Molecular phylogeny and timing of evolution of Anthomyza and related genera (Diptera: Anthomyzidae)

Phylogenetic hypotheses of the relationships of Diptera: Anthomyzidae (61 taxa) are discussed with special reference to the genera Fungomyza Rohá?ek, 1999, Anthomyza Fallén, 1810, Epischnomyia Rohá?ek, 2006 and Arganthomyza Rohá?ek, 2009 based on the analysis of 7 combined mitochondrial + nuclear gene markers in comparison with results of the most recent cladistic analysis of morphological characters. The majority of revealed inter‐ and intrageneric relationships of these genera in both analyses were largely congruent except for the topology of Fungomyza and Arganthomyza being equivocal because of different topologies in phylograms generated by alternative molecular methods and Epischnomyia forming a branch within Anthomyza in the molecular data hypothesis. The formerly unsettled Anthomyza drachma proved to be the sister species of the A. umbrosa group while the affinity of A. flavosterna to the A. bellatrix group has not been confirmed. The first phylogenetic hypothesis of Anthomyzinae to include timing of the nodes of divergence is presented. The origin of the subfamily is dated into the Eocene, ca 37.5 (30.6 – 45.3) MYA, and agrees with the age of the oldest known fossils from Baltic amber. The analysed members of the Anthomyza group of genera were found to have already evolved in the upper Oligocene. The divergences of the Palaearctic and Nearctic sister species in Arganthomyza and Anthomyza were found to occur at several different times during the Neogene (upper Miocene) to lower Quaternary (Pleistocene), from about 7 to 0.7 MYA. These were likely the result of fragmentation of widespread (Holarctic or Sino‐Japanese–Nearctic) ranges of ancestral taxa by vicariance events that were, in turn, caused by multiple interruptions of Beringia Land Bridges or cooling of climate and seem to be consistent with the times of multiple disjunctions of the flora.     

A TOR (target of rapamycin) and nutritional phosphoproteome of fission yeast reveals novel targets in networks conserved in humans

Fluctuations in TOR, AMPK and MAP-kinase signalling maintain cellular homeostasis and coordinate growth and division with environmental context. We have applied quantitative, SILAC mass spectrometry to map TOR and nutrient-controlled signalling in the fission yeast Schizosaccharomyces pombe. Phosphorylation levels at more than 1000 sites were altered following nitrogen stress or Torin1 inhibition of the TORC1 and TORC2 networks that comprise TOR signalling. One hundred and thirty of these sites were regulated by both perturbations, and the majority of these (119) new targets have not previously been linked to either nutritional or TOR control in either yeasts or humans. Elimination of AMPK inhibition of TORC1, by removal of AMPKα (ssp2::ura4⁺), identified phosphosites where nitrogen stress-induced changes were independent of TOR control. Using a yeast strain with an ATP analogue-sensitized Cdc2 kinase, we excluded sites that were changed as an indirect consequence of mitotic control modulation by nitrogen stress or TOR signalling. Nutritional control of gene expression was reflected in multiple targets in RNA metabolism, while significant modulation of actin cytoskeletal components points to adaptations in morphogenesis and cell integrity networks. Reduced phosphorylation of the MAPKK Byr1, at a site whose human equivalent controls docking between MEK and ERK, prevented sexual differentiation when resources were sparse but not eliminated.     

Confirmation of the presence of a Cu(II)/topa quinone active site in the amine oxidase from fenugreek seedlings

Amine oxidase from etiolated seedlings of fenugreek (Trigonellafoenum—graecum) has been isolated by a purification procedureinvolving three chromato—graphic steps. The homogeneousenzyme is of pink colour with a visible absorption maximum at500 nm. The dimeric enzyme (2 75 kDa) is a slightly acidicprotein (pl 6.8) containing 8% neutral sugars. N—ter—minalamino acid sequence of the enzyme shows a high degree of similarityto other plant and microbial copper—containing amine oxidases.The best substrates of the enzyme are aliphatic diamines andsome polyamines, whereas inhibitors are substrate analogues,copper complexing agents, some alkaloids and several other compounds.Spectrophotometric titra—tions with phenylhydrazines demonstratedone reactive carbonyl group per subunit of the enzyme and redox—cyclicquinone staining after native electrophor—esis indicatedthe presence of a quinone cofactor. Differential pulse polarographyshowed the existence of a copper/quinone—containing activesite. The resonance Raman spectroscopy and the pH—dependentshift of the absorption spectrum of the enzyme p—nitrophenylhydrazoneconfirm unambiguously the identity of the cofactor with topaquinone. EPR spectra of the enzyme are in accordance with thoseof tetragonal cupric complexes as known for other copper—containingamine oxidases. Besides the copper, Mn(II)ions were detectedthat partially occupy another metal site in the enzyme, buttheir catalytical importance is unlikely. Key words: Fenugreek, Trigonella foenum—graceum, amine oxidase, topa quinone