Distribution of mitochondrial NADH fluorescence lifetimes: steady-state kinetics of matrix NADH interactions

The lifetimes of fluorescent components of matrix NADH in isolated porcine heart mitochondria were investigated using time-resolved fluorescence spectroscopy. Three distinct lifetimes of fluorescence were resolved: 0.4 (63%), 1.8 (30%), and 5.7 (7%) ns (% total NADH). The 0.4 ns lifetime and the emission wavelength of the short component were consistent with free NADH. In addition to their longer lifetimes, the remaining pools also had a blue-shifted emission spectrum consistent with immobilized NADH. On the basis of emission frequency and lifetime data, the immobilized pools contributed >80% of NADH fluorescence. The steady-state kinetics of NADH entering the immobilized pools was measured in intact mitochondria and in isolated mitochondrial membranes. The apparent binding constants (K(D)s) for NADH in intact mitochondria, 2.8 mM (1.9 ns pool) and >3 mM (5.7 ns pool), were on the order of the estimated matrix [NADH] (approximately 3.5 mM). The affinities and fluorescence lifetimes resulted in an essentially linear relationship between matrix [NADH] and NADH fluorescence intensity. Mitochondrial membranes had shorter emission lifetimes in the immobilized poo1s [1 ns (34%) and 4.1 ns (8%)] with much higher apparent K(D)s of 100 microM and 20 microM, respectively. The source of the stronger NADH binding affinity in membranes is unknown but could be related to high order structure or other cofactors that are diluted out in the membrane preparation. In both preparations, the rate of NADH oxidation was proportional to the amount of NADH in the long lifetime pools, suggesting that a significant fraction of the bound NADH might be associated with oxidative phosphorylation, potentially in complex 1.     

Concordant phylogeography and cryptic speciation in two Western Palaearctic oak gall parasitoid species complexes

Little is known about the evolutionary history of most complex multi‐trophic insect communities. Widespread species from different trophic levels might evolve in parallel, showing similar spatial patterns and either congruent temporal patterns (Contemporary Host‐tracking) or later divergence in higher trophic levels (Delayed Host‐tracking). Alternatively, host shifts by natural enemies among communities centred on different host resources could disrupt any common community phylogeographic pattern. We examined these alternative models using two Megastigmus parasitoid morphospecies associated with oak cynipid galls sampled throughout their Western Palaearctic distributions. Based on existing host cynipid data, a parallel evolution model predicts that eastern regions of the Western Palaearctic should contain ancestral populations with range expansions across Europe about 1.6 million years ago and deeper species‐level divergence at both 8–9 and 4–5 million years ago. Sequence data from mitochondrial cytochrome b and multiple nuclear genes showed similar phylogenetic patterns and revealed cryptic genetic species within both morphospecies, indicating greater diversity in these communities than previously thought. Phylogeographic divergence was apparent in most cryptic species between relatively stable, diverse, putatively ancestral populations in Asia Minor and the Middle East, and genetically depauperate, rapidly expanding populations in Europe, paralleling patterns in host gallwasp species. Mitochondrial and nuclear data also suggested that Europe may have been colonized multiple times from eastern source populations since the late Miocene. Temporal patterns of lineage divergence were congruent within and across trophic levels, supporting the Contemporary Host‐tracking Hypothesis for community evolution.     

Characterization of commercial laminin preparations from human placenta in comparison to recombinant laminins 2 (alpha2beta1gamma1), 8 (alpha4beta1gamma1), 10 (alpha5beta1gamma1).

Laminins, a family of large heterotrimeric (alphabetagamma) proteins, are major components of basement membranes implicated in a variety of cellular functions. Different commercial laminin preparations isolated from human placenta have been widely used in functional studies but their molecular properties are poorly known. In the present study, we characterized several of these preparations by ELISA, silver staining and Western blotting, in comparison to mouse laminin 1 (alpha1beta1gamma1), and recombinant human laminins 2 (alpha2beta1gamma1), 8 (alpha4beta1gamma1) and 10 (alpha5beta1gamma1). The cell migration-promoting activity of different batches was also tested. The placenta laminin preparations differed from one another and consisted of highly fragmented proteins, a mixture of laminin isoforms, and/or contaminating fibronectin. Major functional differences between batches were also observed, reflecting molecular heterogeneity. Previous data obtained in functional studies using these preparations need to be interpreted with caution and may require revision, and future functional studies demand prior molecular characterization of the laminins, particularly their alpha-chain.     

