Crystal structure of human PACRG in complex with MEIG1 reveals roles in axoneme formation and tubulin binding

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Mechanisms of HIV-1 evasion to the antiviral activity of chemokine CXCL12 indicate potential links with pathogenesis

HIV-1 infects CD4 T lymphocytes (CD4TL) through binding the chemokine receptors CCR5 or CXCR4. CXCR4-using viruses are considered more pathogenic, linked to accelerated depletion of CD4TL and progression to AIDS. However, counterexamples to this paradigm are common, suggesting heterogeneity in the virulence of CXCR4-using viruses. Here, we investigated the role of the CXCR4 chemokine CXCL12 as a driving force behind virus virulence. In vitro, CXCL12 prevents HIV-1 from binding CXCR4 and entering CD4TL, but its role in HIV-1 transmission and propagation remains speculative. Through analysis of thirty envelope glycoproteins (Envs) from patients at different stages of infection, mostly treatment-naïve, we first interrogated whether sensitivity of viruses to inhibition by CXCL12 varies over time in infection. Results show that Envs resistant (RES) to CXCL12 are frequent in patients experiencing low CD4TL levels, most often late in infection, only rarely at the time of primary infection. Sensitivity assays to soluble CD4 or broadly neutralizing antibodies further showed that RES Envs adopt a more closed conformation with distinct antigenicity, compared to CXCL12-sensitive (SENS) Envs. At the level of the host cell, our results suggest that resistance is not due to improved fusion or binding to CD4, but owes to viruses using particular CXCR4 molecules weakly accessible to CXCL12. We finally asked whether the low CD4TL levels in patients are related to increased pathogenicity of RES viruses. Resistance actually provides viruses with an enhanced capacity to enter naive CD4TL when surrounded by CXCL12, which mirrors their situation in lymphoid organs, and to deplete bystander activated effector memory cells. Therefore, RES viruses seem more likely to deregulate CD4TL homeostasis. This work improves our understanding of the pathophysiology and the transmission of HIV-1 and suggests that RES viruses’ receptors could represent new therapeutic targets to help prevent CD4TL depletion in HIV+ patients on cART.     

Distribution of neuropeptide Y immunoreactivity in human visual cortex and underlying white matter

Immunocytochemical techniques have been used to study neuropeptide Y (NPY) distribution in the human visual cortex (Brodman's areas 17, 18 and 19) NYP cell bodies belong mostly to inhibitory (multipolar and bitufted) but also to excitatory (bipolar and some pyramidal) neuronal types. Their distribution is similar in the three cortical areas studied: 20 to 40% of the NPY perikarya are located in the cortical gray matter, mostly in the deep layers, while the remaining 60 to 80% are located in the underlying white matter. Immunoreactive NPY processes form a rich network of intersecting fibers throughout the entire visual cortex. A superficial plexus (layers I and II) and a deep plexus (deep layer V and layer VI) of NPY fibers are present in areas 17, 18 and 19. In area 17, an additional well developed plexus is present in layers IVb and IVc. These plexuses receive branches from long parallel fibers arising from deep cortical layers or underlying white matter and terminating in superficial layers. Local or extrinsic NPY terminals wind around vessels in the cortex as well as in the white matter, and either penetrate them or form clusters of club endings on their walls. Our results suggest a role for NPY in human visual circuitry and in cortical blood flow regulation.     

