ABSTRACT
Increase of influenza vaccine production capacity in developing countries has been identified as an important element of global pandemic preparedness. Nevertheless, technology transfer for influenza vaccine production to developing country vaccine manufacturers has proven difficult because of lack of interested technology providers. As an alternative to an individual provider-recipient relationship, a technology and training platform (a "hub") for a generic non-proprietary process was established at a public sector European manufacturer's site. The conditions for setting up such a platform and the potential applicability of this model to other biologicals are discussed.
Subject(s)
Disease Outbreaks/prevention & control , Drug Industry/methods , Influenza Vaccines , Influenza, Human/epidemiology , Influenza, Human/prevention & control , Technology Transfer , Developing Countries , HumansABSTRACT
Short retroposons can be used as natural phylogenetic markers. By means of hybridization and PCR analysis, we demonstrate that B2 retroposon copies are present only in the three rodent families: Muridae, Cricetidae, and Spalacidae. This observation highlights the close phylogenetic relation between these families. Two novel B2-related retroposon families, named DIP and MEN elements, are described. DIP elements are found only in the genomes of jerboas (family Dipodidae) and birch mice (family Zapodidae), demonstrating the close relationship between these rodents. MEN element copies were isolated from the squirrel, Menetes berdmorei, but were not detected in three other species from the family Sciuridae. The MEN element has an unusual dimeric structure: the left and right monomers are B2- and B1-related sequences, respectively. Comparison of the B2, DIP, MEN, and 4.5S1 RNA elements revealed an 80-bp core sequence located at the beginning of the B2 superfamily retroposons. This observation suggests that these retroposon families descended from a common progenitor. A likely candidate for this direct progenitor could be the ID retroposon.
Subject(s)
Evolution, Molecular , Retroelements , Rodentia/physiology , Animals , Base Sequence , In Situ Hybridization , Molecular Sequence Data , Muridae , Polymerase Chain Reaction , Sciuridae , Sequence Homology, Nucleic Acid , Species SpecificityABSTRACT
Lymphocytes regulate their responsiveness to IL-2 through the transcriptional control of the IL-2R alpha gene, which encodes a component of the high affinity IL-2 receptor. In the mouse IL-2R alpha gene this control is exerted via two regulatable elements, a promoter proximal region, and an IL-2-responsive enhancer (IL-2rE) 1.3 kb upstream. In vitro and in vivo functional analysis of the IL-2rE in the rodent thymic lymphoma-derived, CD4- CD8- cell line PC60 demonstrated that three separate elements, sites I, II, and III, were necessary for IL-2 responsiveness; these three sites demonstrate functional cooperation. Site III contains a consensus binding motif for members of the Ets family of transcription factors. Here we demonstrate that Elf-1, an Ets-like protein, binds to site III and participates in IL-2 responsiveness. In vitro site III forms a complex with a protein constitutively present in nuclear extracts from PC60 cells as well as from normal CD4- CD8- thymocytes. We have identified this molecule as Elf-1 according to a number of criteria. The complex possesses an identical electrophoretic mobility to that formed by recombinant Elf-1 protein and is super-shifted by anti-Elf-1 antibodies. Biotinylated IL-2rE probes precipitate Elf-1 from PC60 extracts provided site III is intact and both recombinant and PC60-derived proteins bind with the same relative affinities to different mutants of site III. In addition, by introducing mutations into the core of the site III Ets-like motif and comparing the corresponding effects on the in vitro binding of Elf-1 and the in vivo IL-2rE activity, we provide strong evidence that Elf-1 is directly involved in IL-2 responsiveness. The nature of the functional cooperativity observed between Elf-1 and the factors binding sites I and II remains unresolved; experiments presented here however suggest that this effect may not require direct interactions between the proteins binding these three elements.
Subject(s)
DNA-Binding Proteins/metabolism , Enhancer Elements, Genetic , Interleukin-2/pharmacology , Receptors, Interleukin-2/genetics , T-Lymphocytes/metabolism , Transcription Factors/metabolism , Animals , Base Sequence , Binding Sites , DNA-Binding Proteins/genetics , Ephrin-A2 , Mice , Molecular Sequence Data , Mutation , Nuclear Proteins/metabolism , Protein Binding , Proto-Oncogene Proteins/genetics , Proto-Oncogene Proteins c-ets , Transcription Factors/genetics , Tumor Cells, CulturedSubject(s)
DNA Transposable Elements , Genetic Markers , Phylogeny , Animals , Base Sequence , DNA/genetics , DNA Primers , Molecular Sequence Data , Polymerase Chain Reaction , RodentiaABSTRACT
It is suggested to use short retroposons in the study of phylogenetic relationship between different species. It is likely that the presence of the above mentioned retroposon's copies in different genomes testifies to identical origin of these species. The fact that the genomes of different species include not the same but similar retroposons is considered as evidence for identical phylogenetic origin. A new retroposon was found in genomic DNA of Dipodidae (DIP element). This element has homology with the B2 element of mouse, rat (Muridae) and hamster (Cricetidae). The B2-like elements were not detected in Gliridae and Cavia DNAs, hence we suggest the earlier divergence of these animals from the route leading to Muridae and Cricetidae. The structural analysis of the DIP element supported the role of the nucleotide sequence of 4,5 S(1) RNA in the process of appearing the B2 superfamily of retroposons in Rodents.
