RESUMO
Sjogren's syndrome (SS) is characterized by salivary gland leukocytic infiltrates and impaired salivation (xerostomia). Cox-2 (Ptgs2) is located on chromosome 1 within the span of the Aec2 region. In an attempt to demonstrate that COX-2 drives antibody-dependent hyposalivation, NOD.B10 congenic mice bearing a Cox-2flox gene were generated. A congenic line with non-NOD alleles in Cox-2-flanking genes failed manifest xerostomia. Further backcrossing yielded disease-susceptible NOD.B10 Cox-2flox lines; fine genetic mapping determined that critical Aec2 genes lie within a 1.56 to 2.17Mb span of DNA downstream of Cox-2. Bioinformatics analysis revealed that susceptible and non-susceptible lines exhibit non-synonymous coding SNPs in 8 protein-encoding genes of this region, thereby better delineating candidate Aec2 alleles needed for SS xerostomia.
Assuntos
Cromossomos de Mamíferos , Predisposição Genética para Doença , Síndrome de Sjogren/complicações , Síndrome de Sjogren/genética , Xerostomia/etiologia , Animais , Mapeamento Cromossômico , Ciclo-Oxigenase 2/genética , Modelos Animais de Doenças , Feminino , Expressão Gênica , Humanos , Masculino , Camundongos , Camundongos Congênicos , Camundongos Endogâmicos NOD , Fases de Leitura Aberta , Recombinação Genética , Glândulas Salivares/metabolismo , Glândulas Salivares/patologia , Sialadenite/genética , Sialadenite/imunologia , Sialadenite/patologia , Síndrome de Sjogren/imunologiaRESUMO
Group II introns are mobile retroelements that invade their cognate intron-minus gene in a process known as retrohoming. They can also retrotranspose to ectopic sites at low frequency. Previous studies of the Lactococcus lactis intron Ll.LtrB indicated that in its native host, as in Escherichia coli, retrohoming occurs by the intron RNA reverse splicing into double-stranded DNA (dsDNA) through an endonuclease-dependent pathway. However, in retrotransposition in L. lactis, the intron inserts predominantly into single-stranded DNA (ssDNA), in an endonuclease-independent manner. This work describes the retrotransposition of the Ll.LtrB intron in E. coli, using a retrotransposition indicator gene previously employed in our L. lactis studies. Unlike in L. lactis, in E. coli, Ll.LtrB retrotransposed frequently into dsDNA, and the process was dependent on the endonuclease activity of the intron-encoded protein. Further, the endonuclease-dependent insertions preferentially occurred around the origin and terminus of chromosomal DNA replication. Insertions in E. coli can also occur through an endonuclease-independent pathway, and, as in L. lactis, such events have a more random integration pattern. Together these findings show that Ll.LtrB can retrotranspose through at least two distinct mechanisms and that the host environment influences the choice of integration pathway. Additionally, growth conditions affect the insertion pattern. We propose a model in which DNA replication, compactness of the nucleoid and chromosomal localization influence target site preference.
Assuntos
Elementos de DNA Transponíveis , DNA Bacteriano/genética , Íntrons/genética , Lactococcus lactis/genética , Retroelementos/genética , Proteínas de Bactérias , Replicação do DNA , DNA de Cadeia Simples/genética , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Modelos Biológicos , Modelos GenéticosRESUMO
Initiation codon context is an important determinant of translation initiation rates in both prokaryotes and eukaryotes. Such sequences include the Shine- Dalgarno ribosome-binding site, as well as other motifs surrounding the initiation codon. One proposed interaction is between the base immediately preceding the initiation codon (-1 position) and the nucleotide 3' to the tRNAf(Met) anticodon, at position 37. Adenine is conserved at position 37, and a uridine at -1 has been shown in vitro to favor initiation. We have tested this model in vivo, by manipulating the chloroplast of the green alga Chlamydomonas reinhardtii, where the translational machinery is prokaryotic in nature. We show that translational defects imparted by mutations at the petA -1 position can be suppressed by compensatory mutations at position 37 of an ectopically expressed tRNA(fMet). The mutant tRNAs are fully aminoacylated and do not interfere with the translation of other proteins. Although this extended base pairing is not an absolute requirement for initiation, it may convey added specificity to transcripts carrying non-standard initiation codons, and/or preserve translational fidelity under certain stress conditions.
