ABCB1 and ABCB19 auxin transporters have synergistic effects on early and late Arabidopsis anther development

Arabidopsis abcb1 abcb19 double mutants defective in the auxin transporters ABCB1/PGP1 and ABCB19/PGP19 are altered in stamen elongation, anther dehiscence and pollen maturation. To assess the contribution of these transporters to stamen development we performed phenotypic, histological analyses, and in situ hybridizations on abcb1 and abcb19 single mutant fl owers. We found that pollen maturation and anther dehiscence are precocious in the abcb1 but not in the abcb19 mutant. Accordingly, endothecium ligni fication is altered only in abcb1 anthers. Both abcb1 and abcb1 abcb19 stamens also show altered early development, with asynchronous anther locules and a multilayer tapetum. DAPI staining showed that the timing of meiosis is asynchronous in abcb1 abcb19 anther locules, while only a small percentage of pollen grains are nonviable according to Alexander's staining. In agreement, TAM(TARDY ASYNCHRONOUS MEIOSIS), as well as BAM2(BARELY ANY MERISTEM)—involved in tapetal cell development—are overexpressed in abcb1 abcb19 young fl ower buds. Corre spondingly, ABCB1 and ABCB19 mRNA localization supports the observed phenotypes of abcb1 and abcb1 abcb19 mutant anthers. In conclusion, we provide evidence that auxin transport plays a signi ficant role both in early and late stamen development: ABCB1 plays a major role during anther development, while ABCB19 has a synergistic role.     

Cytology of Dicranostigma and Glaucium (Papaveraceae) from China

The cytological studies were performed in 11 populations of Dicranostigma lactucoides, Dleptopodum, and Derectum and one population for Galucium fimbrilligerum. All three species of Dicranostigma possessed chromosome number 2n=2x=12, of which chromosome numbers for Derectum and Dlactucoides are reported here for the first time. Glaucium fimbrilligerum presented the number 2n=2x=10, which is not only new to the genus but also the known lowest number for Glaucium.     

Assessment of new 6-Cl-HOBt based coupling reagents for peptide synthesis. Part 1: Coupling efficiency study

The efficiency of a series of well-known coupling reagents (TBTU, HATU, and PyBOP) and of new in situ activating reagents (TCTU, HCTU, and DMTMM) was compared by synthesizing the 65–74 fragment of the Acyl Carrier Protein (H-Val-Gln-Ala-Ala-Ile-Asp-Tyr-Ile-Asn-Gly-NH₂₎, containing `a difficult sequence', as a test peptide, in a multiple peptide synthesizer. The longer sequence rMOG(35–55) was also synthesized. It was clear that the aminium salts are more efficient than the phosphonium salt (PyBOP) and that the new 6Cl-HOBt based reagents (HCTU and particularly TCTU) are very efficient, while DMTMM appeared to be not suitable for SPPS.     

Early post‐fire succession in northwestern Patagonia grasslands

Abstract. Large‐scale disturbances, notably fire and grazing, structure grass and shrubland dynamics in semi‐arid environments. We studied early post‐fire succession in two burned grasslands, one unburned grassland, and one shrubland near the burned area. We observed three processes: (1) establishment of a ‘phantom’ community comprised of fugitive species. Although transient, these species increase diversity and recharge the seed bank before the next disturbance; (2) regeneration of the original community by persistence of resprouter species and by auto‐replacement; (3) early stages of invasion by seedlings of the shrub Fabiana imbricata, which germinate next to shrubland and create new F. imbricata patches. Weed invasion was principally due to the ruderal exotic species Verbascum thapsus from the nearby road verge and by rapid increase of Rumex acetosella cover, another exotic species present before the fire. Although post‐fire climatic conditions are particularly important in semi‐arid environments, succession depends greatly on the regeneration strategies and dispersal abilities of the species present in the burned area. The phantom community occurs only at the first stage of succession when there is little competition for resources. We could call this process ‘the race for occupation of the area’. The second stage, when competition for resources becomes progressively more important, could be called ‘the effort to maintain space’.     

