Nodule Organogenesis and Symbiotic Mutants of the Model Legume \u3ci\u3eLotus japonicus\u3c/i\u3e

A detailed microscopical analysis of the morphological features that distinguish different developmental stages of nodule organogenesis in wild-type Lotus japonicus ecotype Gifu B-129-S9 plants was performed, to provide the necessary framework for the evaluation of altered phenotypes of L. japonicus symbiotic mutants. Subsequently, chemical ethyl methanesulfonate (EMS) mutagenesis of L. japonicus was carried out. The analysis of approximately 3,000 M1 plants and their progeny yielded 20 stable L. japonicus symbiotic variants, consisting of at least 14 different symbiosis- associated loci or complementation groups. Moreover, a mutation affecting L. japonicus root development was identified that also conferred a hypernodulation response when a line carrying the corresponding allele (LjEMS102) was inoculated with rhizobia. The phenotype of the LjEMS102 line was characterized by the presence of nodule structures covering almost the entire root length (Nod++), and by a concomitant inhibition of both root and stem growth. A mutation in a single nuclear gene was shown to be responsible for both root and symbiotic phenotypes observed in the L. japonicus LjEMS102 line, suggesting that (a) common mechanism(s) regulating root development and nodule formation exists in legumes

Loss of chloroplast protease SPPA function alters high light acclimation processes in Arabidopsis thaliana L. (Heynh.)

SPPA1 is a protease in the plastids of plants, located in non-appressed thylakoid regions. In this study, T-DNA insertion mutants of the single-copy SPPA1 gene in Arabidopsis thaliana (At1g73990) were examined. Mutation of SPPA1 had no effect on the growth and development of plants under moderate, non-stressful conditions. It also did not affect the quantum efficiency of photosynthesis as measured by dark-adapted Fv/Fm and light-adapted ΦPSII. Chloroplasts from sppA mutants were indistinguishable from the wild type. Loss of SPPA appears to affect photoprotective mechanisms during high light acclimation: mutant plants maintained a higher level of non-photochemical quenching of Photosystem II chlorophyll (NPQ) than the wild type, while wild-type plants accumulated more anthocyanin than the mutants. The quantum efficiency of Photosystem II was the same in all genotypes grown under low light, but was higher in wild type than mutants during high light acclimation. Further, the mutants retained the stress-related Early Light Inducible Protein (ELIP) longer than wild-type leaves during the early recovery period after acute high light plus cold treatment. These results suggest that SPPA1 may function during high light acclimation in the plastid, but is non-essential for growth and development under non-stress conditions

A novel type of macrothrombocytopenia associated with a defect in alpha2,3-sialylation

Item does not contain fulltextWe describe a novel type of human thrombocytopenia characterized by the appearance of giant platelets and variable neutropenia. Searching for the molecular defect, we found that neutrophils had strongly reduced sialyl-Lewis X and increased Lewis X surface expression, pointing to a deficiency in sialylation. We show that the glycosylation defect is restricted to alpha2,3-sialylation and can be detected in platelets, neutrophils, and monocytes. Platelets exhibited a distorted structure of the open canalicular system, indicating defective platelet generation. Importantly, patient platelets, but not normal platelets, bound to the asialoglycoprotein receptor (ASGP-R), a liver cell-surface protein that removes desialylated thrombocytes from the circulation in mice. Taken together, this is the first type of human thrombocytopenia in which a specific defect of alpha2,3-sialylation and an induction of platelet binding to the liver ASGP-R could be detected

Solid-phase synthesis of a pentavalent GalNAc-containing glycopeptide (Tn antigen) representing the nephropathy-associated IgA hinge region

Incomplete or aberrant glycosylation leading to Tn antigen (GalNAca1-Ser/Thr) expression on human glycoproteins is strongly associated with human pathological conditions, including tumors, certain autoimmune diseases, such as the idiopathic IgA nephropathy, and may modulate immune homeostasis. In addition, the Tn antigen is highly expressed by certain pathogens and plays a role in host-pathogen interactions. To enable experimental approaches to study interactions of the Tn antigen with the immune system and analyze anti-Tn antibody responses in infection or disorders, we generated a Tn-expressing resource that can be used for high-throughput screening. In consideration of IgA nephropathy in which the hinge region is incompletely glycosylated, we used this hinge sequence that encodes five potential glycosylation sites as the ideal template for the synthesis of a Tn antigen-expressing glycopeptide. Inclusion of an N-terminal biotin in the peptide enabled binding to streptavidin-coated ELISA plates as monitored using Helix pomatia agglutinin or anti-Tn monoclonal antibody. We also found that the biotinylated IgA-Tn peptide is a functional acceptor for b1-3-galactosylation using recombinant T-synthase (b1-3-galactosyltransferase). Besides its immunochemical functionality as a possible diagnostic tool for IgA nephropathy, the peptide is an excellent substrate for glycan elongation and represents a novel template applicable for glycan-antigen-associated diseases