Nucleotide sequence and organization of potato leafroll virus genomic RNA

The nucleotide sequence of the genomic RNA of potato leafroll virus was determined and its genetic organization deduced. The RNA is 5882 nucleotides long and contains 6 open reading frames (ORFs) encoding proteins of 70, 70, 56, 28, 23 and 17 kDa. The putative genes for the coat protein (23 kDa) and the RNA-dependent RNA polymerase (70 kDa) were identified by interviral amino acid sequence homologies. For expression of the different ORFs, translational frameshift and readthrough events are proposed.     

A U3 RNA pseudogene in mouse: sequence and organization in genomic DNA

A mouse U3 RNA pseudogene has been identified; it corresponds to a U3B full length coding sequence with a 3'-oligo(A) tail, precisely flanked at both ends by a pair of 15 bp direct repeats. These structural features suggest that it has arisen through an RNA-mediated mechanism involving an insertion at staggered nicks in the genome. Sequence data indicate that this mouse specimen has been generated by a different event as compared to the recently described rat pseudogenes. It represents the first reported case, for a pseudogene of this class, to be present at more than one copy per genome.     

Complete cDNA sequence,expression, alternative splicing,and genomic organization of the mouse Nfat5 gene

Members of the NFAT (nuclear factors of activated T cells) gene family have been investigated in numerous organisms, including man and mouse. All NFATs may be synthesized in several isoforms differing in amino or carboxy termini due to 5' and 3' alternative splicing of the corresponding mRNA. Recently, we mapped the murine Nfat5 gene to chromosome 8D. In the present paper we describe for the first time the complete sequence and primary structure of murine Nfat5, two new spliced isoforms, and the expression of murine Nfat5 in embryonic and adult mouse tissues.     

Goose-type lysozyme gene of the chicken: sequence, genomic organization and expression reveals major differences to chicken-type lysozyme gene

This report describes the cloning, sequencing and expression pattern of the chicken goose-type lysozyme gene. The cDNA sequence was found to have no homology to that of the chicken-type lysozyme gene and exhibits a completely different exon-intron organization. In addition, goose-type lysozyme displays an overlapping but different tissue expression pattern in chickens than chicken-type lysozyme.     

RNA recombination in a coronavirus: recombination between viral genomic RNA and transfected RNA fragments.

Mouse hepatitis virus (MHV), a coronavirus, has been shown to undergo a high frequency of RNA recombination both in tissue culture and in animal infection. So far, RNA recombination has been demonstrated only between genomic RNAs of two coinfecting viruses. To understand the mechanism of RNA recombination and to further explore the potential of RNA recombination, we studied whether recombination could occur between a replicating MHV RNA and transfected RNA fragments. We first used RNA fragments which represented the 5' end of genomic-sense sequences of MHV RNA for transfection. By using polymerase chain reaction amplification with two specific primers, we were able to detect recombinant RNAs which incorporated the transfected fragment into the 5' end of the viral RNA in the infected cells. Surprisingly, even the anti-genomic-sense RNA fragments complementary to the 5' end of MHV genomic RNA could also recombine with the MHV genomic RNAs. This observation suggests that RNA recombination can occur during both positive- and negative-strand RNA synthesis. Furthermore, the recombinant RNAs could be detected in the virion released from the infected cells even after several passages of virus in tissue culture cells, indicating that these recombinant RNAs represented functional virion RNAs. The crossover sites of these recombinants were detected throughout the transfected RNA fragments. However, when an RNA fragment with a nine-nucleotide (CUUUAUAAA) deletion immediately downstream of a pentanucleotide (UCUAA) repeat sequence in the leader RNA was transfected into MHV-infected cells, most of the recombinants between this RNA and the MHV genome contained crossover sites near this pentanucleotide repeat sequence. In contrast, when exogenous RNAs with the intact nine-nucleotide sequence were used in similar experiments, the crossover sites of recombinants in viral genomic RNA could be detected at more-downstream sites. This study demonstrated that recombination can occur between replicating MHV RNAs and RNA fragments which do not replicate, suggesting the potential of RNA recombination for genetic engineering.     

The mouse Kell blood group gene (Kel): cDNA sequence, genomic organization, expression, and enzymatic function

The human Kell blood group system is important in transfusion medicine, since Kell is a polymorphic protein and some of its antigens can cause severe reactions if mismatched blood is transfused, while maternal alloimmunization may lead to fetal and neonatal anemia. In humans, Kell is an Mr 93,000 type II membrane glycoprotein with endothelin-3-converting enzyme activity that is linked by a single disulfide bond to another protein, XK, that spans the membrane ten times. An absence of XK leads to clinical symptoms termed the McLeod syndrome. We determined the cDNA sequence of the mouse Kell homologue, the organization of the gene, expression of the protein and its enzymatic function on red cells. Comparison of human and mouse Kell cDNA showed 80% nucleotide and 74% amino acid sequence identity. Notable differences are that the mouse Kell protein has eight probable N-linked carbohydrate side chains, compared to five for human Kell, and that the mouse homologue has one more extracellular cysteine than human Kell protein. The mouse Kell gene (Kel), like its human counterpart, is similarly organized into 19 exons. Kel was located to proximal Chromosome 6. Northern blot analysis showed high expression in spleen and weaker levels in testis and heart. Western blot analysis of red cell membrane proteins demonstrated that mouse Kell glycoprotein has an apparent Mr of 110,000 and, on removal of N-linked sugars, 80,000. As in human red cells, Kell is disulfide-linked to XK and mouse red cells have endothelin-3-converting enzyme activity.     

