Male flowers in Tulipa pumila Moench (Liliaceae) potentially originate from gender diphasy

We found functionally male individuals in an otherwise hermaphroditic population of Tulipa pumila (Liliaceae) located in Tuscany (central Italy). We investigated the sex ratio of this population, followed by morphometric analyses of the scape, leaves and flowers, and tests on pollen germinability and seed number and mass, in order to infer which sexual strategy produced the observed co-occurrence of male and hermaphrodite individuals. We found that sex ratio deviated from 1:1, and functionally male individuals showed a smaller plant size and a reduced pollen fitness (germinability and siring ability) compared to hermaphrodites. These findings point to a resource-dependent sexual allocation strategy, probably associated with gender diphasy.     

Phylogenetic relationships of Italian Bellevalia species (Asparagaceae), inferred from morphology,karyology and molecular systematics

The seven Bellevalia species and subspecies known from Italy, representing about 10% of the genus and three out of six sections, were studied. An integrated morphological, karyological and molecular approach was used to infer phylogenetic and systematic relationships among them. B. romana (the generitype) is the most distinctive species on karyotype asymmetry grounds. B. boissieri and B.dubia, usually considered as subspecies of one species (the latter endemic to Sicily), deserve specific status based on biparental nrDNA markers (internal transcribed spacer, ITS), since they do not form a single clade. The allotetraploid endemic B. pelagica, morphologically similar to B. romana, is sister to the latter under parsimony, both in morphological and ITS trees; it is also related with B. dubia, based on karyotype asymmetry and a uniparental cpDNA marker (trnL^(UAA)–trnF^(GAA) IGS (intergenic spacer)). A second allotetraploid endemic, B. webbiana, is closely related, on morphological, karyological and molecular grounds, with B. boissieri and B. ciliata, and also with B. trifoliata, three species that might all involved in its origin. B. sect. Conicae Feinbr. and sect. Nutantes Feinbr. are here typified, the former (type: B. ciliata) is most likely a synonym of the latter (type: B. trifoliata).     

CDC25B Overexpression Stabilises Centrin 2 and Promotes the Formation of Excess Centriolar Foci

CDK-cyclin complexes regulate centriole duplication and microtubule nucleation at specific cell cycle stages, although their exact roles in these processes remain unclear. As the activities of CDK-cyclins are themselves positively regulated by CDC25 phosphatases, we investigated the role of centrosomal CDC25B during interphase. We report that overexpression of CDC25B, as is commonly found in human cancer, results in a significant increase in centrin 2 at the centrosomes of interphase cells. Conversely, CDC25B depletion causes a loss of centrin 2 from the centrosome, which can be rescued by treatment with the proteasome inhibitor MG132. CDC25B overexpression also promotes the formation of excess centrin 2 “foci”. These foci can accumulate other centrosome proteins, including γ-tubulin and PCM-1, and can function as microtubule organising centres, indicating that these represent functional centrosomes. Formation of centrin 2 foci can be blocked by specific inhibition of CDK2 but not CDK1. CDK2-mediated phosphorylation of Monopolar spindle 1 (Mps1) at the G1/S transition is essential for the initiation of centrosome duplication, and Mps1 is reported to phosphorylate centrin 2. Overexpression of wild-type or non-degradable Mps1 exacerbated the formation of excess centrin 2 foci induced by CDC25B overexpression, while kinase-dead Mps1 has a protective effect. Together, our data suggest that CDC25B, through activation of a centrosomal pool of CDK2, stabilises the local pool of Mps1 which in turn regulates the level of centrin 2 at the centrosome. Overexpression of CDC25B may therefore contribute to tumourigenesis by perturbing the natural turnover of centrosome proteins such as Mps1 and centrin 2, thus resulting in the de novo assembly of extra-numerary centrosomes and potentiating chromosome instability.     

Nitric oxide signaling in yeast

As a cellular signaling molecule, nitric oxide (NO) is widely conserved from microorganisms, such as bacteria, yeasts, and fungi, to higher eukaryotes including plants and mammals. NO is mainly produced by NO synthase (NOS) or nitrite reductase (NIR) activity. There are several NO detoxification systems, including NO dioxygenase (NOD) and S-nitrosoglutathione reductase (GSNOR). NO homeostasis based on the balance between NO synthesis and degradation is important for the regulation of its physiological functions because an excess level of NO causes nitrosative stress due to the high reactivity of NO and NO-derived compounds. In yeast, NO may be involved in stress responses, but NO and its signaling have been poorly understood due to the lack of mammalian NOS orthologs in the genome. Even though the activities of NOS and NIR have been observed in yeast cells, the gene encoding NOS and the NO production mechanism catalyzed by NIR remain unclear. On the other hand, yeast cells employ NOD and GSNOR to maintain an intracellular redox balance following endogenous NO production, exogenous NO treatment, or environmental stresses. This article reviews NO metabolism (synthesis, degradation) and its regulation in yeast. The physiological roles of NO in yeast, including the oxidative stress response, are also discussed here. Such investigations into NO signaling are essential for understanding the NO-dependent genetic and physiological modulations. In addition to being responsible for the pathology and pharmacology of various degenerative diseases, NO signaling may be a potential target for the construction and engineering of industrial yeast strains.     

