Is microtubule disassembly a trigger for cold acclimation?

Cold acclimation was followed in three cultivars of winter wheat (Triticum aestivum L.) that differ in freezing tolerance, using root growth as the indicator. During acclimation (followed through 7 d at 4 degrees C), growth rate progressively recovered. The recovery was fast in the tolerant, slow in the sensitive cultivars. The development of freezing tolerance was followed by a challenging cold shock administered after various time intervals of acclimation. Acclimation proceeded faster in the tolerant cultivars. Microtubules were monitored during the acclimation period. A rapid, but transient partial disassembly in the tolerant cultivars preceded the formation of cold-stable microtubules and the recovery of growth rate. In contrast, this transient disassembly was absent in the sensitive cultivar. When a transient disassembly was artificially generated by a pulse-treatment with the antimicrotubular herbicide pronamide, this could induce freezing tolerance. The appearance of cold-stable microtubules was accompanied by a reduced abundance of type TUA1/2 alpha-tubulin isotypes. These findings are discussed with respect to a role of microtubule disassembly in the sensing of low-temperature stress.     

A new internal mode in F-actin helps explain the remarkable evolutionary conservation of actin's sequence and structure

Actin is one of the most highly conserved eukaryotic proteins. There are no amino acid changes between the chicken and human skeletal muscle isoforms, and the most dissimilar actins still share more than 85% sequence identity [1]. We suggest that large discrete internal modes of freedom within the actin filament may account for a significant component of this conservation, since each subunit must make multiple specific interactions with neighboring subunits. In support of this, we find that the same state of tilt of the actin subunit exists in both yeast and vertebrate striated muscle actin, and that in both the two domains undergo a "propeller rotation." A similar movement of domains has also been seen in hexokinase, Hsc70, and Arp2/3, all structural homologs of actin, suggesting that such an interdomain hinge motion is common to proteins in this superfamily. Subunit-subunit interactions within the actin filament involve sequence insertions that are not present in MreB, a bacterial homolog of actin. Remarkably, we find that in the tilted state actin subunits make new contacts with neighboring subunits that also involve these inserts, suggesting a key role for these elements in F-actin polymorphism.     

TA1/LAT-1/CD98 light chain and system L activity, but not 4F2/CD98 heavy chain, respond to arginine availability in rat hepatic cells. Loss Of response in tumor cells

Tumor associated gene-1/L amino acid transporter-1 (TA1/LAT-1) was recently identified as a light chain of the CD98 amino acid transporter and cellular activation marker. Our previous studies with primary rat hepatocyte cultures demonstrated that TA1 RNA levels were responsive to media amino acid concentrations, suggesting adaptive regulation. High level TA1 expression associated with transformed cells also suggested a role in tumor progression. The present study examined the relationship of TA1/CD98 expression, adaptive response, and associated amino acid transport to neoplastic transformation using a panel of well characterized rat hepatic cell lines. We found 1) increased expression of TA1 in response to amino acid depletion, specific for arginine but not glutamine; 2) loss of TA1 response to arginine in gamma-glutamyl transpeptidase-positive transformed and tumorigenic cells; 3) no appreciable response of 4F2/CD98 heavy chain to arginine levels; and 4) correlation of system L amino acid transport activity in response to arginine with changes in TA1/LAT-1 mRNA but not total immunoreacting protein. Our results suggest this CD98 light chain may act as an environmental sensor, responding to amino acid availability and that its regulation is complex. We hypothesize that altered TA1 expression is an early event in hepatocarcinogenesis giving neoplastic cells a growth or survival advantage, particularly under conditions of limited amino acid availability.     

