l-haloalkanes, and was located 5 kbp upstream of the haloalkane dehalogenase gene (dhaA), Although the second copy of IS2112 in strain NCIMB 13064 was also present on the pRTL1 plasmid, these sequences do not apparently comprise a single composite transposon encoding haloalkane utilization. An analysis of derivatives of NCIMB 13064 revealed that IS2112 was involved in genome rearrangements. IS2112 appeared to change its location as a result of transposition and as a result of other rearrangements of the NCIMB 13064 genome.

Degradation of 1,3-dichloropropene by <i>Pseudomonas cichorii i>170

The gram-negative bacterium Pseudomonas cichorii 170, isolated from soil that was repeatedly treated with the nematocide 1,3-dichloropropene, could utilize low concentrations of 1,3-dichloropropene as a sole carbon and energy source, Strain 170 was also able to grow on 3-chloroallyl alcohol, 3-chloroacrylic acid, and several 1-halo-n-alkanes. This organism produced at least three different dehalogenases: a hydrolytic haloalkane dehalogenase specific for haloalkanes and two 3-chloroacrylic acid dehalogenases, one specific for cis-3-chloroacrylic acid and the other specific for trans-3-chloroacrylic acid. The haloalkane dehalogenase and the trans-3-chloroacrylic acid dehalogenase were expressed constitutively, whereas the cis-3-chloroacrylic acid dehalogenase was inducible, The presence of these enzymes indicates that 1,3-dichloropropene is hydrolyzed to 3-chloroallyl alcohol, which is oxidized in two steps to 3-chloroacrylic acid. The latter compound is then dehalogenated, probably forming malonic acid semialdehyde. The haloalkane dehalogenase gene, which is involved in the conversion of 1,3-dichloropropene to 3-chloroallyl alcohol, was cloned and sequenced, and this gene turned out to be identical to the previously studied dhaA gene of the gram-positive bacterium Rhodococcus rhodochrous NCIMB13063, Mutants resistant to the suicide substrate 1,2-dibromoethane lacked haloalkane dehalogenase activity and therefore could not utilize haloalkanes for growth. PCR analysis showed that these mutants had lost at least part of the dhaA gene.

Cloning of new <i>Rhodococcus i>extradiol dioxygenase genes and study of their distribution in different Rhodococcus strains

Four extradiol dioxygenase genes which encode enzymes active against catechol and substituted catechols were cloned from two different Rhodococcus strains, and their nucleotide sequences were determined. A catechol 2,3-dioxygenase gene (edoC) was shown to be identical to the previously described ipbC gene from the isopropylbenzene operon of Rhodococcus erythropolis. Amino acid sequences deduced from the three other genes (edoA, edoB and edoD) were shown to have various degrees of homology to different extradiol dioxygenases, The EdoA and EdoB dioxygenases were classified as belonging to the third family of type I oxygenases and represented two new subfamilies, whereas the EdoD dioxygenase was a type II enzyme. Analysis of six Rhodococcus strains revealed a wide distribution of the above dioxygenase genes. Rhodococcus sp. I1 was shown to harbour all four of the analysed dioxygenase genes. Nucleotide sequences homologous to the edoB gene were present in all of the strains, including R. erythropolis NCIMB 13065, which did not utilize any of the aromatic compounds analysed. The latter finding points to the existence of a silent pathway(s) for degradation of aromatic compounds in this Rhodococcus strain.