Nucleotide sequence, genomic organization and cell-cycle-dependent expression of a Chlamydomonas 14-3-3 gene

Members of the 14-3-3 protein family have been identified as regulatory elements in intracellular signalling pathways and cell cycle control. Previously we reported the nucleotide sequence of a 14-3-3 cDNA cloned from the unicellular green alga Chlamydomonas reinhardtii. In this communication, we describe the nucleotide sequence, the genomic organization and the cell-cycle-dependent expression of the corresponding gene. The coding sequence of this gene was found to be interrupted by four introns of 124, 116, 81, and 659 bp, respectively. Introns 2-4 were found in conserved positions as compared to the Arabidopsis 14-3-3 genes. A counterpart to intron 1 absent in the Arabidopsis 14-3-3 genes was found in the human 14-3-3 epsilon gene.     

tRNAs in Trypanosoma brucei: genomic organization, expression, and mitochondrial import

The mitochondrial genome of Trypanosoma brucei does not encode tRNAs. Consequently, all mitochondrial tRNAs are imported from the cytosol and originate from nucleus-encoded genes. Analysis of all currently available T. brucei sequences revealed that its genome carries 50 tRNA genes representing 40 different isoacceptors. The identified set is expected to be nearly complete since all but four codons are accounted for. The number of tRNA genes in T. brucei is very low for a eukaryote and lower than those of many prokaryotes. Using quantitative Northern analysis we have determined the absolute abundance in the cell and the mitochondrion of a group of 15 tRNAs specific for 12 amino acids. Except for the initiator type tRNA(Met), which is cytosol specific, the cytosolic and the mitochondrial sets of tRNAs were qualitatively identical. However, the extent of mitochondrial localization was variable for the different tRNAs, ranging from 1 to 7.5% per cell. Finally, by using transgenic cell lines in combination with quantitative Northern analysis it was shown that import of tRNA(Leu)(CAA) is independent of its 5'-genomic context, suggesting that the in vivo import substrate corresponds to the mature, fully processed tRNA.     

Coding sequence, genomic organization, chromosomal localization, and expression pattern of the signalosome component Cops2: the mouse homologue of Drosophila alien

The Drosophila alien gene is highly homologous to the human thyroid receptor interacting protein, TRIP15/COPS2, which is a component of the recently identified signalosome protein complex. We identified the mouse homologue of Drosophila alien through homology searches of the EST database. We found that the mouse cDNA encodes a predicted 443-amino-acid protein, which migrates at approximately 50 kDa. The gene for the mouse alien homologue, named Cops2, includes 12 coding exons spanning approximately 30 kb of genomic DNA on the central portion of mouse chromosome 2. Mouse Cops2 is widely expressed in embryonic, fetal, and adult tissues beginning as early as E7.5. Mouse Cops2 cDNA hybridizes to two mRNA bands in all tissues at approximately 2.3 and approximately 4 kb, with an additional approximately 1.9-kb band in liver. Immunostaining of native and epitope tagged proteins localized the mouse Cops2 protein in both the cytoplasm and the nucleus, with larger amounts in the nucleus in some cells.     

Nucleotide sequence and genomic organization of bird minisatellites.

Two minisatellite loci from a Eurasian songbird, the willow warbler (Phylloscopus trochilus) were isolated, sequenced and used as probes to detect more than 20 related hypervariable loci. In addition, a sequence flanking one of the minisatellite loci was isolated, and used to study a VNTR locus. The bird minisatellites have a repeat unit of either 12 (AGGGAAGGGCTC) or 17 bp (GGGGACAGGGGACACCC), repeated in tandem 40-100 times per locus, and shows partial similarity to the sequence motifs of human minisatellites. These sequences are among the most variable minisatellites known, with the incidence per gamete of new length alleles estimated from family studies of warblers to about 5.6% per locus. The bird minisatellite alleles show mendelian inheritance and segregation analysis indicates that they are derived from families of sequences with members on several autosomal linkage groups. Some of the warbler core sequences cross-hybridize to hypervariable loci in other species of birds, mammals and fishes.     

Cloning,expression, and genomic organization of mouse mp29 gene

The convulsions of approximately 25% of epileptics are inadequately controlled by currently available medication; therefore the preparation of new antiepileptic drugs is of great interest. Aryl semicarbazones can be considered a new class of compounds presenting anticonvulsant activity. In addition, they can be orally administered and are more active as anticonvulsants than mephenytoin or phenobarbital. However, one disadvantage of these compounds is their low water solubility. As a strategy to circumvent this problem, a 1:1 inclusion compound of benzaldehyde semicarbazone (BS) and hydroxypropyl-beta-cyclodextrin (HP-beta-CD) was prepared and characterized. The anticonvulsant activities of the free semicarbazone and of the inclusion compound were evaluated in rats using the maximum electroshock and audiogenic seizures screenings. In both tests the minimum dose of compound necessary to produce activity decreases from 100mg/kg for the free semicarbazone to 35 mg/kg for the inclusion compound, indicating a significant increase in the bio-availability of the drug.