Tyrosine Hydroxylation in Betalain Pigment Biosynthesis Is Performed by Cytochrome P450 Enzymes in Beets (Beta vulgaris)

Yellow and red-violet betalain plant pigments are restricted to several families in the order Caryophyllales, where betacyanins play analogous biological roles to anthocyanins. The initial step in betalain biosynthesis is the hydroxylation of tyrosine to form L-DOPA. Using gene expression experiments in beets, yeast, and Arabidopsis, along with HPLC/MS analysis, the present study shows that two novel cytochrome P450 (CYP450) enzymes, CYP76AD6 and CYP76AD5, and the previously described CYP76AD1 can perform this initial step. Co-expressing these CYP450s with DOPA 4,5-dioxygenase in yeast, and overexpression of these CYP450s in yellow beets show that CYP76AD1 efficiently uses L-DOPA leading to red betacyanins while CYP76AD6 and CYP76AD5 lack this activity. Furthermore, CYP76AD1 can complement yellow beetroots to red while CYP76AD6 and CYP76AD5 cannot. Therefore CYP76AD1 uniquely performs the beet R locus function and beets appear to be genetically redundant for tyrosine hydroxylation. These new functional data and ancestral character state reconstructions indicate that tyrosine hydroxylation alone was the most likely ancestral function of the CYP76AD alpha and beta groups and the ability to convert L-DOPA to cyclo-DOPA evolved later in the alpha group.     

Collagen type I fibril packing in vivo and in vitro

Reviewed are works concerning the mechanisms of collagen (type I) fibril packing and the influence of macromolecular structure and physicochemical parameters of the medium on the process.     

Temperature as a Factor of Niche Separation in Free-Living Mesostigmatid Mites (Mesostigmata: Arachida, Parasitiformes) of Storm Detritus

We carried out a laboratory investigation of temperature effect on survival as well as reproductive and trophic activities of mesostigmatid mites. Representatives of gamasid (three species) and uropodid (two species) mites abundant in storm detritus were used as model species. The upper reproduction limit and the upper survival limit were determined for the mites and their preys in the thermal range from 29 to 47°C. Most of studied species managed to survive and propagate in a wide thermal range corresponding to the thermal gradient in the peripheral zones of algal belts. The role of temperature as a factor of ecological niche separation in saprophytic Mesostigmata and formation of polydominant mite communities in the algal belts were demonstrated.     

Neurogenic cholinergic dilation observed in isolated vessels of the skeletal muscles of cats (the caudal femoral artery)

Neuromediated behaviour of feline caudal femoral arteries was studied in vitro. Transmural stimulation of arteries, previously constricted by noradrenaline, caused an up to 58% relaxation of the induced tone. Atropine (10(-5) g/ml) blocked this response by 93%. The removal of endothelium did not affect the magnitude of the response. It is suggested that the caudal femoral artery is an example of a blood vessel which has muscarinic receptors located directly on its smooth muscles.     

Slow Cortical Potentials Preceeding Visually Guided Saccades in Schizophrenics

The parameters of saccades and presaccadic slow potentials were studied in right-handed men with a dominant right eye, including 19 schizophrenics and 12 healthy subjects. For visual stimulation, three light-emitting diodes were used, which were located in the center of the visual field (the central fixation stimulus) and 10° to the right and left of it (peripheral stimuli). Two stimulation protocols were used: with a simultaneous switching off of the central fixation stimulus and switching on of the peripheral stimuli (test 1) and with an interstimulus gap of 200 ms (test 2). According to the latency, saccades were divided into anticipatory, express, and regular. Slow EEG potentials preceding regular saccades were analyzed. It was found that the proportion of anticipatory saccades is considerably higher than the normal value in schizophrenia. The analysis of the presaccadic potentials demonstrated a significant decrease in the amplitude of negative potentials in the vertex region at early stages of presaccadic preparation and its increase in the occipital region at late stages. Test 2 in the patients demonstrated an increase in the positivity focus in the frontal region of the right hemisphere. It was assumed that the alterations found in schizophrenia result from the deficit of frontal cortical fields.     

Reconstructing labroid evolution with single-copy nuclear DNA.

Fifteen per cent of all living fishes are united in a single suborder (Labroidei) and display a dazzling array of behavioural and ecological traits. The labroids are considered monophyletic and members share a pharyngeal jaw apparatus (PJA) modified for crushing and processing prey. Outside of the explicitly functional PJA, there is no corroborative evidence for a monophyletic Labroidei. Here, we report the first molecular phylogenetic analysis of the suborder. Contrary to morphology-based phylogenies, our single-copy nuclear DNA data do not support labroid families as a natural group. Our data indicate that pharyngognathy has evolved independently among labroid families and that characters of the PJA are not reliable markers of perciform evolution. This work ''crushes'' conventional views of fish phylogeny and should engender novel concepts of piscine life history evolution.     

HPLC photofingerprinting of conformational peculiarities and transitions in oligonucleotide duplexes.

Two self-complementary sequence-isomeric decadeoxyribonucleotides were exposed to UV light under conditions in which they assume duplex structures. After that they were analyzed in the denatured state by reversed-phase high-performance liquid chromatography (HPLC). Characterization of the separated photoproducts allowed localization of cyclobutane pyrimidine dimers in the sequences of the modified oligonucleotides. For [d(GGAAATTTCC)]2, which is known to contain in its central part a stretch of rigid B'-conformation with decreased mobility of constituent bases, lower yields of thymine dimers, as compared with that for ordinary B-form [d(CCTTTAAAGG)]2, were found. On the contrary, mixed thymine-cytosine heterodimers generated in the former oligonucleotide demonstrate the increase in photoreactivity of these residues at the B'-B junction. This is probably due to the peculiar conformation adopted by this decanucleotide. Stimulation of B'-B transition, by increasing the temperature before melting, reduced an inhibition of thymine photodimer formation. During the melting of both oligonucleotides yields of all identified photoinduced cyclobutadipyrimidines were reduced. Possible influences of some metal cations on the stability of the B'-form were also studied by this photoprobing technique. The present study demonstrates the feasibility of HPLC photofingerprinting as a new approach for structural analysis of nucleic acids.     

Application of a new slot comb to electrophoretic analysis.

In the present experiment, a new slot comb was designed in order to form a wide and sloped sample well on the stacking gel of electrophoresis. Using this slot comb, a gradient of the reagent layer of interest can be easily formed transversely on the gel that is perpendicular to the direction of electrophoresis. Thus, the protein sample overlaid on the agent migrates across the gradient layer during electrophoresis to produce a continuous electrophoretic band reflecting the interaction between the protein and reagent. This new slot comb (tentatively called slope comb) was applied to the following two experiments. In the first experiment, in combination with this comb and a reducing agent, 2-mercaptoethanol, the reducing steps of cross-linked axonemal proteins with o-iodosobenzoic acid (OIBA) were analyzed electrophoretically, enabling visualization of the reducing pattern of each axonemal protein in a single experiment. The results obtained indicate that alpha and beta tubulins are cross-linked differently by OIBA. In the second experiment, the formation of the Ca2+ gradient layer using this slope comb could electrophoretically differentiate the Ca2+ sensitivity of three Ca(2+)-binding proteins.