Association of dystrobrevin and regulatory subunit of protein kinase A: a new role for dystrobrevin as a scaffold for signaling proteins

The dystrophin-related and -associated protein dystrobrevin is a component of the dystrophin-associated protein complex, which directly links the cytoskeleton to the extracellular matrix. It is now thought that this complex also serves as a dynamic scaffold for signaling proteins, and dystrobrevin may play a role in this context. Since dystrobrevin involvement in signaling pathways seems to be dependent on its interaction with other proteins, we sought new insights and performed a two-hybrid screen of a mouse brain cDNA library using beta-dystrobrevin, the isoform expressed in non-muscle tissues, as bait. Among the positive clones characterized after the screen, one encodes the regulatory subunit RIalpha of the cAMP-dependent protein kinase A (PKA). We confirmed the interaction by in vitro and in vivo association assays, and mapped the binding site of beta-dystrobrevin on RIalpha to the amino-terminal region encompassing the dimerization/docking domain of PKA regulatory subunit. We also found that the domain of interaction for RIalpha is contained in the amino-terminal region of beta-dystrobrevin. We obtained evidence that beta-dystrobrevin also interacts directly with RIIbeta, and that not only beta-dystrobrevin but also alpha-dystrobrevin interacts with PKA regulatory subunits. We show that both alpha and beta-dystrobrevin are specific phosphorylation substrates for PKA and that protein phosphatase 2A (PP2A) is associated with dystrobrevins. Our results suggest a new role for dystrobrevin as a scaffold protein that may play a role in different cellular processes involving PKA signaling.     

ZAP-70 kinase regulates HIV cell-to-cell spread and virological synapse formation

HIV efficiently spreads in lymphocytes, likely through virological synapses (VSs). These cell-cell junctions share some characteristics with immunological synapses, but cellular proteins required for their constitution remain poorly characterized. We have examined here the role of ZAP-70, a key kinase regulating T-cell activation and immunological synapse formation, in HIV replication. In lymphocytes deficient for ZAP-70, or expressing a kinase-dead mutant of the protein, HIV replication was strikingly delayed. We have characterized further this replication defect. ZAP-70 was dispensable for the early steps of viral cycle, from entry to expression of viral proteins. However, in the absence of ZAP-70, intracellular Gag localization was impaired. ZAP-70 was required in infected donor cells for efficient cell-to-cell HIV transmission to recipients and for formation of VSs. These results bring novel insights into the links that exist between T-cell activation and HIV spread, and suggest that HIV usurps components of the immunological synapse machinery to ensure its own spread through cell-to-cell contacts.     

Infection of bovine oviduct cell cultures with Chlamydophila abortus

Bovine infertility is a major cause of loss in the livestock industry. In the present study bovine oviduct cell cultures were infected with a Chlamydophila abortus strain. A direct evaluation of infection was performed by means of May Grünwald-Giemsa and immunocytochemistry for chlamydial LPS, which revealed inclusion bodies and vacuolisation. SEM and TEM analysis of infected cells showed various degrees of cell damage and conglutination of microvilli. This finding suggests that cattle infertility may result from an alteration of oviduct environment caused by multiplication of C. abortus. This microorganism, among other infectious agents, could be considered a potential causative agent of bovine infertility.     

The phylogeny of polar fishes and the structure,function and molecular evolution of hemoglobin

Fishes thriving in polar habitats offer many opportunities for comparative approaches to understanding protein thermal adaptations. Investigations on the remarkable evolutionary adaptations to these environments of basic proteins such as hemoglobin, the oxygen carrier, can provide new insights into the mechanisms studied in temperate organisms and can shed light on convergent processes evolved in response to thermal adaptations. At the molecular level, hemoglobins are one of the most intriguing systems for studying the relationships between environmental conditions and adaptations. This review summarizes the current knowledge on molecular structure, biological function and phylogeny of hemoglobins of fish species living in both polar habitats but having different evolutionary histories. In benthic, non-migratory, cold-adapted fishes, the stability of thermal conditions may have generated no or few variations in selective pressures on globin sequences through evolutionary time, so that sequences retain the species phylogenetic “signal”. In pelagic, migratory, cold-adapted or temperate fishes, variations in selective pressures on globin sequences caused by variations in temperature accompanying the dynamic life style may have disrupted the phylogenetic “signal” in phenetic trees.     

Isolation and functional characterization of a cDNA coding a hydroxycinnamoyltransferase involved in phenylpropanoid biosynthesis in <Emphasis Type="Italic">Cynara cardunculus</Emphasis> L

Background  

Cynara cardunculus L. is an edible plant of pharmaceutical interest, in particular with respect to the polyphenolic content of its leaves. It includes three taxa: globe artichoke, cultivated cardoon, and wild cardoon. The dominating phenolics are the di-caffeoylquinic acids (such as cynarin), which are largely restricted to Cynara species, along with their precursor, chlorogenic acid (CGA). The scope of this study is to better understand CGA synthesis in this plant.