The roles of IL-12 and IL-23 in CD8 T cell-mediated immunity against Listeria monocytogenes: Insights from a DC vaccination model

Listeria monocytogenes infection induces a strong inflammatory response characterized by the production of IL-12 and IFN-γ and protective immunity against this pathogen is dependent on CD8⁺ T cells (CTL). Recent studies have suggested that these inflammatory cytokines affect the rate of memory CD8⁺ T cell generation as well as the number of short-lived effector cells generated. The role of the closely related cytokine, IL-23, in this response has not been examined. We hypothesized that IL-12 and IL-23 produced by dendritic cells collectively enhance the generation and function of memory cells. To test this hypothesis, we employed a DC vaccination approach. Mice lacking IL-12 and IL-23 were vaccinated with wild-type (WT), IL-12^−/−, or IL-12/23^−/− DC and protection to Lm was monitored. Mice vaccinated with WT and IL-12^−/− DC were resistant to lethal challenge with Lm. Surprisingly, mice vaccinated with IL-12/23^−/− DC exhibited significantly reduced protection when challenged. Protection correlated with the relative size of the memory pools generated. In summary, these data indicate that IL-23 can partially compensate for the lack of IL-12 in the generation protective immunity against Lm.     

New insights into the interaction of ribosomal protein L1 with RNA

The RNA-binding ability of ribosomal protein L1 is of profound interest, since L1 has a dual function as a ribosomal structural protein that binds rRNA and as a translational repressor that binds its own mRNA. Here, we report the crystal structure at 2.6 A resolution of ribosomal protein L1 from the bacterium Thermus thermophilus in complex with a 38 nt fragment of L1 mRNA from Methanoccocus vannielii. The conformation of RNA-bound T.thermophilus L1 differs dramatically from that of the isolated protein. Analysis of four copies of the L1-mRNA complex in the crystal has shown that domain II of the protein does not contribute to mRNA-specific binding. A detailed comparison of the protein-RNA interactions in the L1-mRNA and L1-rRNA complexes identified amino acid residues of L1 crucial for recognition of its specific targets on the both RNAs. Incorporation of the structure of bacterial L1 into a model of the Escherichia coli ribosome revealed two additional contact regions for L1 on the 23S rRNA that were not identified in previous ribosome models.     

Interaction between programmed cell death 5 and calcineurin B-like interacting protein kinase 23 in Oryza sativa

Programmed cell death (PCD) is crucial for plants during development and stress survival. OsPDCD5, an ortholog to mammalian-programmed cell death 5, was previously cloned from rice (Oryza sativa, cv Zhenxian 97A), and its overexpression can induce PCD in transgenic rice. In the present study, immunoblotting analysis revealed that the OsPDCD5 protein was widely expressed in the tassel, leaf, leaf sheath, and different parts of the stem but not in the anther. RT-PCR analysis showed that OsPDCD5 was related to the senescence of leaf and root tissues as well as the development of stem tissues. Furthermore, OsPDCD5 was up-regulated by UV-B irradiation. Calcineurin B-like interacting protein kinase 23 (OsCIPK23), which is involved in the calcineurin B-like proteins (CLBs)/CBL-interacting protein kinases (CIPKs) signaling network, was identified as interacting with OsPDCD5 by yeast two-hybrid screening and subsequently confirmed by pull-down assay in vitro. Present findings may shed light on the investigation of the biochemical function of OsPDCD5 in rice.     

Disruption of ste23 gene affects composition profile and bioactivity of exopolysaccharide produced by Streptomyces sp. 139

AIMS: To study the function of the gene ste23 involved in the biosynthesis of Ebosin. METHODS AND RESULTS: In search of databases, the deduced product of the gene ste23 showed high homology to dTDP-4-dehydrorhamnose 3,5-epimerases. ste23 was replaced by a kanamycin resistance gene through double crossover. Compared with Ebosin, an exopolysaccharide (EPS) produced by wild-type Streptomyces sp. 139, the EPS produced by the ste23 mutant (designated EPS1) had a remarkably different monosaccharide composition and significantly diminished rhamnose content, though the molecular mass of EPS1 was similar to that of Ebosin. In addition, EPS1 lost the interleukin 1 (IL-1) antagonist activity in vitro. CONCLUSIONS: ste23 may be involved in the Ebosin biosynthesis in S. sp. 139. and its bioactivity. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first genetic work investigating functions of genes involved in EPS production in streptomyces by gene replacement of the pathway genes.     