Siglec-9 defines and restrains a natural killer subpopulation highly cytotoxic to HIV-infected cells

Siglec-9 is an MHC-independent inhibitory receptor expressed on a subset of natural killer (NK) cells. Siglec-9 restrains NK cytotoxicity by binding to sialoglycans (sialic acid-containing glycans) on target cells. Despite the importance of Siglec-9 interactions in tumor immune evasion, their role as an immune evasion mechanism during HIV infection has not been investigated. Using in vivo phenotypic analyses, we found that Siglec-9⁺ CD56^dim NK cells, during HIV infection, exhibit an activated phenotype with higher expression of activating receptors and markers (NKp30, CD38, CD16, DNAM-1, perforin) and lower expression of the inhibitory receptor NKG2A, compared to Siglec-9^- CD56^dim NK cells. We also found that levels of Siglec-9⁺ CD56^dim NK cells inversely correlate with viral load during viremic infection and CD4⁺ T cell-associated HIV DNA during suppressed infection. Using in vitro cytotoxicity assays, we confirmed that Siglec-9⁺ NK cells exhibit higher cytotoxicity towards HIV-infected cells compared to Siglec-9^- NK cells. These data are consistent with the notion that Siglec-9⁺ NK cells are highly cytotoxic against HIV-infected cells. However, blocking Siglec-9 enhanced NK cells’ ability to lyse HIV-infected cells, consistent with the known inhibitory function of the Siglec-9 molecule. Together, these data support a model in which the Siglec-9⁺ CD56^dim NK subpopulation is highly cytotoxic against HIV-infected cells even whilst being restrained by the inhibitory effects of Siglec-9. To harness the cytotoxic capacity of the Siglec-9⁺ NK subpopulation, which is dampened by Siglec-9, we developed a proof-of-concept approach to selectively disrupt Siglec/sialoglycan interactions between NK and HIV-infected cells. We achieved this goal by conjugating Sialidase to several HIV broadly neutralizing antibodies. These conjugates selectively desialylated HIV-infected cells and enhanced NK cells’ capacity to kill them. In summary, we identified a novel, glycan-based interaction that may contribute to HIV-infected cells’ ability to evade NK immunosurveillance and developed an approach to break this interaction.     

Neutralizing Activity Induced by the Attenuated Coxsackievirus B3 Sabin3-like Strain Against CVB3 Infection

Coxsackievirus B3 (CVB3) causes viral myocarditis, and can ultimately result in dilated cardiomyopathy. There is no vaccine available for clinical use. In the present work, we assessed whether the Sabin3-like mutant of CVB3 could induce a protective immunity against virulent CVB3 Nancy and CVB4 E2 strains in mice by both oral and intraperitoneal (IP) routes. Serum samples, taken from mice inoculated with Sabin3-like, were assayed in vitro for their anti-CVB3 neutralizing activity. CVB3 Sabin3-like was highly attenuated in vivo and was able to induce an anti-CVB3 activity of the serum. However, at 4 days post-CVB3 challenge, significant increased titers of CVB3 neutralizing antibodies were detectable in the sera of immunized mice over the next 6 days. Non-immunized mice challenged with CVB3 Nancy had no anti-CVB3 activity in their sera until 10 days post-infection. CVB3 Nancy induced higher viral titers than did the mutant strain. There was no variation of the neutralizing activity of serum taken from mice immunized with CVB3 Sabin3-like and challenged with CVB4 E2, compared to non-immunized mice. Despite the fact that CVB3 and CVB4 are closely related viruses, virus-neutralizing activity clearly distinguish between these viruses. A variable and limited amount of pancreatic inflammation was seen in some mice 10 days after Sabin3-like inoculation by IP route, whereas there was no evidence of pancreatic damage in mice inoculated by oral route. All immunized mice were protected from myocarditis and pancreatitis at 8 days post-challenge with CVB3 or CVB4 E2. These findings strongly suggest that the mutant strain could be considered a candidate for an attenuated CVB3 vaccine.     

Solution conformation of a pectin fragment disaccharide using molecular modelling and nuclear magnetic resonance

In the present study, the conformational behaviour of methylated pectic disaccharide 4-O-alpha-D-galactopyranurosyl 1-O-methyl-alpha-D-galactopyranuronic 6,6'-dimethyl diester 1 has been completely characterized through combined n.m.r. and molecular modelling studies. The 1H-1H n.O.e. across the glycosidic bond was measured by both steady-state and transient 1D and 2D experiments. In parallel, the complete conformational analysis of the disaccharide has been achieved with the MM3 molecular mechanics method. The conformation of the pyranose ring is confirmed by the excellent agreement between the experimental and calculated intracyclic scalar coupling constants. The iso-energy contours displayed on the 'relaxed' map indicate an important flexibility about the glycosidic linkage. There is no significant influence of the methoxyl group on the conformational behaviour of the disaccharide. The theoretical n.m.r. data were calculated taking into account all the accessible conformations and using the averaging methods appropriate for slow internal motions. 3JC-H coupling constants were calculated using an equation suitable for C-O-C-H segments. The agreement between experimental and theoretical data is excellent. Within the potential energy surface calculated for the disaccharide, several conformers can be identified. When these conformations are extrapolated to a regular polymer structure, they generate pectins with right- and left-handed chirality along with a two-fold helix. These different types of helical structure are the result of small changes in conformation, without any drastic variation of the fibre repeat.