Interplay between RNA interference and heat shock response systems in Drosophila melanogaster.

The genome expression pattern is strongly modified during the heat shock response (HSR) to form an adaptive state. This may be partly achieved by modulating microRNA levels that control the expression of a great number of genes that are embedded within the gene circuitry. Here, we investigated the cross-talk between two highly conserved and universal house-keeping systems, the HSR and microRNA machinery, in Drosophila melanogaster We demonstrated that pronounced interstrain differences in the microRNA levels are alleviated after heat shock (HS) to form a uniform microRNA pattern. However, individual strains exhibit different patterns of microRNA expression during the course of recovery. Importantly, HS-regulated microRNAs may target functionally similar HS-responsive genes involved in the HSR. Despite the observed general downregulation of primary microRNA precursor expression as well as core microRNA pathway genes after HS, the levels of many mature microRNAs are upregulated. This indicates that the regulation of miRNA expression after HS occurs at transcriptional and post-transcriptional levels. It was also shown that deletion of all hsp70 genes had no significant effect on microRNA biogenesis but might influence the dynamics of microRNA expression during the HSR.

Disruption of SF3B1 results in deregulated expression and splicing of key genes and pathways in myelodysplastic syndrome hematopoietic stem and progenitor cells.

The splicing factor SF3B1 is the most commonly mutated gene in the myelodysplastic syndrome (MDS), particularly in patients with refractory anemia with ring sideroblasts (RARS). We investigated the functional effects of SF3B1 disruption in myeloid cell lines: SF3B1 knockdown resulted in growth inhibition, cell cycle arrest and impaired erythroid differentiation and deregulation of many genes and pathways, including cell cycle regulation and RNA processing. MDS is a disorder of the hematopoietic stem cell and we thus studied the transcriptome of CD34(+) cells from MDS patients with SF3B1 mutations using RNA sequencing. Genes significantly differentially expressed at the transcript and/or exon level in SF3B1 mutant compared with wild-type cases include genes that are involved in MDS pathogenesis (ASXL1 and CBL), iron homeostasis and mitochondrial metabolism (ALAS2, ABCB7 and SLC25A37) and RNA splicing/processing (PRPF8 and HNRNPD). Many genes regulated by a DNA damage-induced BRCA1-BCLAF1-SF3B1 protein complex showed differential expression/splicing in SF3B1 mutant cases. This is the first study to determine the target genes of SF3B1 mutation in MDS CD34(+) cells. Our data indicate that SF3B1 has a critical role in MDS by affecting the expression and splicing of genes involved in specific cellular processes/pathways, many of which are relevant to the known RARS pathophysiology, suggesting a causal link.

[InterPro as a new tool for whole genome analysis. A comparative analysis of Mycobacterium tuberculosis, Bacillus subtilis and Escherichia coli as a case study].

InterPro was developed as a new integrated documentation resource for protein families, domains and functional sites to rationalize the complementary efforts of the PROSITE, PRINTS, Pfam and ProDom database projects and has applications in computational functional classification of newly determined sequences lacking biochemical characterization and in comparative genome analysis. InterPro contains over 3500 entries, with more than 1000000 hits in SWISS-PROT and TrEMBL. The database is accessible for text- and sequence-based searches at http://www.ebi.ac.uk/interpro/. InterPro was used for whole proteome analysis of the pathogenic microorganism, Mycobacterium tuberculosis, and comparison with the predicted protein coding sequences of the complete genomes of Bacillus subtilis and Escherichia coli. 64.8% of the M. tuberculosis proteins in the proteome matched InterPro entries, and these could be classified according to function. The comparison with B. subtilis and E. coli provided information on the most common protein families and domains, and the most highly represented families in each organism. InterPro thus provides a useful tool for global views of whole proteomes and their compositions.

Analysis of the mouse transcriptome based on functional annotation of 60,770 full-length cDNAs.

Only a small proportion of the mouse genome is transcribed into mature messenger RNA transcripts. There is an international collaborative effort to identify all full-length mRNA transcripts from the mouse, and to ensure that each is represented in a physical collection of clones. Here we report the manual annotation of 60,770 full-length mouse complementary DNA sequences. These are clustered into 33,409 'transcriptional units', contributing 90.1% of a newly established mouse transcriptome database. Of these transcriptional units, 4,258 are new protein-coding and 11,665 are new non-coding messages, indicating that non-coding RNA is a major component of the transcriptome. 41% of all transcriptional units showed evidence of alternative splicing. In protein-coding transcripts, 79% of splice variations altered the protein product. Whole-transcriptome analyses resulted in the identification of 2,431 sense-antisense pairs. The present work, completely supported by physical clones, provides the most comprehensive survey of a mammalian transcriptome so far, and is a valuable resource for functional genomics.

Interactive InterPro-based comparisons of proteins in whole genomes.

