BcSAK1, a stress-activated mitogen-activated protein kinase, is involved in vegetative differentiation and pathogenicity in Botrytis cinerea

The gene bcsak1, encoding a mitogen-activated protein kinase (MAPK) of Botrytis cinerea, was cloned and characterized. The protein has high homology to the yeast Hog1 and to corresponding MAPKs from filamentous fungi, but it shows unique functional features. The protein is phosphorylated under osmotic stress, specific fungicides, and oxidative stress mediated by H(2)O(2) and menadione. Northern blot analyses indicate that only a subset of typical oxidative stress response genes is regulated by BcSAK1. In contrast to most other fungal systems, Deltabcsak1 mutants are significantly impaired in vegetative and pathogenic development: they are blocked in conidia formation, show increased sclerotial development, and are unable to penetrate unwounded plant tissue. These data indicate that in B. cinerea the stress-activated MAPK cascade is involved in essential differentiation programs.     

Minireview: Recent progress in hemocyanin research

This review summarizes recent highlights of our joint work on the structure, evolution, and function of a family of highly complex proteins, the hemocyanins. They are blue-pigmented oxygen carriers, occurring freely dissolved in the hemolymph of many arthropods and molluscs. They are copper type-3 proteins and bind one dioxygen molecule between two copper atoms in a side-on coordination. They possess between 6 and 160 oxygen-binding sites, and some of them display the highest molecular cooperativity observed in nature. The functional properties of hemocyanins can be convincingly described by either the Monod-Wyman-Changeux (MWC) model or its hierarchical extension, the Nested MWC model; the latter takes into account the structural hierarchies in the oligomeric architecture. Recently, we applied these models to interpret the influence of allosteric effectors in detailed terms. Effectors shift the allosteric equilibria but have no influence on the oxygen affinities characterizing the various conformational states. We have shown that hemocyanins from species living at different environmental temperatures have a cooperativity optimum at the typical temperature of their natural habitat. Besides being oxygen carriers, some hemocyanins function as a phenoloxidase (tyrosinase/catecholoxidase) which, however, requires activation. Chelicerates such as spiders and scorpions lack a specific phenoloxidase, and in these animals activated hemocyanin might catalyse melanin synthesis in vivo. We propose a similar activation mechanism for arthropod hemocyanins, molluscan hemocyanins and tyrosinases: amino acid(s) that sterically block the access of phenolic compounds to the active site have to be removed. The catalysis mechanism itself can now be explained on the basis of the recently published crystal structure of a tyrosinase. In a series of recent publications, we presented the complete gene and primary structure of various hemocyanins from different molluscan classes. From these data, we deduced that the molluscan hemocyanin molecule evolved ca. 740 million years ago, prior to the separation of the extant molluscan classes. Our recent advances in the 3D cryo-electron microscopy of hemocyanins also allow considerable insight into the oligomeric architecture of these proteins of high molecular mass. In the case of molluscan hemocyanin, the structure of the wall and collar of the basic decamers is now rapidly becoming known in greater detail. In the case of arthropod hemocyanin, a 10-? structure and molecular model of the Limulus 8 × 6mer shows the amino acids at the various interfaces between the eight hexamers, and reveals histidine-rich residue clusters that might be involved in transferring the conformational signals establishing cooperative oxygen binding.     

Extinction pattern of Alpine cave bears - new data and climatological interpretation

ABSTRACT

Cave bears have disappeared from the Alps from different altitudes at different times. The temporal progression of the HDEL (Height Dependent Extinction Line) – a compilation of the geologically most recent radiocarbon dates per altitude level – is not consistent with the general cooling of the temperatures from about 45 ka BP. The cave bear sites of the Northern Alps with the most recent radiocarbon ages are not situated in the lowlands but in caves in altitudes of 1,500 m to 1,700 m above sea level (a.s.l.).

Cave bears fed almost exclusively on herbs and leaves. It was assumed that with the general cooling in the OIS 3 since about 45 ka BP also the migration of the alpine elements into the lowlands took place. It could be recognized that the populations in the lower situated cave bear site became earlier extinct than the cave bear population in the higher altitudes.

With new radiocarbon dates, done at the Curt-Engelhorn-Center Archaeometry at the Reiss-Engelhorn-Museen in Mannheim (Germany), the HDEL can be determined much more precisely and the causes of gradual extinction are also better understood.     

Phylogeography and population genetic structure of the European roe deer in Switzerland following recent recolonization

In the early 1800s, the European roe deer (Capreolus capreolus) was probably extirpated from Switzerland, due to overhunting and deforestation. After a federal law was enacted in 1875 to protect lactating females and young, and limiting the hunting season, the roe deer successfully recovered and recolonized Switzerland. In this study, we use mitochondrial DNA and nuclear DNA markers to investigate the recolonization and assess contemporary genetic structure in relation to broad topographic features, in order to understand underlying ecological processes, inform future roe deer management strategies, and explore the opportunity for development of forensic traceability tools. The results concerning the recolonization origin support natural, multidirectional immigration from neighboring countries. We further demonstrate that there is evidence of weak genetic differentiation within Switzerland among topographic regions. Finally, we conclude that the genetic data support the recognition of a single roe deer management unit within Switzerland, within which there is a potential for broad‐scale geographic origin assignment using nuclear markers to support law enforcement.     

