Complete genome sequence of Salmonella enterica serovar Typhimurium LT2.

Salmonella enterica subspecies I, serovar Typhimurium (S. typhimurium), is a leading cause of human gastroenteritis, and is used as a mouse model of human typhoid fever. The incidence of non-typhoid salmonellosis is increasing worldwide, causing millions of infections and many deaths in the human population each year. Here we sequenced the 4,857-kilobase (kb) chromosome and 94-kb virulence plasmid of S. typhimurium strain LT2. The distribution of close homologues of S. typhimurium LT2 genes in eight related enterobacteria was determined using previously completed genomes of three related bacteria, sample sequencing of both S. enterica serovar Paratyphi A (S. paratyphi A) and Klebsiella pneumoniae, and hybridization of three unsequenced genomes to a microarray of S. typhimurium LT2 genes. Lateral transfer of genes is frequent, with 11% of the S. typhimurium LT2 genes missing from S. enterica serovar Typhi (S. typhi), and 29% missing from Escherichia coli K12. The 352 gene homologues of S. typhimurium LT2 confined to subspecies I of S. enterica-containing most mammalian and bird pathogens-are useful for studies of epidemiology, host specificity and pathogenesis. Most of these homologues were previously unknown, and 50 may be exported to the periplasm or outer membrane, rendering them accessible as therapeutic or vaccine targets.

Complete sequence and genomic analysis of murine gammaherpesvirus 68.

Murine gammaherpesvirus 68 (gammaHV68) infects mice, thus providing a tractable small-animal model for analysis of the acute and chronic pathogenesis of gammaherpesviruses. To facilitate molecular analysis of gammaHV68 pathogenesis, we have sequenced the gammaHV68 genome. The genome contains 118,237 bp of unique sequence flanked by multiple copies of a 1,213-bp terminal repeat. The GC content of the unique portion of the genome is 46%, while the GC content of the terminal repeat is 78%. The unique portion of the genome is estimated to encode at least 80 genes and is largely colinear with the genomes of Kaposi's sarcoma herpesvirus (KSHV; also known as human herpesvirus 8), herpesvirus saimiri (HVS), and Epstein-Barr virus (EBV). We detected 63 open reading frames (ORFs) homologous to HVS and KSHV ORFs and used the HVS/KSHV numbering system to designate these ORFs. gammaHV68 shares with HVS and KSHV ORFs homologous to a complement regulatory protein (ORF 4), a D-type cyclin (ORF 72), and a G-protein-coupled receptor with close homology to the interleukin-8 receptor (ORF 74). One ORF (K3) was identified in gammaHV68 as homologous to both ORFs K3 and K5 of KSHV and contains a domain found in a bovine herpesvirus 4 major immediate-early protein. We also detected 16 methionine-initiated ORFs predicted to encode proteins at least 100 amino acids in length that are unique to gammaHV68 (ORFs M1 to 14). ORF M1 has striking homology to poxvirus serpins, while ORF M11 encodes a potential homolog of Bcl-2-like molecules encoded by other gammaherpesviruses (gene 16 of HVS and KSHV and the BHRF1 gene of EBV). In addition, clustered at the left end of the unique region are eight sequences with significant homology to bacterial tRNAs. The unique region of the genome contains two internal repeats: a 40-bp repeat located between bp 26778 and 28191 in the genome and a 100-bp repeat located between bp 98981 and 101170. Analysis of the gammaHV68, HVS, EBV, and KSHV genomes demonstrated that each of these viruses have large colinear gene blocks interspersed by regions containing virus-specific ORFs. Interestingly, genes associated with EBV cell tropism, latency, and transformation are all contained within these regions encoding virus-specific genes. This finding suggests that pathogenesis-associated genes of gammaherpesviruses, including gammaHV68, may be contained in similarly positioned genome regions. The availability of the gammaHV68 genomic sequence will facilitate analysis of critical issues in gammaherpesvirus biology via integration of molecular and pathogenetic studies in a small-animal model.

PRBI gene variants coding for length and null polymorphisms among human salivary Ps, PmF, PmS, and Pe proline-rich proteins (PRPs).

Six closely linked PRP (proline-rich protein) genes code for salivary PRPs that show frequent length and null polymorphisms. We report assignment of Ps proteins to the PRB1 gene, the derived primary structures of Ps 1 and Ps 2 proteins, and the molecular basis for some null alleles among PRB1-coded PRPs (Ps, PmF, PmS, and Pe). The derived primary structures of Ps 1 and Ps 2 proteins were determined by sequencing exon 3 of the different-length PRB1M (medium) and PRB1L (large) copies from subject C.J. with the Ps 1-2 phenotype. The PRB1L copy (coding for Ps 2) contained three additional tandem repeats within the Ps coding region, and the different-length Ps 1 and Ps 2 proteins can be explained on this basis. The molecular basis for the Ps 0 and the Pe- phenotypes was determined in another individual (M.V.O., a PRB2/1 fusion-gene heterozygote) with a single PRB1L copy. A premature stop mutation (CGA [Arg]-->TGA [stop]) occurred at residue 61 in the Ps-coding region. The identical mutation was found in the PRB1L and PRB1/2S (small) copies of a second individual (E.A.) with reduced Pe protein and the Ps 0 phenotype. This individual is a PRB1/2 fusion-gene heterozygote (Azen et al. 1992) with probably three mutated PRB1 copies (PRB1L-PRB1L-PRB1/2S). DNA sequences of the postulated crossover region of the PRB1/2S fusion-gene copy supported the postulated crossover. The PmF- and PmS- phenotypes in the three subjects were due to both the stop mutation and the lack of suitable proteolytic cleavage sites in the PRB1-coded precursor proteins.

Alleles at the PRB3 locus coding for a disulfide-bonded human salivary proline-rich glycoprotein (Gl 8) and a null in an Ashkenazi Jew.

From electrophoretic analysis, we identified in the saliva of an Ashkenazi Jew a disulfide-bonded major glycoprotein variant (Gl 8) that is a product of the proline-rich protein (PRP) locus PRB3. A previous study of this variant protein misidentified it as Pa 2 and as a product of a different PRP locus. The other PRB3 allele in this individual is an apparent null. To identify the mutations, we sequenced the tandemly repetitious exon 3 (the major protein-coding portions) of both alleles. A CGT----TGT (Arg----Cys) mutation was found in one allele (PRB3Scys), which accounts for the disulfide-bonded and peroxidase-modifying properties of Gl 8. A single nucleotide insertion was found in the other allele (PRB3Mnull) that leads to a frameshift with a premature termination codon that causes an apparent lack of gene expression. Null alleles are frequent at PRP loci coding for basic and glycosylated PRPs, and the mechanism described might explain other null phenotypes among PRPs. From nucleotide comparisons, a model of intragenic unequal crossing-over is proposed to explain, in part, the generation of the PRB3Mnull allele. The Gl 8 protein variant is found in Ashkenazi Jews (gene frequency around .008) but not in the general white, black, or Japanese populations. It is interesting that products of different PRP genes, Gl 8 from PRB3 and Pa 1 from PRH1, are both disulfide bonded and probably modify salivary peroxidase (part of an important intraoral antibacterial system) through formation of disulfide-bonded heterodimers.