Yeast (ADPribosyl)ation: revisiting a controversial question

The controversy about the occurrence of an (ADPribosyl)ating activity in yeast is still standing up. Here we discuss this topic on the basis of results obtained with classic experiments proposed over years as basis to characterize an (ADPribosyl)ation system in any organism. Independent results obtained in two different laboratories were in line with each other and went towards the occurrence of an active (ADPribosyl)ating system in Saccharomyces cerevisiae. In fact data collected from nuclear preparations of cultured cells matched those from baker's yeast and lyophilized yeast cells. Yeast (ADPribosyl)ating enzyme is a protein of 80-90 kDa, as determined by electrophoresis on polyacrylamide gel in sodium dodecyl sulphate, followed by immunoblotting with antibodies against anti-poly(ADPribose) polymerase catalytic site. It synthesizes products, that, after digestion with phosphodiesterase, co-migrates mainly with phosphoribosyl adenosine monophosphate after thin layer chromatography on silica gel plate.     

Primi Dati Sull'andamento Stagionale delle Temperature del Troneo di Olivo

Abstract

Temperatures recorded in a olive-tree stem in 1961–1962. — In the Botanical Garden of Bari a temperature recorder has been placed, bearing 6 electrothermometers with platinum resistences having a sensitivity of 2,4 mm/C[ddot], with a recording approximation of 0,1 C[ddot].

The olive tree under examination is a grown-up tree, which has been pruned according to the local methode, and it belongs to a local breed named « Ogliarola leccese ».

— Five thermometers have been employed as follows: 1^rst in the air, hanging from the stem, m. 1,30, high; 2^nd into the outer bark, cm, 0,5 deep; 3^rd into the cambium, cm. 1,5 deep; 4^th into the sap-wood, cm. 6 deep; 5^th into the hearth-wood, cm. 22 deep.

— The thermometers were all placed on the east side of the stem.

— The figures and the graphs of the paper show that:

a) The whole stem of the specimen under examination has a mean temperature which is higher than that of the surrounding air.

b) The outer bark gets warmer than the air, during the day-time; during the night it gives up heat outside and inside, namely towards the stem tissues, particularly to the bark-cambium-sapwood zone, and also as far as the hearth wood zone.

c) The whole living zone of the stem (bark-cambium-sapwood) generally keeps temperature values intermediate between those of the outer bark and those of the air. The temperature values vary along the year.

d) In spring the cambium is warmer than the outer bark.

e) In winter cambium and sapwood generally have the same temperature. The temperature values recorded in this season are much lower than those of the outer bark and almost the same of those of the air.

f) The behaviour of temperature in cambium and sapwood is probably ruled by biological factors.

g) The hearth-wood keeps generally warmer than the air and cooler than other outer tissues. Its thermic rhytm is almost reversed in comparison with that of the outer tissues.

Il seems that hearth wood acts as an useful heat collector and distributor towards the living outer tissues during the diurnal and seasonal temperature remissions, at least in mild climates as that of Bari.     

Searching algorithm for type IV secretion system effectors 1.0: a tool for predicting type IV effectors and exploring their genomic context

Type IV effectors (T4Es) are proteins produced by pathogenic bacteria to manipulate host cell gene expression and processes, divert the cell machinery for their own profit and circumvent the immune responses. T4Es have been characterized for some bacteria but many remain to be discovered. To help biologists identify putative T4Es from the complete genome of α- and γ-proteobacteria, we developed a Perl-based command line bioinformatics tool called S4TE (searching algorithm for type-IV secretion system effectors). The tool predicts and ranks T4E candidates by using a combination of 13 sequence characteristics, including homology to known effectors, homology to eukaryotic domains, presence of subcellular localization signals or secretion signals, etc. S4TE software is modular, and specific motif searches are run independently before ultimate combination of the outputs to generate a score and sort the strongest T4Es candidates. The user keeps the possibility to adjust various searching parameters such as the weight of each module, the selection threshold or the input databases. The algorithm also provides a GC% and local gene density analysis, which strengthen the selection of T4E candidates. S4TE is a unique predicting tool for T4Es, finding its utility upstream from experimental biology.     