MOTIVATION: The SWISS-PROT group at the EBI has developed the Proteome Analysis Database utilizing existing resources and providing comprehensive and integrated comparative analysis of the predicted protein coding sequences of the complete genomes of bacteria, archaea and eukaryotes. The Proteome Analysis Database is accompanied by a program that has been designed to carry out interactive InterPro proteome comparisons for any one proteome against any other one or more of the proteomes in the database.

Profiling the malaria genome: a gene survey of three species of malaria parasite with comparison to other apicomplexan species.

We have undertaken the first comparative pilot gene discovery analysis of approximately 25,000 random genomic and expressed sequence tags (ESTs) from three species of Plasmodium, the infectious agent that causes malaria. A total of 5482 genome survey sequences (GSSs) and 5582 ESTs were generated from mung bean nuclease (MBN) and cDNA libraries, respectively, of the ANKA line of the rodent malaria parasite Plasmodium berghei, and 10,874 GSSs generated from MBN libraries of the Salvador I and Belem lines of Plasmodium vivax, the most geographically wide-spread human malaria pathogen. These tags, together with 2438 Plasmodium falciparum sequences present in GenBank, were used to perform first-pass assembly and transcript reconstruction, and non-redundant consensus sequence datasets created. The datasets were compared against public protein databases and more than 1000 putative new Plasmodium proteins identified based on sequence similarity. Homologs of previously characterized Plasmodium genes were also identified, increasing the number of P. vivax and P. berghei sequences in public databases at least 10-fold. Comparative studies with other species of Apicomplexa identified interesting homologs of possible therapeutic or diagnostic value. A gene prediction program, Phat, was used to predict probable open reading frames for proteins in all three datasets. Predicted and non-redundant BLAST-matched proteins were submitted to InterPro, an integrated database of protein domains, signatures and families, for functional classification. Thus a partial predicted proteome was created for each species. This first comparative analysis of Plasmodium protein coding sequences represents a valuable resource for further studies on the biology of this important pathogen.

Application of InterPro for the functional classification of the proteins of fish origin in SWISS-PROT and TrEMBL.

InterPro (http://www.ebi.ac.uk/interpro/) is an integrated documentation resource for protein families, domains and sites, developed initially as a means of rationalizing the complementary efforts of the PROSITE, PRINTS, Pfam and ProDom database projects. It is a useful resource that aids the functional classification of proteins. Almost 90% of the actinopterygii protein sequences from SWISS-PROT and TrEMBL can be classified using InterPro. Over 30% of the actinopterygii protein sequences currently in SWISS-PROT and TrEMBL are of mitochondrial origin, the majority of which belong to the cytochrome b/b6 family. InterPro also gives insights into the domain composition of the classified proteins and has applications in the functional classification of newly determined sequences lacking biochemical characterization, and in comparative genome analysis. A comparison of the actinopterygii protein sequences against the sequences of other eukaryotes confirms the high representation of eukaryotic protein kinase in the organisms studied. The comparisons also show that, based on InterPro families, the trans-species evolution of MHC class I and II molecules in mammals and teleost fish can be recognized.

The InterPro database, an integrated documentation resource for protein families, domains and functional sites.

Signature databases are vital tools for identifying distant relationships in novel sequences and hence for inferring protein function. InterPro is an integrated documentation resource for protein families, domains and functional sites, which amalgamates the efforts of the PROSITE, PRINTS, Pfam and ProDom database projects. Each InterPro entry includes a functional description, annotation, literature references and links back to the relevant member database(s). Release 2.0 of InterPro (October 2000) contains over 3000 entries, representing families, domains, repeats and sites of post-translational modification encoded by a total of 6804 different regular expressions, profiles, fingerprints and Hidden Markov Models. Each InterPro entry lists all the matches against SWISS-PROT and TrEMBL (more than 1,000,000 hits from 462,500 proteins in SWISS-PROT and TrEMBL). The database is accessible for text- and sequence-based searches at http://www.ebi.ac.uk/interpro/. Questions can be emailed to interhelp@ebi.ac.uk.

Proteome Analysis Database: online application of InterPro and CluSTr for the functional classification of proteins in whole genomes.

The SWISS-PROT group at EBI has developed the Proteome Analysis Database utilising existing resources and providing comparative analysis of the predicted protein coding sequences of the complete genomes of bacteria, archaea and eukaryotes (http://www.ebi.ac. uk/proteome/). The two main projects used, InterPro and CluSTr, give a new perspective on families, domains and sites and cover 31-67% (InterPro statistics) of the proteins from each of the complete genomes. CluSTr covers the three complete eukaryotic genomes and the incomplete human genome data. The Proteome Analysis Database is accompanied by a program that has been designed to carry out InterPro proteome comparisons for any one proteome against any other one or more of the proteomes in the database.

InterPro--an integrated documentation resource for protein families, domains and functional sites.

