The 'lipid raft' microdomain proteins reggie-1 and reggie-2 (flotillins) are scaffolds for protein interaction and signalling

Reggie-1 and reggie-2 are two evolutionarily highly conserved proteins which are up-regulated in retinal ganglion cells during regeneration of lesioned axons in the goldfish optic nerve. They are located at the cytoplasmic face of the plasma membrane and are considered to be 'lipid raft' constituents due to their insolubility in Triton X-100 and presence in the 'floating fractions'; hence they were independently named flotillins. According to our current view, the reggies subserve functions as protein scaffolds which form microdomains in neurons, lymphocytes and many other cell types across species as distant as flies and humans. These microdomains are of a surprisingly constant size of less than or equal to 0.1 mm in all cell types, whereas the distance between them is variable. The microdomains co-ordinate signal transduction of specific cell-surface proteins and especially of GPI (glycosylphosphatidylinositol)-anchored proteins into the cell, as is demonstrated for PrP(c) (cellular prion protein) in T-lymphocytes. These cells possess a pre-formed reggie cap scaffold consisting of densely packed reggie microdomains. PrP(c) is targeted to the lymphocyte reggie cap when activated by antibody cross-linking, and induces a distinct Ca(2+) signal. In developing zebrafish, reggies become concentrated in neurons and axon tracts, and their absence, after morpholino antisense RNA-knockdown, results in deformed embryos with reduced brains. Likewise, defects in Drosophila eye morphogenesis occur upon reggie overexpression in mutant flies. The defects observed in the organism, as well as in single cells in culture, indicate a morphogenetic function of the reggies, with emphasis on the nervous system. This complies with their role as scaffolds for the formation of multiprotein complexes involved in signalling across the plasma membrane.     

Synthesis and biological investigation of S-aryl-S-DABO derivatives as HIV-1 inhibitors

S-Aryl-S-DABO derivatives, a novel subclass of S-DABO anti-HIV-1 agents, were synthesized via Ullmann type reaction starting from the corresponding 2-thiouracils by the aid of microwave irradiation. The results of their evaluation as inhibitors of RT are reported together with their antiviral activity in cellular assays.     

Allogeneic Non-Adherent Bone Marrow Cells Facilitate Hematopoietic Recovery but Do Not Lead to Allogeneic Engraftment

Background

Non adherent bone marrow derived cells (NA-BMCs) have recently been described to give rise to multiple mesenchymal phenotypes and have an impact in tissue regeneration. Therefore, the effects of murine bone marrow derived NA-BMCs were investigated with regard to engraftment capacities in allogeneic and syngeneic stem cell transplantation using transgenic, human CD4⁺, murine CD4^−/−, HLA-DR3⁺ mice.

Methodology/Principal Findings

Bone marrow cells were harvested from C57Bl/6 and Balb/c wild-type mice, expanded to NA-BMCs for 4 days and characterized by flow cytometry before transplantation in lethally irradiated recipient mice. Chimerism was detected using flow cytometry for MHC-I (H-2D[b], H-2K[d]), mu/huCD4, and huHLA-DR3). Culturing of bone marrow cells in a dexamethasone containing DMEM medium induced expansion of non adherent cells expressing CD11b, CD45, and CD90. Analysis of the CD45⁺ showed depletion of CD4⁺, CD8⁺, CD19⁺, and CD117⁺ cells. Expanded syngeneic and allogeneic NA-BMCs were transplanted into triple transgenic mice. Syngeneic NA-BMCs protected 83% of mice from death (n = 8, CD4⁺ donor chimerism of 5.8±2.4% [day 40], P<.001). Allogeneic NA-BMCs preserved 62.5% (n = 8) of mice from death without detectable hematopoietic donor chimerism. Transplantation of syngeneic bone marrow cells preserved 100%, transplantation of allogeneic bone marrow cells 33% of mice from death.

Conclusions/Significance

NA-BMCs triggered endogenous hematopoiesis and induced faster recovery compared to bone marrow controls. These findings may be of relevance in the refinement of strategies in the treatment of hematological malignancies.     

A comparison of the wood anatomy of 11 species from two cerrado habitats (cerrado s.s. and adjacent gallery forest)

A comparative study of the secondary xylem (wood) anatomy of 11 species (38 specimens) occurring in cerrado s.s. and the adjacent gallery forest (both cerrado s.l. habitat) was made with the aim of identifying the anatomical characteristics of ecological value and correlating them with the environmental conditions. The anatomical features that vary, in general, between the two habitats are: growth ring distinctness (well or poorly defined); tyloses and deposits (more abundant in cerrado specimens); gelatinous fibres (more evident in cerrado specimens and in different patterns between habitats); variation in paratracheal and banded parenchyma (more abundant in cerrado); and more cells per parenchyma strand in cerrado. In general, gallery forest specimens have wider vessels, fewer vessels per square millimetre and larger intervessel pits, indicating more efficient water conduction, whereas cerrado s.s. specimens are the opposite, with low vulnerability and mesomorphy indices, demonstrating greater safety under conditions of water stress. © 2012 The Linnean Society of London, Botanical Journal of the Linnean Society, 2012, ?? , ??–??.     

TF — A novel cell-permeable and selective inhibitor of human protein kinase CK2 induces apoptosis in the prostate cancer cell line LNCaP

BackgroundAbnormally high activity of protein kinase CK2 is linked to various diseases including cancer. Therefore, the inhibition of CK2 is a promising therapeutic strategy to fight this disease.MethodsWe screened a library of synthetic molecules concerning their capacity to inhibit CK2. The activity of CK2 and their IC₅₀ and K_i values were determined by a capillary electrophoresis assay. The effects of the inhibitor in a cell culture model were analyzed by cell counting, a viability assay, cytofluorimetry and Western blot.ResultsThe best CK2 inhibitor found in this screen was 6,7-dichloro-1,4-dihydro-8-hydroxy-4-[(4-methylphenylamino)methylen]dibenzo [b,d]furan-3(2H)-one, which we refer to as “TF”. TF showed tight binding to CK2 with low IC₅₀ (29 nM) and K_i (15 nM) values. TF inhibited only seven out of 61 human kinases tested (> 70% inhibition). Incubation of LNCaP cells with 50 μM TF for 48 h decreased the intracellular CK2 activity by 50%, confirming that the inhibitor is membrane permeable. The decrease in activity was correlated with a severe reduction in cell viability. The reduction in cell viability is at least partly due to the induction of apoptosis.General significanceIn many cancers the protein kinase CK2 is significantly up-regulated and supports the neoplastic phenotype. New therapeutic strategies should be based on diverse reliable inhibitors to reverse the abnormal high levels to normal settings.     

3-(7-Azaindolyl)-4-arylmaleimides as potent, selective inhibitors of glycogen synthase kinase-3

A novel series of acyclic 3-(7-azaindolyl)-4-(aryl/heteroaryl)maleimides was synthesized and evaluated for activity against GSK-3beta and selectivity versus PKC-betaII, as well as a broad panel of protein kinases. Compounds 14 and 17c potently inhibited GSK-3beta (IC(50)=7 and 26 nM, respectively) and exhibited excellent selectivity over PKC-betaII (325 and >385-fold, respectively). Compound 17c was also highly selective against 68 other protein kinases. In a cell-based functional assay, both 14 and 17c effectively increased glycogen synthase activity by inhibiting GSK-3beta.