Photorhabdus luminescens is an enterobacterium that is symbiotic with soil entomopathogenic nematodes and pathogenic to a wide range of insects. P. luminescens promotes its own transmission among susceptible insect populations using its nematode host as vector 1 . Its life cycle comprises a symbiotic stage in the nematode's gut and a virulent stage in the insect larvae, which it kills through toxemia and septicemia. After the nematode attacks a prey insect and P. luminescens is released, the bacterium produces a wide variety of virulence factors ensuring rapid insect killing. Bioconversion of the insect cadaver by exoenzymes produced by the bacteria allows the bacteria to multiply and the nematode to reproduce. During this process P. luminescens produces antibiotics to prevent invasion of the insect cadaver by bacterial or fungal competitors. Finally, elimination of competitors allows P. luminescens and the nematode to reassociate specifically before leaving the insect cadaver 2,3 .To better understand this complex life style, we determined the genome sequence of P. luminescens subspecies laumondii strain TT01 4 , a symbiont of the nematode Heterorhabditis bacteriophora isolated on Trinidad and Tobago. RESULTS General featuresStrain TT01 possesses a single circular chromosome of 5,688,987 bp with an average GC content of 42.8%. No plasmid replicon was found.A total of 4,839 protein-coding genes, including 157 pseudogenes, seven complete sets (23S, 5S and 16S) of ribosomal RNA operons and 85 tRNA genes, were predicted ( Fig. 1; Supplementary Table 1 online). Toxins against insectsMore toxin genes were predicted in the P. luminescens genome than in any other bacterial genome sequenced yet. A large number of these toxins may be involved in the killing of a wide variety of insects. Some may act synergistically or use redundancy for 'overkill' 5 , ensuring a quick death of the host. In addition, some may kill insects by interfering with their development. In the TT01 genome, two paralogs, plu4092 and plu4436, encode proteins similar to juvenile hormone esterases (JHEs) of the insect Leptinotarsa decemlineata 6 . Juvenile hormone maintains the insect in a larval state. Its inactivation by JHE allows metamorphosis to proceed. JHEs may be used to trigger the insect endocrine machinery at an inappropriate time and thus represents a promising approach for insect control 7 . These genes are located downstream of highly related orphan genes (plu4093 and plu4437), suggesting a locus duplication.The toxicity of the proteins encoded by these two loci was verified experimentally. Two Escherichia coli clones, containing the recombinant BAC1A02 and BAC8C11, were shown to be toxic toward insects. BAC1A02, which contains the locus plu4093-plu4092, exhibited substantial oral toxicity toward three mosquito species, Aedes aegypti,

Class II genes of the human major histocompatibility complex (MHC) are polymorphic. Allelic variation of the coding region of these genes is involved in the antigen presentation and is associated with susceptibility to certain autoimmune diseases. The DR region is unique among human class II regions in that multiple DRB genes are expressed. Differential expression of the different DRB loci has been demonstrated, and we sequenced the proximal promoter region of the HLA-DRB genes, known to be involved in the regulation of these genes. We found locus-specific and allele-specific nucleotide variations in their regulatory regions and we determined the relationship between the regulatory regions of HLA-DRB genes. This polymorphism found in the regulatory conserved boxes could be involved in the observed differential expression of DRB loci. In addition, we found a polymorphism between the regulatory regions of DRB1 alleles which might be involved in an allele-specific regulation and therefore could be considered as an additional factor in susceptibility to autoimmune diseases.