MOTIVATION: InterPro is a new integrated documentation resource for protein families, domains and functional sites, developed initially as a means of rationalising the complementary efforts of the PROSITE, PRINTS, Pfam and ProDom database projects. RESULTS: Merged annotations from PRINTS, PROSITE and Pfam form the InterPro core. Each combined InterPro entry includes functional descriptions and literature references, and links are made back to the relevant parent database(s), allowing users to see at a glance whether a particular family or domain has associated patterns, profiles, fingerprints, etc. Merged and individual entries (i.e. those that have no counterpart in the companion resources) are assigned unique accession numbers. Release 1.2 of InterPro (June 2000) contains over 3000 entries, representing families, domains, repeats and sites of post-translational modification (PTMs) encoded by 6581 different regular expressions, profiles, fingerprints and Hidden Markov Models (HMMs). Each InterPro entry lists all the matches against SWISS-PROT and TrEMBL (more than 1000000 hits from 264333 different proteins out of 384572 in SWISS-PROT and TrEMBL).

[Determination and comparative analysis of the nucleotide sequence for minireplicon fragments of the cryptic plasmid p1414 from the soil strain of Bacillus subtilis]. / Opredelenie i sravnitel'nyi analiz nukleotidnoi posledovatel'nosti fragmentov minireplikona kripticheskoi plazmidy p1414 iz pochvennogo shtamma Bacillus subtilis.

Nucleotide sequences of two minireplicon fragments of the p1414 cryptic plasmid of Bacillus subtilis were determined. The fragments corresponded to the region containing ori(+) and the gene coding for Rep protein. Comparing sequences of the fragments with corresponding sequences of other ss+ plasmids suggested that ori(+) of p1414 belongs to the family of endogenous cryptic B. subtilis plasmids, which form an individual, closely related subgroup in the group of ori(+) sequences of the pC194 type. It was found that the amino acid sequence of a conservative FLTLTV motif located, together with its flanking sequences, at the N ends of Rep proteins encoded by different ss+ plasmids, is similar to those of several transmembrane proteins and signal peptides. These results, together with computer data on predicting the secondary and tertiary structure of the N-terminal domain of the p1414 Rep protein, suggest that the domain can serve as a "membrane anchor" during plasmid replication.

[Conserved regions of potential ORF1 protein products of mobile elements and retroviral proteins, encoded by the gag gene]. / Konservativnye oblasti potentsial'nykh belkovykh produktov ORF1 mobil'nykh élementov i belkov retrovirusov, kodiruemykh genom gag.

Computer search for homology regions of retroelement ORF1 protein products and retroviral gag-encoded proteins was performed using nucleotide and protein sequences of 113 retrotransposons, conservative domains were found in amino acid sequences encoded by viral gene gag: 22 and 17 amino acid residues in capsid and nucleocapsid proteins, respectively. Similar regions were found in potential protein products of ORF1. Another conservative region was revealed by multiple alignment in amino acid sequences of potential protein products of ORF1 of mobile elements and in p19 matrix protein of some retroviruses; this region is different in mobile elements from different classes, but within each class it shows high similarity. The secondary structure of this region is an alpha-helix.

The pss4 gene from Rhizobium leguminosarum by viciae VF39: cloning, sequence and the possible role in polysaccharide production and nodule formation.

The Tn5 insertion into the genome of Rhizobium leguminosarum bv viciae VF39, resulting in non-mucoid growth and formation of non-N2-fixing nodule-like structures on Vicia faba plants, was mapped within a 1.4-kb EcoRV-SacI fragment. Nucleotide sequence analysis revealed an ORF (pss4) of 263 amino acids (aa). Three transcription start points (tsp) were determined. Two of them were localized upstream from the first GTG codon; the third tsp was mapped in front of the second putative start codon (GTG) corresponding to Val64 of the Pss4 aa sequence. The expression of pss4 in a T7 RNA polymerase/promoter system produced a single approx. 29-kDa protein. Pss4 reveals similarity to several proteins involved in polysaccharide biosynthesis in various Rhizobium species. A nearly complete homology was found with PssA from Rl biovar phaseoli 8002 [Borthakur et al., Mol. Gen. Genet. 213 (1988) 155-162], except that Pss4 has an additional 63 aa on its N terminus.

[Translocation of chromosome 1/29 and its various phenotypic manifestations in Ala-Tau breed cattle]. / Translokatsiia khromosomy 1/29 i ee nekotorye fenotipicheskie proiavleniia u skota ala-tau porody.

117 heads of Ala-Tau breed cattle were studied cytogenetically. The 1/29 translocation was present in 13 animals of unusual for this breed light-grey colour. Chimerism 2n = 59/2n = 60 was found in twins. Dominant mutation transforming dark brown colour to the light-grey seemed to be present in the 1/29 chromosome locus, near the centromere region.