A Medicago truncatula NADPH oxidase is involved in symbiotic nodule functioning

The plant plasma membrane-localized NADPH oxidases, known as respiratory burst oxidase homologues (RBOHs), appear to play crucial roles in plant growth and development. They are involved in important processes, such as root hair growth, plant defence reactions and abscisic acid signalling. Using sequence similarity searches, we identified seven putative RBOH-encoding genes in the Medicago truncatula genome. A phylogenetic reconstruction showed that Rboh gene duplications occurred in legume species. We analysed the expression of these MtRboh genes in different M. truncatula tissues: one of them, MtRbohA, was significantly up-regulated in Sinorhizobium meliloti-induced symbiotic nodules. MtRbohA expression appeared to be restricted to the nitrogen-fixing zone of the functional nodule. Moreover, using S. meliloti bacA and nifH mutants unable to form efficient nodules, a strong link between nodule nitrogen fixation and MtRbohA up-regulation was shown. MtRbohA expression was largely enhanced under hypoxic conditions. Specific RNA interference for MtRbohA provoked a decrease in the nodule nitrogen fixation activity and the modulation of genes encoding the microsymbiont nitrogenase. These results suggest that hypoxia, prevailing in the nodule-fixing zone, may drive the stimulation of MtRbohA expression, which would, in turn, lead to the regulation of nodule functioning.     

Improvement of liver cancer detection with simultaneous assessment of circulating levels of free alpha-fetoprotein (AFP) and AFP-IgM complexes

We assessed the presence of alpha-fetoprotein (AFP) complexed with IgM (AFP-IgM IC) in serum of patients affected by hepatocellular carcinoma (HCC), cirrhosis and chronic hepatitis as well as in healthy subjects by means of a dedicated ELISA assay. The amount of AFP-IgM IC was expressed in arbitrary units (AU) on a reference standard curve. Free AFP (FAFP) levels were determined in parallel in each sample by means of an automated immunoassay system. The mean serum concentration of AFP-IgM IC was significantly higher in HCC patients (mean +/- SD: 1378.3 +/- 2935.7 AU/mL) than in cirrhotic patients (129.8 +/- 261.4 AU/mL) and in patients with chronic hepatitis (80.9 +/- 168.9 AU/mL) (p < 0.01). HCC patients had FAFP values above the 20 ng/mL cutoff in 44% of cases (22/50) and AFP-IgM IC values above the 120 AU/mL cutoff in 60% of cases (30/50). The occurrence of the free and IgM-complexed form of circulating AFP did not overlap, and 82% of patients (41/50) were positive for at least one marker. The results indicate that AFP-IgM IC is a complementary serological marker to FAFP and that the combination of these biomarkers may be useful in the diagnosis of liver cancer.     

Caveolin-stabilized membrane domains as multifunctional transport and sorting devices in endocytic membrane traffic

Endocytosis comprises several routes of internalization. An outstanding question is whether the caveolar and endosomal pathways intersect. Following transport of the caveolar protein Caveolin-1 and two cargo complexes, Simian Virus 40 and Cholera toxin, in live cells, we uncovered a Rab5-dependent pathway in which caveolar vesicles are targeted to early endosomes and form distinct and stable membrane domains. In endosomes, the low pH selectively allowed the toxin to diffuse out of the caveolar domains into the surrounding membrane, while the virus remained trapped. Thus, we conclude that, unlike cyclic assembly and disassembly of coat proteins in vesicular transport, oligomeric complexes of caveolin-1 confer permanent structural stability to caveolar vesicles that transiently interact with endosomes to form subdomains and release cargo selectively by compartment-specific cues.     

Identification of novel chromosomal regions associated with airway hyperresponsiveness in recombinant congenic strains of mice