Unique Characteristics of Recombinant Hybrid Manganese Superoxide Dismutase from <Emphasis Type="Italic">Staphylococcus equorum and S. saprophyticus</Emphasis>

A recombinant hybrid of manganese dependent-superoxide dismutase of Staphylococcus equorum and S. saprophyticus has successfully been overexpressed in Escherichia coli BL21(DE3), purified, and characterized. The recombinant enzyme suffered from degradation and aggregation upon storage at ?20 °C, but not at room temperature nor in cold. Chromatographic analysis in a size exclusion column suggested the occurrence of dimeric form, which has been reported to contribute in maintaining the stability of the enzyme. Effect of monovalent (Na⁺, K⁺), divalent (Ca²⁺, Mg²⁺), multivalent (Mn^2+/4+, Zn^2+/4+) cations and anions (Cl^?, SO₄ ^2?) to the enzyme stability or dimeric state depended on type of cation or anion, its concentration, and pH. However, tremendous effect was observed with 50 mM ZnSO₄, in which thermostability of both the dimer and monomer was increased. Similar situation was not observed with MnSO₄, and its presence was detrimental at 200 mM. Finally, chelating agent appeared to destabilize the dimer around neutral pH and dissociate it at basic pH. The monomer remained stable upon addition of ethylene diamine tetraacetic acid. Here we reported unique characteristics and stability of manganese dependent-superoxide dismutase from S. equorum/saprophyticus.     

MAPK Pathway Activation Delays G2/M Progression by Destabilizing Cdc25B

Activation of the mitogen-activated protein kinase (MAPK) pathway by growth factors or phorbol esters during G₂ phase delays entry into mitosis; however, the role of the MAPK pathway during G₂/M progression remains controversial. Here, we demonstrate that activation of the MAPK pathway with either epidermal growth factor or 12-O-tetradecanoylphorbol-13-acetate induces a G₂ phase delay independent of known G₂ phase checkpoint pathways but was specifically dependent on MAPK/extracellular signal-regulated kinase kinase (MEK1). Activation of MAPK signaling also blocked exit from a G₂ phase checkpoint arrest. Both the G₂ phase delay and blocked exit from the G₂ checkpoint arrest were mediated by the MEK1-dependent destabilization of the critical G₂/M regulator cdc25B. Reintroduction of cdc25B overcame the MEK1-dependent G₂ phase delay. Thus, we have demonstrated a new function for MEK1 that controls G₂/M progression by regulating the stability of cdc25B. This represents a novel mechanism by which factors that activate MAPK signaling can influence the timing of entry into mitosis, particularly exit from a G₂ phase checkpoint arrest.     

Mitotic Phosphorylation of Cdc25B Ser321 Disrupts 14-3-3 Binding to the High Affinity Ser323 Site

Cdc25B is a key regulator of entry into mitosis, and its activity and localization are regulated by binding of the 14-3-3 dimer. There are three 14-3-3 binding sites on Cdc25B, with Ser³²³ being the highest affinity binding and is highly homologous to the Ser²¹⁶ 14-3-3 binding site on Cdc25C. Loss of 14-3-3 binding to Ser³²³ increases cyclin/Cdk substrate access to the catalytic site, thereby increasing its activity. It also affects the localization of Cdc25B. Thus, phosphorylation and 14-3-3 binding to this site is essential for down-regulating Cdc25B activity, blocking its mitosis promoting function. The question of how this inhibitory signal is relieved to allow Cdc25B activation and entry into mitosis is yet to be resolved. Here, we show that Ser³²³ phosphorylation is maintained into mitosis, but phosphorylation of Ser³²¹ disrupts 14-3-3 binding to Ser³²³, mimicking the effect of inhibiting Ser³²³ phosphorylation on both Cdc25B activity and localization. The unphosphorylated Ser³²¹ appears to have a role in stabilizing 14-3-3 binding to Ser³²³, and loss of the Ser hydroxyl group appears to be sufficient to significantly reduce 14-3-3 binding. A consequence of loss of 14-3-3 binding is dephosphorylation of Ser³²³. Ser³²¹ is phosphorylated in mitosis by Cdk1. The mitotic phosphorylation of Ser³²¹ acts to maintain full activation of Cdc25B by disrupting 14-3-3 binding to Ser³²³ and enhancing the dephosphorylation of Ser³²³ to block 14-3-3 binding to this site.