Specific recognition of the viral protein UL18 by CD85j/LIR-1/ILT2 on CD8+ T cells mediates the non-MHC-restricted lysis of human cytomegalovirus-infected cells

Immune evasion mechanisms of human CMV are known; however, the immune control of infection remains poorly elucidated. We show that interaction between the viral protein UL18 on infected cells and the invariant receptor CD85j/LIR-1/ILT2 expressed on CTL is relevant for the control of infection. Resting and activated CD8(+) T cells lysed UL18 expressing cells, whereas cells infected with CMV defective for UL18 were not killed. Lysis was not dependent on CD8(+) T cell Ag specificity, MHC-unrestricted and specifically blocked by anti-CD85j and anti-UL18 mAb. Moreover, soluble recombinant UL18Fc immunoprecipitated CD85j from T cells. Activation is mediated by CD85j and its pathway is unrelated to CD3/TCR engagement. UL18 is detected in immunocompromised patients with productive infection and the mechanism used in vivo by human CMV to ensure survival of the immunocompetent host may be mediated by activation signals delivered by infected cells to T lymphocytes via UL18/CD85j interactions.     

CD85j (leukocyte Ig-like receptor-1/Ig-like transcript 2) inhibits human osteoclast-associated receptor-mediated activation of human dendritic cells

Immature dendritic cells (DCs) derived from freshly isolated human monocytes were used to evaluate the effect of the inhibiting receptor CD85j (leukocyte Ig-like receptor-1/ILT2) on activation induced by cross-linking of the human osteoclast-associated receptor (hOSCAR). CD85j and hOSCAR were expressed consistently at the same density on monocytes and on monocyte-derived DCs (both immature and mature). Cross-linking of hOSCAR, which activates via the FcR-associated gamma-chain, induced Ca(2+) flux in DCs. Concomitant cross-linking of anti-CD85j mAb abolished this early activation event. Likewise, CD85j stimulation strongly reduced IL-8 and IL-12 production by hOSCAR-activated DCs. Inhibition of DCs via CD85j also impaired their ability to enhance Ag-specific T cell proliferation induced by hOSCAR. Finally, because hOSCAR prevents apoptosis of DCs in the absence of growth/survival factors, CD85j cross-linking was able to counteract completely this antiapoptotic effect and to reduce Bcl-2 expression enhanced by hOSCAR stimulation. Thus, CD85j is an inhibiting receptor that is functional in human DCs.     

Serum response factor, an enriched cardiac mesoderm obligatory factor, is a downstream gene target for Tbx genes

We tested the idea that T-box factors direct serum response factor (SRF) gene activity early in development. Analysis of SRF-LacZ "knock-in" mice showed highly restricted expression in early embryonic cardiac and skeletal muscle mesoderm and neuroectoderm. Examination of the SRF gene for regulatory regions by linking the promoter and 5'-flanking sequences, up to 5.5 kb, failed to target LacZ transgene activity to the heart and the tail pre-somitic mesenchyme. However, linkage of a minimal SRF promoter with the SRF 3'-untranslated region (UTR), inundated with multimeric T-box binding sites (TBEs), restored robust reporter gene activity to embryonic heart and tail. Finer dissection of the 3'-UTR to a small cluster of TBEs also stimulated transgene activity in the cardiac forming region and the tail, however, when the TBEs contained within these DNA sequences were mutated, preventing Tbx binding, transgene activity was lost. Tbx2, Tbx5, and the cardiac-enriched MYST family histone acetyltransferase TIP60, were observed to be mutual interactive cofactors through the TIP60 zinc finger and the T-box of the Tbx factors. In SRF-null ES cells, TIP60, Tbx2, and Tbx5 were sufficient to stimulate co-transfected SRF reporter activity, however this activity required the presence of the SRF 3'-UTR. SRF gene transactivation was blocked by two distinct TIP60 mutants, in which either the histone acetyltransferase domain was inactivated or the Zn finger-protein binding domain was excised. Our study supports the idea that SRF embryonic cardiac gene expression is dependent upon the SRF 3'-UTR enhancer, Tbx2, Tbx5, and TIP60 histone acetyltransferase activity.     

Isolation of competition-defective mutants of Rhizobium fredii.

We coupled Tn5 mutagenesis with a competition assay to isolate mutants of Rhizobium fredii USDA 257 that are defective in competition for nodulation of soybeans. Two mutants with single Tn5 inserts in the chromosome showed reduced competitiveness in vermiculite but were identical to the wild-type strain in symbiotic properties when inoculated alone. Recombination of Tn5 and flanking genomic regions cloned from the mutants into the parent strain showed that Tn5 was responsible for the mutant phenotype.     

Evidence that the phytochrome gene family in black cottonwood has one PHYA locus and two PHYB loci but lacks members of the PHYC/F and PHYE subfamilies

The phytochrome photoreceptors play important roles in the photoperiodic control of vegetative bud set, growth cessation, dormancy induction, and cold-hardiness in trees. Interestingly, ecotypic differences in photoperiodic responses are observed in many temperate- zone tree species. Northern and southern ecotypes of black cottonwood (Populus trichocarpa Torr. & Gray), for example, exhibit marked differences in the timing of short-day-induced bud set and growth cessation, and these responses are controlled by phytochrome. Therefore, as a first step toward determining the molecular genetic basis of photoperiodic ecotypes in trees, we characterized the phytochrome gene (PHY) family in black cottonwood. We recovered fragments of one PHYA and two PHYB using PCR-based cloning and by screening a genomic library. Results from Southern analyses confirmed that black cottonwood has one PHYA locus and two PHYB loci, which we arbitrarily designated PHYB1 and PHYB2. Phylogenetic analyses which included PHY from black cottonwood, Arabidopsis thaliana and tomato (Solanum lycopersicum) suggest that the PHYB/D duplications in these species occurred independently. When Southern blots were probed with PHYC, PHYE, and PHYE heterologous probes, the strongest bands that we detected were those of black cottonwood PHYA and/or PHYB. These results suggest that black cottonwood lacks members of the PHYC/F and PHYE subfamilies. Although black cottonwood could contain additional PHY that are distantly related to known angiosperm PHY, our results imply that the PHY family of black cottonwood is less complex than that of other well-characterized dicot species such as Arabidopsis and tomato. Based on Southern analyses of five black cottonwood genotypes representing three photoperiodic ecotypes, substantial polymorphism was detected for at least one of the PHYB loci but not for the PHYA locus. The novel character of the PHY family in black cottonwood, as well as the differences in polymorphism we observed between the PHYA and PHYB subfamilies, indicates that a number of fundamental macro- and microevolutionary questions remain to be answered about the PHY family in dicots.      

Cerebellar neurones: Differentiation and modulation of sensitivity to excitotoxic treatment

The neurite outgrowth and adhesion complex (NOAC), isolated from rabbit sera has been dissociated in its major components by reverse-phase chromatography in HPLC by using a C₁₈ column. SDS-PAGE analisys of the active fractions revealed the presence of three major bands of approximately 100, 70 and 50 kDa. Studies on the biological activity of NOAC were carried out on rat cerebellar granule cells. NOAC-cultured cells exhibit a marked resistance to excitotoxic stimuli carried by glutamate.