RGS7 is palmitoylated and exists as biochemically distinct forms

Regulator of G protein signaling (RGS) proteins are GTPase-activating proteins that modulate neurotransmitter and G protein signaling. RGS7 and its binding partners Galpha and Gbeta5 are enriched in brain, but biochemical mechanisms governing RGS7/Galpha/Gbeta5 interactions and membrane association are poorly defined. We report that RGS7 exists as one cytosolic and three biochemically distinct membrane-bound fractions (salt-extractable, detergent-extractable, and detergent-insensitive) in brain. To define factors that determine RGS7 membrane attachment, we examined the biochemical properties of recombinant RGS7 and Gbeta5 synthesized in Spodoptera frugiperda insect cells. We have found that membrane-bound but not cytosolic RGS7 is covalently modified by the fatty acid palmitate. Gbeta5 is not palmitoylated. Both unmodified (cytosolic) and palmitoylated (membrane-derived) forms of RGS7, when complexed with Gbeta5, are equally effective stimulators of Galpha(o) GTPase activity, suggesting that palmitoylation does not prevent RGS7/Galpha(o) interactions. The isolated core RGS domain of RGS7 selectively binds activated Galpha(i/o) in brain extracts and is an effective stimulator of both Galpha(o) and Galpha(i1) GTPase activities in vitro. In contrast, the RGS7/Gbeta5 complex selectively interacts with Galpha(o) only, suggesting that features outside the RGS domain and/or Gbeta5 association dictate RGS7-Galpha interactions. These findings define previously unrecognized biochemical properties of RGS7, including the first demonstration that RGS7 is palmitoylated.     

The use of the Insect Growth Regulators fenoxycarb and epofenonane against leafrollers in Integrated Pest Management in apple orchards

The applicability of the Insect Growth Regulators (IGR) with juvenile-hormone activity, epofenonane and fenoxycarb, was investigated in an Integrated Pest Management (IPM) program in six apple orchards during 1977–1983, against tortricid larvae. In some of these orchards Adoxophyes orana (F.v.R.) was the main pest species, and in others Pandemis heparana (Denn. & Schiff), Archips podana (Scop.) and Archips rosana (L.) were predominant.The IGR was sprayed twice at the time of emergence of last-instar larvae in spring, and resulted in adequate reduction of the numbers of the leafroller complex during the rest of the year. Reinfestation by moths never resulted in an increase to a harmful population level. IGR treatments kept fruit damage by leafrollers at a low level.

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Utilisation de deux régulateurs de croissance d'insectes (RCI), l'épofénonane et le fénoxycarbe, en lutte intégrée contre les tordeuses des vergers de pommiers

Résumé L'épofénonane et le fénoxycarbe, deux régulateurs de croissance d'insectes (RCI) analogues de l'hormone juvénile, ont été testés contre les tordeuses de 1977 à 1983 dans 6 vergers de pommiers conduits en lutte intégrée. Dans certains vergers, l'espèce la plus abondante était Adoxophyes orana, dans d'autres par contre Pandemis heparana, Archips podana ou Archips rosana étaient dominantes.Cette étude vise à déterminer: (a) si l'immigration à partir de vergers voisins, en particulier de l'espèce bivoltine A. orana, peut compromettre le succès de la lutte; (b) si deux applications annuelles de RCI suffisent pour lutter simultanément contre plusieurs espèces différentes de tordeuses.L'importance de l'immigration est estimée en comparant les résultats obtenus avec les RCI dans des vergers isolés et non isolés.Deux traitements aux RCI sont effectués au printemps au moment où les larves de la génération hivernante sont au dernier stade. Ces traitements conduisant à une réduction remarquable pour le reste de l'année du complexe des tordeuses y compris A. orana, quelque soit la situation du verger. Une réinfestation ultérieure n'a jamais provoqué un dépassement du seuil économique de tolérance. L'application des RCI a permis de maintenir les attaques de tordeuses sur fruits à un bas niveau. Les RCI preentent un intérêt considérable pour la lutte intégrée contre les tordeuses car les autres moyens sélectifs disponibles actuellement sont peu efficaces.

     

Drug screening assay based on the interaction of intact Keap1 and Nrf2 proteins in cancer cells

Background

The Nrf2–Keap1 interaction is the major regulatory pathway for cytoprotective responses against oxidative and electrophilic stresses. Keap1, a substrate protein of a Cul3-dependent E3 ubiquitin ligase complex, is a negative regulator of Nrf2. The use of chemicals to regulate the interaction between Keap1 and Nrf2 has been proposed as a strategy for the chemoprevention of degenerative diseases and cancers.

Modeled structure of the whole regulator G-protein signaling-2

There is an increasing interest towards the mechanism by which regulators of G-protein signaling regulate signals of G-protein-coupled receptors. RGS2 is a regulator of Gq protein signaling (RGS), the N-terminal region of which is known to contain determinants for G protein-coupled receptor recognition, but its structure is still unknown. To understand the molecular basis for this recognition, the three-dimensional model of RGS2, including N-terminal region and RGS box, was modeled. For this, RGS4 box structure and data from circular dichroism study of RGS2 N-terminal region were used. Then, membrane-targeting activity of the RGS2 amphipathic helix contained in the N-terminal region was investigated. Furthermore, in cellulo study provided first evidence that an internal sequence within the N-terminal region of RGS2 is involved in RGS2 regulation of cholecystokinin receptor-2 signal. RGS2 modeled structure can now serve to study molecular recognition of RGS2 by signaling molecules.     

Domain Structure of Virulence-associated Response Regulator PhoP of Mycobacterium tuberculosis: ROLE OF THE LINKER REGION IN REGULATOR-PROMOTER INTERACTION(S)

The PhoP and PhoR proteins from Mycobacterium tuberculosis form a highly specific two-component system that controls expression of genes involved in complex lipid biosynthesis and regulation of unknown virulence determinants. The several functions of PhoP are apportioned between a C-terminal effector domain (PhoPC) and an N-terminal receiver domain (PhoPN), phosphorylation of which regulates activation of the effector domain. Here we show that PhoPN, on its own, demonstrates PhoR-dependent phosphorylation. PhoPC, the truncated variant bearing the DNA binding domain, binds in vitro to the target site with affinity similar to that of the full-length protein. To complement the finding that residues spanning Met¹ to Arg¹³⁸ of PhoP constitute the minimal functional PhoPN, we identified Arg¹⁵⁰ as the first residue of the distal PhoPC domain capable of DNA binding on its own, thereby identifying an interdomain linker. However, coupling of two functional domains together in a single polypeptide chain is essential for phosphorylation-coupled DNA binding by PhoP. We discuss consequences of tethering of two domains on DNA binding and demonstrate that linker length and not individual residues of the newly identified linker plays a critical role in regulating interdomain interactions. Together, these results have implications for the molecular mechanism of transmission of conformation change associated with phosphorylation of PhoP that results in the altered DNA recognition by the C-terminal domain.