The complexities of G-protein-coupled receptor kinase function in Hedgehog signaling

Hedgehog (Hh) signaling is essential for proper tissue patterning and maintenance and has a substantial impact on human disease. While many of the main components and mechanisms involved in transduction of the Hh signal have been identified, the details of how the pathway functions are continually being refined. One aspect that has attracted much attention recently is the involvement of G-protein-coupled receptor kinases (GRKs) in the pathway. These regulators of G-protein-coupled receptor (GPCR) signaling have an evolutionarily-conserved function in promoting high-threshold Hh target gene expression through regulation of Smoothened (Smo), a GPCR family member that activates intracellular Hh signaling. Several models of how GRKs impact on Smo to increase downstream signaling have been proposed. Recently, we demonstrated that these kinases have surprisingly complex and conflicting roles, acting to limit signaling through the pathway while also promoting Smo activity. In addition to the previously described direct effects of Gprk2 on Smo activation, Gprk2 also indirectly affects Hh signaling by controlling production of the second messenger cyclic AMP to influence Protein kinase A activity.     

Mitochondrial haplotypes reveal olive fly (Bactrocera oleae) population substructure in the Mediterranean

The olive fly (Bactrocera oleae) is the most important olive tree (Olea europaea) pest. In the Mediterranean basin, where 98?% of its main hosts are concentrated, it causes major agricultural losses, due to its negative effect on production and quality of both olive and olive oil. Previous phylogeographic analyses have established that Mediterranean olive fly populations are distinct from other Old World populations, but did not agree on the specific population substructure within this region. In order to achieve a higher resolution of the diversity of olive fly populations, particularly in Central and Western Mediterranean (home to 70?% of the world production), we comparatively analyzed a set of samples from Portugal in the context of published mitochondrial sequences across the species' worldwide range. Strong evidence of population substructure was found in the Central and Western Mediterranean area, with two clearly separate phylogenetic branches. Together with previously published data, our results strongly support the existence of at least three distinct Mediterranean populations of the olive fly, raise the possibility of additional regional substructure and suggest specific avenues for future research. This knowledge can be instrumental in the development of better management and control strategies for a major pest of Mediterranean agriculture.     

An Extracellular Ion Pathway Plays a Central Role in the Cooperative Gating of a K2P K+ Channel by Extracellular pH

Proton-gated TASK-3 K⁺ channel belongs to the K_2P family of proteins that underlie the K⁺ leak setting the membrane potential in all cells. TASK-3 is under cooperative gating control by extracellular [H⁺]. Use of recently solved K_2P structures allows us to explore the molecular mechanism of TASK-3 cooperative pH gating. Tunnel-like side portals define an extracellular ion pathway to the selectivity filter. We use a combination of molecular modeling and functional assays to show that pH-sensing histidine residues and K⁺ ions mutually interact electrostatically in the confines of the extracellular ion pathway. K⁺ ions modulate the pK_a of sensing histidine side chains whose charge states in turn determine the open/closed transition of the channel pore. Cooperativity, and therefore steep dependence of TASK-3 K⁺ channel activity on extracellular pH, is dependent on an effect of the permeant ion on the channel pH_o sensors.     

The Compatibility of d-Pinitol and 1d-1-O-Methyl-Muco-Inositol with Malate Dehydrogenase Activity

Pinitol (1d -3-O-methyl-chiro-inositol) and 1d -1-O-methyl-muco-inositol, two cyclitols wide-spread in the plant kingdom, were isolated from plant sources in order to test their compatibility with malate dehydrogenase activity. Both compounds had no inhibitory effect on malate dehydrogenase from Rhizophora mangle in a range of 100 to 1000 mol . m^?3. Their influence on malate dehydrogenase activity from different plant sources (Rh. mangle L., Mesembryanthemum crystallinum L., Cicer arietinum L. and Spinacia oleracea L.) was also small and similar to that observed for a number of well established compatible solutes (e.g. proline, glycine betaine). A possible role of cyclitols as cryoprotectants or radical scavengers is discussed.     

Contribution of de novo protein synthesis to the hypertrophic effect of IGF-1 but not of thyroid hormones in adult ventricular cardiomyocytes

Background: Enhanced expression of IGF-1 occurs in left ventricular hypertrophy (LVH) associated with systemic hypertension. Cardiac dysfunction accompanied by LVH is also observed in hyperthyroidism. Objective: to assess the relative contributions of de novo protein synthesis and attenuated protein degradation to increased protein mass associated with cardiomyocyte hypertrophy elicited by IGF-1 and thyroid hormones (tri-iodo thyronine T₃, and l-thyroxine T₄), respectively. Methods: total mass of protein, and both the incorporation, and removal of previously incorporated l-U-¹⁴C-phenylalanine, indices of protein synthesis and degradation, respectively, were assessed in quiescent adult rat ventricular cardiomyocytes maintained in short-term culture, and corrected for DNA content, as a index of cell number. Results: IGF-1 (1 pM-100 nM) increased cell protein significantly, maximally at 1 nM and by 38% above basal value after 24 h. T₃ (10 pM-2 M) and T₄ (10 pM-2 M) increased cell protein significantly maximally at 1 M and by 33.2 and 30.5%, respectively, above basal value. IGF-1 ( 10 pM), T₃ (10 pM-2 M) and T₄ (10 pM-2 M) did not increase incorporation of l-U-₁₄C-phenylalanine above basal values. IGF-1 (100 pM-100 nM) increased incorporation of radiolabel significantly maximally at 100 nM and by 56%. T₄ (100 pM) and IGF-1 (10 pM), concentrations that did not stimulate de novo protein synthesis, attenuated the degradation of radiolabelled protein by 13.6 and 11.8%, respectively, compared to control values after 48 h. Conclusion: These data indicate that the acute hypertrophic response to (i) thyroid hormones cannot be attributed to initiation of de novo protein synthesis; (ii) IGF-1 comprises two components; the response elicited by IGF-1 (< 10 pM) is independent of, while the response elicited by IGF-1 (> 100 pM) is due to de novo protein synthesis.     

Variation of growth form parameters inCapsella (Cruciferae)

Growth form parameters ofCapsella bursa-pastoris populations, including a wide range of different environments, have been analyzed from random block field and growth chamber experiments. Changes in one character are often correlated with changes in another. Of special interest are correlations detected with the onset of first flowering. Variation in each of the characters is clearly influenced by both phenotypic and genotypic components. However, genotype — environment interactions are also subject to variation. Therefore, the adaptive significance of a given parameter is not found to be constant over the entire geographical range of the genus. Alpine populations tend to shift from annual to biannual life cycles.Part of a series Adaptation in life history traits of colonizing plant species. Part of a doctoral thesis by the first author. Dedicated to Prof. Dr.Karl Mägdefrau on the occasion of his 80th birthday.