Genetic diversity of phenazine- and pyoluteorin-producing pseudomonads isolated from green pepper rhizosphere

The genetic diversity among indigenous phenazine-1-carboxylic acid (PCA)-producing and pyoluteorin (Plt)-producing isolates of pseudomonads screened from green pepper rhizosphere was exploited in this study. A total of 48 bacterium isolates producing one or both of these antibiotics were screened from green pepper rhizosphere in diverse regions in China. Among these isolates, 45 could produce PCA, 3 could produce both PCA and Plt, and none could produce Plt only. Based on the restriction patterns of partial 16S and 16S-23S internal transcribed spacer (ITS) PCR fragments generated by enzyme HaeIII or HinfI, these isolates fell into 19 or 17 distinct groups respectively, indicating that there was a significant diversity among them. Polygenetic analysis of the partial 16S rDNA and 16S-23S ITS sequence from the representative in each group in the context of similar sequence from previously described bacterial species indicated that most isolates were closely related to the species of Pseudomonas fluorescens, P. putida, and Stenotrophomonas maltophilia. Some of these representatives of these isolates, then, are likely to be novel strains or species in these two genera. The GenBank accession numbers for DNA sequences of the partial 16S rDNA with ITS region in each isolate determined in this study were: GP30 DQ003219; GP127 DQ003220; GP83 DQ003221; GP42, DQ003222; GP59 DQ003223; GP50 DQ003224; GP36 DQ003225; GP110 DQ003226; GP26 DQ003227; GP37 DQ003228; GP60 DQ003229; GP31 DQ003230; GP57 DQ003231; GP75 DQ003232; GP115 DQ003233; GP65 DQ003234; GP32 DQ003235; GP76 DQ003236; GP78 DQ003237.     

Immune enhancement and anti-tumour activity of IL-23

Immunotherapy, including the use of cytokines and/or modified tumour cells immune stimulatory cytokines, can enhance the host anti-tumour immune responses. Interleukin-23 (IL-23) is a relative novel cytokine, which consists of a heterodimer of the IL-12p40 subunit and a novel p19 subunit. IL-23 has biological activities similar to but distinct from IL-12. IL-23 can enhance the proliferation of memory T cells and the production of IFN-γ, IL-12 and TNF-α from activated T cells. IL-23 activates macrophages to produce TNF-α and nitric oxide. IL-23 can also act directly on dendritic cells and possesses potent anti-tumour and anti-metastatic activity in murine models of cancer. IL-23 can also induce a lower level of IFN-γ production compared with that induced by IL-12. This may make IL-23 an alternative and safer therapeutic agent for cancer, as IL-12 administration can lead to severe toxic side effects because of the extremely high levels of IFN-γ it induces.This article is a symposium paper from the Annual Meeting of the “International Society for Cell and Gene Therapy of Cancer”, held in Shenzhen, China, on 9–11 December 2005.     

Evidence for distinct functions for human DNA repair factors hHR23A and hHR23B

Rad23 proteins bind ubiquitinated substrates and the proteasome, consistent with an important role in protein degradation. Although human Rad23 proteins (hHR23A and hHR23B) have redundant roles in DNA repair, we determined they formed distinct interactions with proteasomes and multiubiquitinated proteins, but similar binding to Ataxin-3. Threonine-79 contributed to the weak proteasome-binding property of hHR23A, and its conversion to proline (T79P), which is the residue present in hHR23B, increased proteasome interaction. We also determined that hHR23A and hHR23B could be co-purified with unique proteolytic and stress-responsive factors from human breast cancer tissues, indicating that they have unique functions in vivo.     

Photosensitization of skin fibroblasts and HeLa cells by three chlorin derivatives: Role of chemical structure and delivery vehicle

The chemical nature of the sensitizer and its selective uptake by malignant cells are decisive to choose an appropriate biocompatible carrier, able to preserve the photosensitizing characteristics of the dye. In this paper we demonstrate the photodynamic properties of three chlorins, derived from chlorophyll a, and the usefulness of liposomal carriers to design pharmaceutical formulations. The chlorins have been quantitatively incorporated into stable liposomes obtained from a mixture of l-α-palmitoyloleoylphosphatidylcholine and l-α-dioleoylphosphatidylserine in a 13.5:1.5 molar ratio (POPC/OOPS-liposomes). The chlorin uptake by skin fibroblasts increases steadily, reaching in all cases a plateau level dependent on both the chlorin structure and the vehicle employed. The photophysical properties of the three chlorins in THF are nearly identical and fulfill the requirements for a PDT photosensitizer. Incorporation of chlorins into liposomes induces important changes in their photophysics, but does not impair their cellular uptake or their cell photosensitization ability. In fact we observe in the cells the same photophysical behavior as in THF solution. Specifically, we demonstrate, by recording the near-IR phosphorescence of ¹O₂, that the chlorins are able to photosensitize the production of ¹O₂ in the cell membrane. The cell-photosensitization efficiency depended on the chlorin and cell line nature, the carrier, and the length of pre-incubation and post-irradiation periods. The high photodynamic activity of chlorin-loaded liposomes and the possibility to design liposomal carriers to achieve a specific target site favors this approach to obtain an eventual pharmaceutical formulation.