Sexual behavior and hormonal estrus cycles in captive aged lowland gorillas (Gorilla gorilla)

To evaluate whether observed cycles in proceptive behavior in aging lowland gorilla females (age 40+) at Brookfield Zoo were driven by ovarian activity, we compared monthly behavioral data to estradiol and progestogen cycles based on fecal hormone assessments. Progestogen peaks showed regularity and close coincidence with monthly sexual behaviors. Estradiol was more variable. Progestogen peaks varied between 22+/-5 days for the control female (29 years old), to 24+/-2.5 and 29+/-8 for the two aged subjects. In the first aged female, which was housed with other females and a silverback, the high degree of cyclicity in sexual behavior, regularity of progestogen cycles, and close concordance between hormonal cycling and sexual behavior strongly compared to patterns found (in this and other studies) in gorilla females <35 years old. Cyclical progestogen peaks were longer and more variable in the second aged female-perhaps because she lacked the social mediation of other females or a male. For husbandry reasons she is not housed with the gorilla group, behavioral data were not collected from her. The value of our longitudinal study is in obtaining reproductive profiles of primate females that are approaching maximum lifespan. This pilot study is part of a larger research project on reproductive senescence that will include other captive females >35 years old, a population that is rapidly increasing in North American zoos as gorillas continue to age.     

The MHC class I-like IgG receptor controls perinatal IgG transport,IgG homeostasis,and fate of IgG-Fc-coupled drugs

Abs of the IgG isotype are efficiently transported from mother to neonate and have an extended serum t(1/2) compared with Abs of other isotypes. Circumstantial evidence suggests that the MHC class I-related protein, the neonatal FcR (FcRn), is the FcR responsible for both in vivo functions. To understand the phenotypes imposed by FcRn, we produced and analyzed mice with a defective FcRn gene. The results provide direct evidence that perinatal IgG transport and protection of IgG from catabolism are mediated by FcRn, and that the latter function is key to IgG homeostasis, essential for generating a potent IgG response to foreign Ags, and the basis of enhanced efficacy of Fc-IgG-based therapeutics. FcRn is therefore a promising therapeutic target for enhancing protective humoral immunity, treating autoimmune disease, and improving drug efficacy.     

Development of an in vitro integration assay for the Bacteroides conjugative transposon CTnDOT

Integrated self-transmissible elements called conjugative transposons (CTns) are responsible for the transfer of antibiotic resistance genes in many different species of bacteria. One of the best characterized of these newly recognized elements is the Bacteroides CTn, CTnDOT. CTnDOT is thought to have a circular transfer intermediate that transfers to and integrates into the genome of the recipient cell. Previous investigations of the mechanism of CTnDOT integration have been hindered by the lack of an in vitro system for checking this model of integration and determining whether the CTnDOT integrase alone was sufficient to catalyze the integration reaction or whether host factors might be involved. We report here the development of an in vitro system in which a plasmid containing the joined ends of CTnDOT integrates into a plasmid carrying a CTnDOT target site. To develop this in vitro system, a His-tagged version of the integrase gene of CTnDOT was cloned and shown to be active in vivo. The protein produced by this construct was partially purified and then added to a reaction mixture that contained the joined ends of the circular form of CTnDOT and a plasmid carrying one of the CTnDOT target sites. Integration was demonstrated by using a fairly simple mixture of components, but integration was stimulated by a Bacteroides extract or by purified Escherichia coli integration host factor. The results of this study demonstrate both that the circular form of CTnDOT is the form that integrates into the target site and that host factors are involved in the integration process.     

Mitochondria-rich cells as experimental model in studies of epithelial chloride channels

The mitochondria-rich (mr) cell of amphibian skin epithelium is differentiated as a highly specialised pathway for passive transepithelial transport of chloride. The apical membrane of mr cells expresses several types of Cl(-) channels, of which the function of only two types has been studied in detail. (i) One type of channel is gated by voltage and external chloride concentration. This intriguing type of regulation leads to opening of channels only if [Cl(-)](o) is in the millimolar range and if the electrical potential is of a polarity that secures an inwardly directed net flux of this ion. Reversible voltage activations of the conductance proceed with long time constants, which depend on V in such a way that the rate of conductance activation increases when V is clamped at more negative values (serosal bath grounded). The gating seems to involve processes that are dependent on F-actin localised in the submembrane domain in the neck region of the flask-shaped mr cell. (ii) The other identified Cl(-) pathway of mr cells is mediated by small-conductance apical CFTR chloride channels as concluded from its activation via beta-adrenergic receptors, ion selectivity, genistein stimulation and inhibition by glibenclamide. bbCFTR has been cloned, and immunostaining has shown that the gene product is selectively expressed in mr cells. There is cross-talk between the two pathways in the sense that activation of the conductance of the mr cell by voltage clamping excludes activation via receptor occupation, and vice versa. The mechanism of this cross-talk is unknown.