The role of fruit drinks in daily diet of some Osijek inhabitants, Croatia

The aim of the study was to assess the role of fruit drinks in daily diet of Osijek inhabitants. A special questionnaire was administered to 199 patients visiting a family physician's office during December 2003. The concentration of vitamin C (L-ascorbic acid) was determined in 42 commercially available fruit drinks. Only 11% (22/199) of study subjects did not take fruit drinks. The mean concentration of vitamin C in all samples was 150.10 +/- 166.83 mg/L. The questionnaire revealed that 89% (177/199) of study subjects using fruit drinks were taking a mean of 0.4 L of fruit drink per day, yielding a mean of 60.04 mg of vitamin C, i.e. 100% of the recommended daily allowance of 60 mg. Study results indicated fruit drinks to be a significant source of vitamin C in daily diet, however, great variation in vitamin C intake according to socioeconomic status of study subjects and type of fruit drink should be noted.     

Synaptic Synthesis,Dephosphorylation, and Degradation: A NOVEL PARADIGM FOR AN ACTIVITY-DEPENDENT NEURONAL CONTROL OF CDKL5*

Mutations in the X-linked CDKL5 (cyclin-dependent kinase-like 5) gene have been associated with several forms of neurodevelopmental disorders, including atypical Rett syndrome, autism spectrum disorders, and early infantile epileptic encephalopathy. Accordingly, loss of CDKL5 in mice results in autistic-like features and impaired neuronal communication. Although the biological functions of CDKL5 remain largely unknown, recent pieces of evidence suggest that CDKL5 is involved in neuronal plasticity. Herein, we show that, at all stages of development, neuronal depolarization induces a rapid increase in CDKL5 levels, mostly mediated by extrasomatic synthesis. In young neurons, this induction is prolonged, whereas in more mature neurons, NMDA receptor stimulation induces a protein phosphatase 1-dependent dephosphorylation of CDKL5 that is mandatory for its proteasome-dependent degradation. As a corollary, neuronal activity leads to a prolonged induction of CDKL5 levels in immature neurons but to a short lasting increase of the kinase in mature neurons. Recent results demonstrate that many genes associated with autism spectrum disorders are crucial components of the activity-dependent signaling networks regulating the composition, shape, and strength of the synapse. Thus, we speculate that CDKL5 deficiency disrupts activity-dependent signaling and the consequent synapse development, maturation, and refinement.     

Heterologous expression and solution structure of defensin from lentil Lens culinaris

A new defensin Lc-def, isolated from germinated seeds of the lentil Lens culinaris, has molecular mass 5440.4 Da and consists of 47 amino acid residues. Lc-def and its ¹⁵N-labeled analog were overexpressed in Escherichia coli. Antimicrobial activity of the recombinant protein was examined, and its spatial structure, dynamics, and interaction with lipid vesicles were studied by NMR spectroscopy. It was shown that Lc-def is active against fungi, but does not inhibit the growth of Gram-positive and Gram-negative bacteria. The peptide is monomeric in aqueous solution and contains one α-helix and triple-stranded β-sheet, which form cysteine-stabilized αβ motif (CSαβ) previously found in other plant defensins. The sterically neighboring loop1 and loop3 protrude from the defensin core and demonstrate significant mobility on the μs–ms timescale. Lc-def does not bind to the zwitterionic lipid (POPC) vesicles but interacts with the partially anionic (POPC/DOPG, 7:3) membranes under low-salt conditions. The Lc-def antifungal activity might be mediated through electrostatic interaction with anionic lipid components of fungal membranes.     

Direct interaction of actin filaments with F‐BAR protein pacsin2

Two mechanisms have emerged as major regulators of membrane shape: BAR domain‐containing proteins, which induce invaginations and protrusions, and nuclear promoting factors, which cause generation of branched actin filaments that exert mechanical forces on membranes. While a large body of information exists on interactions of BAR proteins with membranes and regulatory proteins of the cytoskeleton, little is known about connections between these two processes. Here, we show that the F‐BAR domain protein pacsin2 is able to associate with actin filaments using the same concave surface employed to bind to membranes, while some other tested N‐BAR and F‐BAR proteins (endophilin, CIP4 and FCHO2) do not associate with actin. This finding reveals a new level of complexity in membrane remodeling processes.