Construction of a cosmid library of Spirulina platensis as an approach to DNA physical mapping

Abstract A Spirulina platensis gene library has been constructed using cosmid vector pMMB34. The cosmid bank was controlled for its random gene distribution by colony hybridization. Genes were identified using either homologous or heterologous probes of genes involved in photosynthesis (large and small subunit of d -ribulose 1,5-bisphosphate carboxylase, 32 kDa thylakoid protein, α, β subunits of C-phycocyanin) and protein synthesis (elongation factors EF-Tu, EF-G).     

Origin of the sarcosine molecules of actinomycins

1. Streptomyces V–187 produces on minimal medium a mixture composed mainly of actinomycin C₁ (actinomycin D) and actinomycin A₁ (actinomycin I). If sarcosine is added to the medium, the micro-organism produces, in addition to actinomycins C₁ and A₁, actinomycin F₈ (actinomycin II) and actinomycin F₉ (actinomycin (III), characterized by the substitution by sarcosine of one or both the proline molecules present in actinomycin C₁. 2. Exogenous sarcosine seems to be incorporated as such by Streptomyces V–187 only in the sarcosine molecule(s) that replace proline in the actinomycins of the F group, whereas, for the synthesis of the other sarcosine molecules, the amino acid is first demethylated to glycine. 3. The incorporation of sarcosine and glycine into actinomycin by Streptomyces antibioticus appears to follow a similar pattern, except that a portion of the methyl group produced in the degradation of sarcosine is utilized as a source of the methyl groups of the antibiotic. This explains the previously reported lack of cross-dilution between glycine and sarcosine observed in the incorporation of these amino acids into actinomycin.     

Assunzione di Fosforo32 da Parte di Cuscuta Epithymum Parassita di Trifolium Repens

Abstract

Uptake of Phosporus-P³² by CUSCUTA EPITHYMUM parasitic of TRIFOLIUM REPENS. — Filaments of C. epithymum, parasitic on T. repens, partially immersed in a solution containing KH₂P³²O₄, uptake from the host solely, or mainly, radioactive ortophosphate. Radioactive hexose phosphates and radioactive nucleotides present in the extracts prepared from C. epithymum, appear to be the result of the metabolization by the dodder of the orthophosphate uptaken from the host.     

Fissazione di CO2 in Cuscuta epithymum

Abstract

CO₂ FIXATION IN CUSCUTA EPITHYMUM. — Seedlings of Cuscuta epithymum fixe approximately the same amount of C¹⁴O₂ irrespectively of age, pigmentation, presence or absence of light. Examination by paper chromatography of the extracts of plants exposed to C¹¹O₂ revealed that most, or all, of the radioactivity is concentrated in the area of the organic acids and of the acidic amino acids. It is tentatively concluded that C. epithymum fixes carbon dioxide through a mechanism different from that involving ribulose-1,5-diphosphate carboxylase and carboxydismutase.     

Assunzione Di Metaboliti Da Parte Di « Cuscuta Epithymum » Parassita Di « Trifolium Repens »

Abstract

Uptake of metabolites by CUSCUTA. — The distribution of the radioactivity in the filaments of Cuscuta epithymum attached to the host (Trifolium repens) growing in an atmosphere of C¹⁴O₂ indicates that the dodder uptakes from the host solely, or mainly, sucrose. The other radioactive compounds present in the extracts of Cuscuta appear to be derived from the sucrose rather than uptaken from the host.     

Effetto Dell'attinomicina Sulla Crescita di Alcune Specie di Alghe

Abstract

Action of Actinomycin on the growth of Algae. — Actinomycin C inhibits the growth of two strains of Euglena gracilis, of Chlorella vulgaris, of Prototheca zopfii and of Scenedesmus sp. The growth inhibitory effect is evident on both autotrophically and heterotrophically grown cultures. DNA extracted from C. vulgaris appears to form a complex with Actinomycin C similar to that observed in the case of other organisms.     

An essay on the classification of Moniliaceous medical fungi

Summary A key and a plan of tentative classification of the fundamental families of medical fungi are given. It is based on the presence, in tissues or cultures, of different categories of true spores, conidia and vegetative spores (thallospores). The most important medical genera of fungi, with type species, basonym, author and data for each family of fungi are listed. The apparent morphological affinity between certain families of fungi is suggested.     