SUMMARYSusceptibility to rheumatoid arthritis (RA) is associated with defined HLA-DRB1 alleles. However the molecular basis of this association is not known. Peculiarities in the expression of disease-linked DRB1 alleles could be involved since in healthy controls HLA-DRB1 gene expression varies according to the alleles in B cells. Peripheral blood B cells of healthy controls and RA patients were examined for their level of allelic DRB1 transcripts using a competitive PCR approach. Levels of DRB1 transcripts were greatly modified in RA and influenced by HLA-DRB1 genotype: patients with double dose of RAassociated alleles displayed up-regulated amounts of DRB1 gene transcripts whereas patients carrying either a single or no at risk allele had low levels of DRB1 transcripts, compared to control individuals. These differential levels of DRB1 gene expression were not influenced in any way by clinical, biological or therapeutic features of the patients. Various amounts of DRB1 mRNA may be related to variations of the density of DR molecules on B cells and consequently could influence the response of CD4 T cells. This particular regulation of DRB1 gene expression in RA patients could therefore represent one of the molecular mechanisms involved in the association of HLA DRB1 genes to RA.

The genome of the Autographa californica Multinucleocapsid Polyhedrosis Virus (AcMNPV) contains nine interspersed homologous regions (hrs) that function as potent enhancer sequences when linked in cis to either viral or heterologous RNA polymerase II-dependent promoters. Their activity is strongly increased by the binding of the major immediate early viral transregulator IE1 on 28-mer palindromic sites present in hrs. We show that hrs of AcMNPV additionally carry, in the interpalindromic sequences, a large number of cAMP response elements (CRE) and TPA response elements (TRE), known to bind ubiquitous cellular transcription factors of the bZIP family. Moreover, these clusters of CRE and TRE motifs are concentrated in hrs. Analysis of the 25 baculovirus genomes sequenced so far reveals that these motifs are evolutionary conserved in Lepidoptera NPVs, suggesting a functional role in the hr enhancer function. Consistently, EMSA experiments indicate that CRE and on a lesser extent TRE sites specifically bind insect host factors. Moreover, reporter assays reveal that these CRE sites have an additive stimulatory effect on RNAPol II-dependent transcription in Sf9 cells and are potentially able to synergize with the IE1-binding palindrome.

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In addition to polymorphism of the peptide-binding site, the density of the MHC class II molecules expressed on the membrane of APC could well play a significant role in the MHC-peptide-TCR interaction during the immune response. We therefore investigated the regulation of the expression of the HLA-DRB genes at the transcriptional level. A competitive PCR approach was used to estimate the quantities of the HLA-DRB transcripts in peripheral blood B cells. When comparing the amounts of steady-state mRNAs among the different DRB1 alleles, the DRB1 transcripts in the DR52 haplotype group were found to be 2.5 to 3.5 times more abundant than the DRB1*01 transcripts, 1.5 to 2 times more abundant than the DRB1*04 transcripts, and 7 times more abundant than the DRB1*08 transcripts. Within the DR52 haplotype group, the DRB1 and DRB3 transcripts had the same abundance. Taken together, these results are in good agreement with the previously reported transcriptional activities of the DRB promoters except for DRB1*04, thus suggesting a differential post-transcriptional regulation among the DRB1 mRNAs.

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The regulation of HLA class II genes is of particular interest with regard to the modulation of the immune response. The polymorphism of their coding regions is directly involved in the specificity of the Ag presentation, and their level of expression affects the extent of T cell activation. Previously, we have described an allelic polymorphism in the proximal promoter regions of HLA-DRB genes. The aim of this study was to compare the transcriptional activities of the promoters of the DRB genes and DRB1 alleles in a transient expression system. We have demonstrated a marked difference in their promoter strengths, as determined by their relative abilities to initiate transcription of the chloramphenicol acetyltransferase reporter gene in human B cell lines. The polymorphism of the promoter regions has been mapped to the regulatory boxes, and, by using gel retardation experiments, we found a differential ability of the nuclear proteins to bind to the partially conserved X box regions. Taken together, our results demonstrate the functional consequences of the allelic polymorphism of the proximal promoter regions of the DRB genes. These findings strongly suggest the existence, for the HLA-DR genes, of an interdependence between the polymorphism of the coding regions, which directly affects the capacity of peptide binding, and the polymorphism of the regulatory regions, which influences the transcriptional activities of the promoters.