Crystal structure of a novel mid-gut procarboxypeptidase from the cotton pest Helicoverpa armigera.

The cotton bollworm Helicoverpa armigera (Hubner) (Lepidoptera: Noctuidae) is one of the most serious insect pests in Australia, India and China. The larva causes substantial economical losses to legume, fibre, cereal oilseed and vegetable crops. This pest has proven to be difficult to control by conventional means, mainly due to the development of pesticide resistance. We present here the 2.5 A crystal structure from the novel procarboxypeptidase (PCPAHa) found in the gut extracts from H. armigera larvae, the first one reported for an insect. This metalloprotease is synthesized as a zymogen of 46.6 kDa which, upon in vitro activation with Lys-C endoproteinase, yields a pro-segment of 91 residues and an active carboxypeptidase moiety of 318 residues. Both regions show a three-dimensional structure quite similar to the corresponding structures in mammalian digestive carboxypeptidases, the most relevant structural differences being located in the loops between conserved secondary structure elements, including the primary activation site. This activation site contains the motif (Ala)(5)Lys at the C terminus of the helix connecting the pro- and the carboxypeptidase domains. A remarkable feature of PCPAHa is the occurrence of the same (Ala)(6)Lys near the C terminus of the active enzyme. The presence of Ser255 in PCPAHa instead of Ile and Asp found in the pancreatic A and B forms, respectively, enlarges the S1' specificity pocket and influences the substrate preferences of the enzyme. The C-terminal tail of the leech carboxypeptidase inhibitor has been modelled into the PCPAHa active site to explore the substrate preferences and the enzymatic mechanism of this enzyme.     

Molecular phylogeny of the fungus gnat subfamilies Gnoristinae and Mycomyinae,and their position within Mycetophilidae (Diptera)

The phylogeny of the fungus gnat family Mycetophilidae (Diptera) is reconstructed with a focus on the species‐rich and taxonomically difficult subfamilies Gnoristinae and Mycomyinae. The multigene phylogenetic analyses are based on five nuclear (18S, 28S, CAD, MCS, ITS2) and four mitochondrial (12S, 16S, COI, CytB) gene markers. The analyses strongly support the monophyly of Mycetophilidae and the subfamilies Manotinae, Sciophilinae, Leiinae, and Mycomyinae, although Gnoristinae is paraphyletic with respect to Mycetophilinae. All the genera and groups of genera included are supported as monophyletic, except for Acomoptera Vockeroth, Boletina Staeger, Dziedzickia Johannsen, Ectrepesthoneura Enderlein, and Neoempheria Osten Sacken. Ancestral character state reconstructions were applied to two morphological features present in Gnoristinae and Mycomyinae (i.e. presence of setae on wing membrane and wing vein R₄) in order to assess their evolution. The wing vein R₄ appears as an unstable character, spread throughout different clades. A dated phylogeny of the family Mycetophilidae showed that most of the subfamilies of Mycetophilidae originated and diversified during the Cretaceous. The youngest subfamilies, originated in the Paleogene, appear to be Mycomyinae and Mycetophilinae.     

Identifying common prognostic factors in genomic cancer studies: A novel index for censored outcomes

Background  

With the growing number of public repositories for high-throughput genomic data, it is of great interest to combine the results produced by independent research groups. Such a combination allows the identification of common genomic factors across multiple cancer types and provides new insights into the disease process. In the framework of the proportional hazards model, classical procedures, which consist of ranking genes according to the estimated hazard ratio or the p-value obtained from a test statistic of no association between survival and gene expression level, are not suitable for gene selection across multiple genomic datasets with different sample sizes. We propose a novel index for identifying genes with a common effect across heterogeneous genomic studies designed to remain stable whatever the sample size and which has a straightforward interpretation in terms of the percentage of separability between patients according to their survival times and gene expression measurements.     

S100 alpha, CAPL, and CACY: molecular cloning and expression analysis of three calcium-binding proteins from human heart.

Elevated levels of intracellular calcium are a major cause of myocardial dysfunction. To find possible mediators of the deregulated calcium we searched for EF-hand calcium-binding proteins of the S100 family. By PCR technology we identified three members of the S100 protein family (S100 alpha, CACY, and CAPL) in the human heart. We cloned the corresponding cDNAs and examined their expression levels in various human tissues by Northern blot analysis. All three proteins are expressed at high levels in the human heart. Whereas CACY and CAPL mRNAs are expressed ubiquitously, S100 alpha mRNA is restricted to heart, skeletal muscle, and brain. Interestingly, the expression pattern of S100 alpha, CACY, and CAPL in human tissues differs significantly from that in rodent tissues.     

Erythrocytes and age. Disintegration of erythrocytes by brilliant cresyl blue in correlation to age

The author describes changes in the disintegration of erythrocytes by brilliant cresyl blue in correlation to age, in rats aged 21, 42, 90-105, 340-360 and 690-720 days. The erythrocytes were incubated for 4 hours in an isotonic NaCl solution, in Krebs-Ringer solution and in each of these solutions plus brilliant cresyl blue. Disintegration in plain NaCl solution was found to be the greatest in the case of erythrocytes from 690- to 720-day-old rats. In the same solution plus brilliant cresyl blue, the rate of disintegration was very high in 21-day-old, 42-day-old and 690- to 720-day-old animals; at 90-105 days it was lower and at 340-360 days it was the lowest. Disintegration of erythrocytes in plain Krebs-Ringer solution was the lowest at 21 and 42 days; in the other age groups it was slightly higher. On adding brilliant cresyl blue, the rate of disintegration rose significantly in 21-, 42- and 690- to 720-day-old animals; at 90-105 days and 340-360 days it was no different from disintegration in plain Krebs-Ringer solution. It can be seen from the results that the rate of brilliant cresyl blue-induced erythrocyte disintegration is dependent on the age of the animals from which the erythrocytes are taken.     

Thiostrepton resistance mutations in the gene for 23S ribosomal RNA of halobacteria

Mutants of Halobacterium (H.) halobium and H. cutirubrum were isolated which are resistant to the 70S ribosome inhibitor thiostrepton. Using primer extension analysis, resistance was shown to correlate with base changes at position 1159, which corresponds to position 1067 of the E. coli 23S rRNA. In four mutants, A1159 was replaced by U, in one mutant by G. The results show that not only methylation (Cundliffe & Thompson (1979) Nature 278, 859-861) of A1067 (E. coli nomenclature), but also base changes at this position cause high-level resistance to thiostrepton.