Airway responsiveness is the ability of the airways to respond to bronchoconstricting stimuli by reducing their diameter. Airway hyperresponsiveness has been associated with asthma susceptibility in both humans and murine models, and it has been shown to be a complex and heritable trait. In particular, the A/J mouse strain is known to have hyperresponsive airways, while the C57BL/6 strain is known to be relatively refractory to bronchoconstricting stimuli. We analyzed recombinant congenic strains (RCS) of mice generated from these hyper- and hyporesponsive parental strains to identify genetic loci underlying the trait of airway responsiveness in response to methacholine as assessed by whole-body plethysmography. Our screen identified 16 chromosomal regions significantly associated with airway hyperresponsiveness (genome-wide P ≤ 0.05): 8 are supported by independent and previously published reports while 8 are entirely novel. Regions that overlap with previous reports include two regions on chromosome 2, three on chromosome 6, one on chromosome 15, and two on chromosome 17. The 8 novel regions are located on chromosome 1 (92–100 cM), chromosome 5 (>73 cM), chromosome 7 (>63 cM), chromosome 8 (52–67 cM), chromosome 10 (3–7 cM and >68 cM), and chromosome 12 (25–38 cM and >52 cM). Our data identify several likely candidate genes from the 16 regions, including Ddr2, Hc, Fbn1, Flt3, Utrn, Enpp2, and Tsc.     

Photoinduced electron transfer and chemical alpha-deoxygenation of D-galactono-1,4-lactone. Synthesis of 2-deoxy-D-lyxo-hexofuranosides

Two simple procedures for the synthesis of 2-deoxy-D-lyxo-hexono-1,4-lactone are described. Reductive cleavage of a 2-O-tosyl derivative of D-galactono-1,4-lactone in the presence of sodium iodide afforded the 2-deoxy derivative. On the other hand, alpha-deoxygenation of D-galactono-1,4-lactone was easily achieved by photochemical electron transfer deoxygenation of HO-2 as the 3-(trifluoromethyl)benzoate. Methyl 2-deoxy-beta-D-lyxo-hexafuranoside ('methyl 2-deoxy-beta-D-galactofuranoside') was synthesized and tested as substrate for exo beta-D-galactofuranosidase from Penicillium fellutanum. The reaction was followed by HPAEC, showing that methyl 2-deoxy-beta-D-galactofuranoside was not hydrolyzed by incubation with the enzyme. Neither the 2-deoxy lactone, nor the 2-deoxy-beta-D-galactofuranoside acted as inhibitors of the reaction with the 4-nitrophenyl beta-D-galactofuranoside. The present and our previous results show that the hydroxyl groups at C-2, C-3 and C-6 of the galactofuranoside are essential for interaction with the exo beta-D-galactofuranosidase.     

Effect of deamidation on folding of ribonuclease A

The folding of ribonuclease A (RNase A) has been extensively studied by characterizing the disulfide containing intermediates using different experimental conditions and analytical techniques. So far, some aspects still remain unclear such as the role of the loop 65-72 in the folding pathway. We have studied the oxidative folding of a RNase A derivative containing at position 67 the substitution Asn --> isoAsp where the local structure of the loop 65-72 has been modified keeping intact the C65-C72 disulfide bond. By comparing the folding behavior of this mutant to that of the wild-type protein, we found that the deamidation significantly decreases the folding rate and alters the folding pathway of RNase A. Results presented here shed light on the role of the 65-72 region in the folding process of RNase A and also clarifies the effect of the deamidation on the folding/unfolding processes. On a more general ground, this study represents the first characterization of the intermediates produced along the folding of a deamidated protein.     

Summer and spring trophic niche of larval and juvenile <Emphasis Type="Italic">Pleuragramma antarcticum</Emphasis> in the Western Ross Sea,Antarctica

The aim of this paper is to contribute to the knowledge on the feeding habits of larvae and juvenile Pleuragramma antarcticum in the western Ross Sea. In summer, the diet of P. antarcticum postlarvae (8–17 mm) was dominated by calanoid eggs (35.5%), Limacina (32.1%) and tintinnids (17.6%), while the principal food of juveniles consisted mainly of copepods (98.2%), with Oncaea curvata being the most abundant (85.1%) and the most frequently consumed prey. The food composition of P. antarcticum postlarvae (24–29 mm), collected in spring, suggest that they fed actively under the sea ice. Stephos longipes, Harpacticus furcifer and Paralabidocera antarctica sea ice copepods represent, in all their different developmental stages, the most abundant biomass food in Terra Nova Bay in this period. Our results therefore suggest that the diet of younger Pleuragramma specimens shifted in prey composition from the first summer to the following spring. This study draws attention to the key role of the copepod, P. antarctica, in the food web of Terra Nova Bay. This article belongs to a special topic: Five articles coordinated by L. Guglielmo and V. Saggiomo appear in this issue of Polar Biology and are a result of a workshop on Sea-ice communities in Terra Nova Bay (Ross Sea) held in August 2007 in Capo Calavà, Messina, Italy.