Morphological correlates of serotonin-neuropeptide Y interactions in the rat suprachiasmatic nucleus: Combined radioautographic and immunocytochemical data

Summary The morphological substrate of putative serotonin (5-HT)/neuropeptide Y (NPY) interactions in thé suprachiasmatic nucleus (SCN) was investigated by combined radioautography and immunocytochemistry after intraventricular administration of (³H)5-HT in the rat. In the ventral portion of the SCN, the distribution of (³H)5-HT uptake sites overlapped closely the NPY-immunoreactive terminals. Previous investigations have shown that the dense 5-HT and NPY innervations of the SCN originate in different structures, i.e., the midbrain raphe nuclei and the ventral lateral geniculate nucleus, respectively. Accordingly, in the present study, destruction of 5-HT afferents by 5,7-dihydroxytryptamine was not found to induce any modification in NPY staining and, in ultrastructural immuno-radioautographic preparations, two distinct pools of axonal varicosities could be identified. Both 5-HT and NPY terminals established morphologically defined synaptic junctions, sometimes on the same neuronal target. Some cases of direct axo-axonic appositions between the two types of terminals were also encountered. These data constitute additional criteria for characterizing the cytological basis of the multiple transmitter interactions presumably involved in the function of the SCN as a central regulator of circadian biological rhythms.     

Selection of somatic hybrids between diploid clones of potato (Solanum tuberosum L.) transformed by direct gene transfer

Summary Five diploid potato clones have been transformed by electroporation of protoplasts with different selectable markers. The resulting diploid regenerated plants have been used in somatic hybridization. It has been shown that hybrid cell selection on the basis of antibiotic or herbicide resistances brought by the two parents of fusion is an efficient method for the recovery of tetraploid somatic hybrids.     

Poliovirus translation: a paradigm for a novel initiation mechanism

All eukaryotic cellular mRNAs, and most viral mRNAs, are blocked at their 5' ends with a cap structure (m7GpppX, where X is any nucleotide). Poliovirus, along with a small number of other animal and plant viral mRNAs, does not contain a 5' cap structure. Since the cap structure functions to facilitate ribosome binding to mRNA, translation of polio-virus must proceed by a cap-independent mechanism. Consistent with this, recent studies have shown that ribosomes can bind to an internal region within the long 5' noncoding sequence of poliovirus RNA. Possible mechanisms for cap-independent translation are discussed. Cap-independent translation of poliovirus RNA is of major importance to the mechanism of shut-off of host protein synthesis after infection. Moreover, it is likely to play a role in determining poliovirus neurovirulence and attenuation.     

Purification and characterization of a methionine-specific aminopeptidase from Salmonella typhimurium

An aminopeptidase specific for methionine (peptidase M) has been purified from wild-type and mutant Salmonella typhimurium strains. Recombinant peptidase M was also purified from Escherichia coli. These preparations were characterized with respect to their physicochemical properties using analytical ultracentrifugation, SDS/PAGE, isoelectric focusing, titration curve analysis, amino acid analysis, N-and C-terminal sequencing and various spectroscopic methods. Peptidase M activity is stimulated by Co2+, in agreement with previous studies using crude extracts of Salmonella. The purified preparations did not contain significant amounts of any metal. Enzymically important metal is loosely associated and lost during enzyme purification. Peptidase M was shown to contain seven free sulphydryl residues none of which are involved in either intra-or inter-molecular disulphide bonds. Most appear solvent-accessible as evidenced by their reactivity under native conditions. Limited modification of the sulphydryl residues with either iodoacetamide or 5,5'-dithiobis(2-nitrobenzoic acid) led to inactivation. Several cysteines were shown to be labelled to various degrees by peptide mapping of inactivated S-[14C]carboxymethylated protein. Whether cysteine modification affects enzymic activity directly (blocking an active site) or indirectly (by causing conformational change) remains to be established.     

Porcine D-amino acid oxidase: production of the biologically active enzyme in Escherichia coli

DNA molecules coding either for mature porcine D-amino acid oxidase or for truncated forms of the enzyme have been obtained by stepwise addition of synthetic oligonucleotides to a partial cDNA. Under the control of the lambda PL thermoregulatable promoter, these DNAs were respectively expressed in Escherichia coli as 36, 28 and 25 kilodalton polypeptides, specifically recognised by antibodies raised against the natural enzyme. None of the truncated proteins were biologically active whereas the mature recombinant species was able to hydrolyze D-alanine in vitro as efficiently as the natural product.