Abnormal kinetochore-generated pulling forces from expressing a N-terminally modified Hec1

Background

Highly Expressed in Cancer protein 1 (Hec1) is a constituent of the Ndc80 complex, a kinetochore component that has been shown to have a fundamental role in stable kinetochore-microtubule attachment, chromosome alignment and spindle checkpoint activation at mitosis. HEC1 RNA is found up-regulated in several cancer cells, suggesting a role for HEC1 deregulation in cancer. In light of this, we have investigated the consequences of experimentally-driven Hec1 expression on mitosis and chromosome segregation in an inducible expression system from human cells.

Methodology/Principal Findings

Overexpression of Hec1 could never be obtained in HeLa clones inducibly expressing C-terminally tagged Hec1 or untagged Hec1, suggesting that Hec1 cellular levels are tightly controlled. On the contrary, a chimeric protein with an EGFP tag fused to the Hec1 N-terminus accumulated in cells and disrupted mitotic division. EGFP- Hec1 cells underwent altered chromosome segregation within multipolar spindles that originated from centriole splitting. We found that EGFP-Hec1 assembled a mutant Ndc80 complex that was unable to rescue the mitotic phenotypes of Hec1 depletion. Kinetochores harboring EGFP-Hec1 formed persisting lateral microtubule-kinetochore interactions that recruited the plus-end depolymerase MCAK and the microtubule stabilizing protein HURP on K-fibers. In these conditions the plus-end kinesin CENP-E was preferentially retained at kinetochores. RNAi-mediated CENP-E depletion further demonstrated that CENP-E function was required for multipolar spindle formation in EGFP-Hec1 expressing cells.

Conclusions/Significance

Our study suggests that modifications on Hec1 N-terminal tail can alter kinetochore-microtubule attachment stability and influence Ndc80 complex function independently from the intracellular levels of the protein. N-terminally modified Hec1 promotes spindle pole fragmentation by CENP-E-mediated plus-end directed kinetochore pulling forces that disrupt the fine balance of kinetochore- and centrosome-associated forces regulating spindle bipolarity. Overall, our findings support a model in which centrosome integrity is influenced by the pathways regulating kinetochore-microtubule attachment stability.     

Mechanism of Aurora B activation by INCENP and inhibition by hesperadin

Aurora family serine/threonine kinases control mitotic progression, and their deregulation is implicated in tumorigenesis. Aurora A and Aurora B, the best-characterized members of mammalian Aurora kinases, are approximately 60% identical but bind to unrelated activating subunits. The structure of the complex of Aurora A with the TPX2 activator has been reported previously. Here, we report the crystal structure of Aurora B in complex with the IN-box segment of the inner centromere protein (INCENP) activator and with the small molecule inhibitor Hesperadin. The Aurora B:INCENP complex is remarkably different from the Aurora A:TPX2 complex. INCENP forms a crown around the small lobe of Aurora B and induces the active conformation of the T loop allosterically. The structure represents an intermediate state of activation of Aurora B in which the Aurora B C-terminal segment stabilizes an open conformation of the catalytic cleft, and a critical ion pair in the kinase active site is impaired. Phosphorylation of two serines in the carboxyl terminus of INCENP generates the fully active kinase.     

Implications for kinetochore-microtubule attachment from the structure of an engineered Ndc80 complex

Kinetochores are proteinaceous assemblies that mediate the interaction of chromosomes with the mitotic spindle. The 180 kDa Ndc80 complex is a direct point of contact between kinetochores and microtubules. Its four subunits contain coiled coils and form an elongated rod structure with functional globular domains at either end. We crystallized an engineered "bonsai" Ndc80 complex containing a shortened rod domain but retaining the globular domains required for kinetochore localization and microtubule binding. The structure reveals a microtubule-binding interface containing a pair of tightly interacting calponin-homology (CH) domains with a previously unknown arrangement. The interaction with microtubules is cooperative and predominantly electrostatic. It involves positive charges in the CH domains and in the N-terminal tail of the Ndc80 subunit and negative charges in tubulin C-terminal tails and is regulated by the Aurora B kinase. We discuss our results with reference to current models of kinetochore-microtubule attachment and centromere organization.