Lectin-like components in the macronuclear replication bands of Euplotes eurystomus

Upon incubation with fluoresceinylated neoglycoproteins, isolated macronuclei from the ciliated protozoan Euplotes eurystomus display different labelling patterns depending on the nature of the sugar bound to the neoglycoproteins. Specific sugar-binding components (i.e., lectin-like molecules) are associated with presumed nucleoli and with the macronuclear replication bands. This is the first demonstration that DNA synthesis and sugar-binding components are co-localized in an eukaryotic cell.     

Lichtabhängige Neusynthese von δ-Aminolaevulinsäure-Dehydratase in isolierten Avena-Etioplasten

Summary A light dependent increase of the activity of -aminolaevulinate dehydratase (E.C. 4.2.1.24) in isolated etioplasts of Avena sativa L. was shown. This increase can be assumed to be due to a de-novo synthesis of the enzyme. Chloramphenicol was found to inhibit this synthesis, whereas cycloheximide did not have any effect. Illumination with red light (660 nm) was followed by the same increase of porphobilinogen production as illumination with white light; far-red (731 nm) did not induce such an effect. It is concluded that a phytochrome-mediated mechanism is involved in the induction of -aminolaevulinate dehydratase synthesis.

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Abkürzungen ALA -Aminolaevulinsäure - CAP Chloramphenicol - CHI Cycloheximid - PBG Porphobilinogen     

Gene silencing quantitatively controls the function of a developmental trans-activator

How a single cell gives rise to progeny with differing fates remains poorly understood. We examined cells lacking methyl CpG binding domain protein-2 (MBD2), a molecule that has been proposed to link DNA methylation to silent chromatin. Helper T cells from Mbd2(-/-) mice exhibit disordered differentiation. IL-4, the signature of a restricted set of progeny, is expressed ectopically in Mbd2(-/-) parent and daughter cells. Loss of MBD2-mediated silencing renders the normally essential activator, Gata-3, dispensable for IL-4 induction. Gata-3 and MBD2 act in competition, wherein each factor independently, and quantitatively, regulates the binary choice of whether heritable IL-4 expression is established. Gata-3 functions, in part, to displace MBD2 from methylated DNA. These results suggest that activating and silencing signals integrate to provide spatially and temporally restricted patterns of gene activity.     

Role of C-terminal domain and transmembrane helices 5 and 6 in function and quaternary structure of major intrinsic proteins: analysis of aquaporin/glycerol facilitator chimeric proteins

We previously observed that aquaporins and glycerol facilitators exhibit different oligomeric states when studied by sedimentation on density gradients following nondenaturing detergent solubilization. To determine the domains of major intrinsic protein (MIP) family proteins involved in oligomerization, we constructed protein chimeras corresponding to the aquaporin AQPcic substituted in the loop E (including the proximal part of transmembrane domain (TM) 5) and/or the C-terminal part (including the distal part of TM 6) by the equivalent domain of the glycerol channel aquaglyceroporin (GlpF) (chimeras called AGA, AAG, and AGG). The analogous chimeras of GlpF were also constructed (chimeras GAG, GGA, and GAA). cRNA corresponding to all constructs were injected into Xenopus oocytes. AQPcic, GlpF, AAG, AGG, and GAG were targeted to plasma membranes. Water or glycerol membrane permeability measurements demonstrated that only the AAG chimera exhibited a channel function corresponding to water transport. Analysis of all proteins expressed either in oocytes or in yeast by velocity sedimentation on sucrose gradients following solubilization by 2% n-octyl glucoside indicated that only AQPcic and AAG exist in tetrameric forms. GlpF, GAG, and GAA sediment in a monomeric form, whereas GGA and AGG were found mono/dimeric. These data bring new evidence that, within the MIP family, aquaporins and GlpFs behave differently toward nondenaturing detergents. We demonstrate that the C-terminal part of AQPcic, including the distal half of TM 6, can be substituted by the equivalent domain of GlpF (AAG chimera) without modifying the transport specificity. Our results also suggest that interactions of TM 5 of one monomer with TM 1 of the adjacent monomer are crucial for aquaporin tetramer stability.     

Porphyrias: Animal models and prospects for cellular and gene therapy

The rapid progress in the development of molecular technology has resulted in the identification of most of the genes of the heme biosynthesis pathway. Important problems in the pathogenesis and treatment of porphyrias now seem likely to be solved by the possibility of creating animal models and by the transfer of normal genes or cDNAs to target cells. Animal models of porphyrias naturally occur for erythropoietic protoporphyria and congenital erythropoietic porphyria, and different murine models have been or are being created for erythropoietic and hepatic porphyrias. The PBGD knock-out mouse will be useful for the understanding of nervous system dysfunction in acute porphyrias. Murine models of erythropoietic porphyrias are being used for bone-marrow transplantation experiments to study the features of erythropoietic and hepatic abnormalities. Gene transfer experiments have been startedin vitro to look at the feasibility of somatic gene therapy in erythropoietic porphyrias. In particular, we have documented sufficient gene transfer rate and metabolic correction in different CEP disease cells to indicate that this porphyria is a good candidate for treatment by gene therapy in hematopoietic stem cells. With the rapid advancement of methods that may allow more precise and/or efficient gene targeting, gene therapy will become a new therapeutic option for porphyrias.     

Ant-repellent terpenoids from Melampodium divaricatum

The leaves of Melampodium divaricatum have been systematically fractionated by following biological activity in an assay which measures repellency to the leafcutter ant Atta cephalotes (Formicidae, Attini). Several terpenoids have been isolated which show significant ant-repellency.     

Characterization of 79 microsatellite DNA markers in the Pacific oyster Crassostrea gigas

We characterized 79 microsatellite DNA markers, which were obtained from genomic libraries enriched for CA, GA, ATG and TAGA motif repeats, in the Pacific oyster Crassostrea gigas. For eight F₁ grandparents or great‐grandparents of mapping families, the average heterozygosity, 0.705, and average number of alleles per locus, 5.7, did not vary among motif‐repeat or motif‐complexity categories. Non‐amplifying polymerase chain reaction null alleles, which were confirmed by segregation in the mapping families, were detected at 41 (51.9%) of the 79 loci. Cross‐species amplifications from C. angulata, C. sikamea, C. ariakensis and C. virginica showed a precipitous decline with distance from the focal species C. gigas.     

Bypass of A- and B-signaling requirements for Myxococcus xanthus development by mutations in spdR

Mutations in spdR, previously reported to bypass the developmental requirement for B-signaling in Myxococcus xanthus, also bypass the requirement for A-signaling but not C-, D-, or E-signaling. Mutations in spdR restored nearly wild-type levels of sporulation to representative A-signal-deficient mutants carrying asgA476, asgB480, and asgC767 and improved the quality of fruiting body formation in the asgB480 mutant. The defect in A-factor production by the asgB480 mutant was not restored in the spdR2134 asgB480 double mutant.