A generalization of the van-der-Pol oscillator underlies active signal amplification in Drosophila hearing

The antennal hearing organs of the fruit fly Drosophila melanogaster boost their sensitivity by an active mechanical process that, analogous to the cochlear amplifier of vertebrates, resides in the motility of mechanosensory cells. This process nonlinearly improves the sensitivity of hearing and occasionally gives rise to self-sustained oscillations in the absence of sound. Time series analysis of self-sustained oscillations now unveils that the underlying dynamical system is well described by a generalization of the van-der-Pol oscillator. From the dynamic equations, the underlying amplification dynamics can explicitly be derived. According to the model, oscillations emerge from a combination of negative damping, which reflects active amplification, and a nonlinear restoring force that dictates the amplitude of the oscillations. Hence, active amplification in fly hearing seems to rely on the negative damping mechanism initially proposed for the cochlear amplifier of vertebrates.     

Chaperones that cure yeast artificial [PSI+] and their prion-specific effects

The [PSI(+)] nonsense-suppressor determinant of Saccharomyces cerevisiae results from the ability of Sup35 (eRF3) translation termination factor to undergo prion-like aggregation [1]. Although this process is autocatalytic, in vivo it depends on the chaperone Hsp104, whose lack or overexpression can cure [PSI(+)] [2]. Overproduction of the chaperone protein Ssb1 increased the [PSI(+)] curing by excess Hsp104, although it had no effect on its own, and excess chaperone protein Ssa1 protected [PSI(+)] against Hsp104 [3,4]. We used an artificial [PSI(+)(PS)] based on the Sup35 prion-forming domain from yeast Pichia methanolica [5] to find other prion-curing factors. Both [PSI(+)(PS)] and [PSI(+)] have prion 'strains', differing in their suppressor efficiency and mitotic stability. We show that [PSI(+)(PS)] and a 'weak' strain of [PSI(+)] can be cured by overexpression of chaperones Ssa1, Ssb1 and Ydj1. The ability of different chaperones to cure [PSI(+)(PS)] showed significant prion strain specificity, which could be related to variation in Sup35 prion structure. Our results imply that homologs of these chaperones may be active against mammalian prion and amyloid diseases.     

Serosurvey of free-ranging Amur tigers in the Russian Far East

Wild Amur tigers (Panthera tigris altaica, n=44) from the Russian Far East were tested for antibodies to feline leukemia virus, feline corona virus (FCoV), feline immunodeficiency virus, feline parvovirus (FPV), canine distemper virus (CDV), Toxoplasma gondii, and Bartonella henselae. Antibodies to FCoV, CDV, FPV, and T. gondii were detected in 43, 15, 68, and 42% of tigers, respectively. No differences were detected in antibody prevalence estimates between tigers captured as part of a research program and those captured to mitigate human-tiger conflicts. Domestic dogs (Canis familiaris) were tested as a potential source for CDV; 16% were vaccinated against CDV and 58% of unvaccinated dogs were antibody positive for CDV. A high percentage of tigers were exposed to potential pathogens that could affect the survival of this species. We recommend continued monitoring of wild tigers throughout Asia, development of standardized sampling and postmortem examination procedures, and additional research to better understand potential domestic and wild animal sources for these pathogens.     

Plant genera and species new to China recently found in northwest Xinjiang

Two genera new to China, Fibigia and Pachyneurum and 10 species found in China for the first time, Puccinellia kalininae, Stellaria pulvinata, Nanophyton mongolicum, Capsella orienalis, Pachyneurum grandiflorum, Fibigia spathulata, Craniospermum tuvinicum, Euphrasia schischkinii, E. syreitschikovii, Veronica schmakovii along with 3 species previously unknown in Xinjiang, Rorippa indica, Acalypha australis, and Phalaris canariensis, found in northwest Xinjiang are reported. In addition, the distribution of two species in China is corrected: Lepidium densiflorum is firstly reported for Nei Mongol while the occurrence of Draba sibirica in Gansu is not confirmed. Results of floristic studies by the Chinese–Russian Altai expedition during 2004–2007 are also summarized including 34 species and 1 subspecies revealed as new for China; 7 species confirmed to occur in China; one genus and 5 species not confirmed for China, as well as a number of new records for various provinces, mostly Xinjiang.