TRPV4 calcium entry channel: a paradigm for gating diversity

The vanilloid receptor-1 (VR1, now TRPV1) was the founding member of a subgroup of cation channels within the TRP family. The TRPV subgroup contains six mammalian members, which all function as Ca2+ entry channels gated by a variety of physical and chemical stimuli. TRPV4, which displays 45% sequence identity with TRPV1, is characterized by a surprising gating promiscuity: it is activated by hypotonic cell swelling, heat, synthetic 4alpha-phorbols, and several endogenous substances including arachidonic acid (AA), the endocannabinoids anandamide and 2-AG, and cytochrome P-450 metabolites of AA, such as epoxyeicosatrienoic acids. This review summarizes data on TRPV4 as a paradigm of gating diversity in this subfamily of Ca2+ entry channels.     

Bcl-2 family member Bfl-1/A1 sequesters truncated bid to inhibit is collaboration with pro-apoptotic Bak or Bax

Following caspase-8 mediated cleavage, a carboxyl-terminal fragment of the BH3 domain-only Bcl-2 family member Bid transmits the apoptotic signal from death receptors to mitochondria. In a screen for possible regulators of Bid, we defined Bfl-1/A1 as a potent Bid interacting protein. Bfl-1 is an anti-apoptotic Bcl-2 family member, whose preferential expression in hematopoietic cells and endothelium is controlled by inflammatory stimuli. Its mechanism of action is unknown. We find that Bfl-1 associates with both full-length Bid and truncated (t)Bid, via the Bid BH3 domain. Cellular expression of Bfl-1 confers protection against CD95- and Trail receptor-induced cytochrome c release. In vitro assays, using purified mitochondria and recombinant proteins, demonstrate that Bfl-1 binds full-length Bid, but does not interfere with its processing by caspase-8, or with its mitochondrial association. Confocal microscopy supports that Bfl-1, which at least in part constitutively localizes to mitochondria, does not impede tBid translocation. However, Bfl-1 remains tightly and selectively bound to tBid and blocks collaboration between tBid and Bax or Bak in the plane of the mitochondrial membrane, thereby preventing mitochondrial apoptotic activation. Lack of demonstrable interaction between Bfl-1 and Bak or Bax in the mitochondrial membrane suggests that Bfl-1 generally prevents the formation of a pro-apoptotic complex by sequestering BH3 domain-only proteins.     

Six related nucleoside/nucleobase transporters from Trypanosoma brucei exhibit distinct biochemical functions

Purine nucleoside and nucleobase transporters are of fundamental importance for Trypanosoma brucei and related kinetoplastid parasites because these protozoa are not able to synthesize purines de novo and must salvage the compounds from their hosts. In the studies reported here, we have identified a family of six clustered genes in T. brucei that encode nucleoside/nucleobase transporters. These genes, TbNT2/927, TbNT3, TbNT4, TbNT5, TbNT6, and TbNT7, have predicted amino acid sequences that show high identity to each other and to TbNT2, a P1 type nucleoside transporter recently identified in our laboratory. Expression in Xenopus laevis oocytes revealed that TbNT2/927, TbNT5, TbNT6, and TbNT7 are high affinity adenosine/inosine transporters with K(m) values of <5 microm. In addition, TbNT5, and to a limited degree TbNT6 and TbNT7, also mediate the uptake of the nucleobase hypoxanthine. Ribonuclease protection assays showed that mRNA from all of the six members of this gene family are expressed in the bloodstream stage of the T. brucei life cycle but that TbNT2/927 and TbNT5 mRNAs are also expressed in the insect stage of the life cycle. These results demonstrate that T. brucei expresses multiple purine transporters with distinct substrate specificities and different patterns of expression during the parasite life cycle.     

DNA Repair 2008: Tenth Biennial Meeting of the German Society for Research on DNA Repair

The Tenth Meeting of the German Society for Research on DNA Repair was held in Berlin in September 2008. Invited presentations by Yosef Shiloh, Stanton L. Gerson, Sacha Beneke, Patrick Concannon, Jochen Dahm-Daphi, Thilo Dörk, Friedrike Eckardt-Schupp, Bernd Epe, Ian Hickson, Ulrich Hübscher, Penny Jeggo, Malik Lutzmann, Christof Niehrs, Primo Schär and Predrag Slijepcevic together with over 80 selected oral and poster presentations generated an inspiring scientific program, which documented the impressive progress of the community and defined future challenges in the field.     

A validated method for the quantification of enterodiol and enterolactone in plasma using isotope dilution liquid chromatography with tandem mass spectrometry

Enterolactone and enterodiol are phytoestrogens with structural similarity to endogenous estrogens. Because of their biological activities, they may affect the development of several diseases. To quantify enterodiol and enterolactone in plasma, we developed and validated a liquid chromatography-tandem mass spectrometry method with electrospray ionization using 13C3 labeled isotopes. The method consists of a simple enzymatic hydrolysis and ether extraction followed by a rapid LC separation (run-time of 11 min). Detection limits as low as 0.15 nM for enterodiol and 0.55 nM for enterolactone were achieved. The within-run R.S.D. ranges from 3 to 6% and the between-run R.S.D. ranges from 10 to 14% for both enterolignans. This method allows simple, rapid, and sensitive quantification, and is suitable for measuring large numbers of samples.     

Competitive strategies in adult beech and spruce: space-related foliar carbon investment versus carbon gain

In Central Europe, Fagus sylvatica and Picea abies represent contrasting extremes in foliage type, crown structure and length of growing season. In order to examine the competitive strategies of these two co-occurring species, we tested the following hypotheses: (1) the space occupied by the foliage of sun branches is characterized by greater foliar mass investment compared to shade branches, (2) the carbon (C) gain per unit of occupied space is greater in sun than in shade branches, and (3) annual C and water costs of the foliage for sustaining the occupied space are low, wherever C gain per unit of occupied space is low. These were investigated in a mature forest in Southern Germany. The examination was based on the annual assessment of space-related resource investments and gains of the foliage. The foliated space around branches was regarded as the relevant volume with respect to aboveground resource availability. Occupied crown space per standing foliage mass was higher in shade compared to sun branches of beech, whereas no difference existed in crown volume per foliage mass between sun and shade branches of spruce (hypothesis 1 accepted for beech but rejected for spruce). However, beech occupied more space per foliage mass than spruce. The C gain per occupied crown volume was greater in sun than in shade branches (hypothesis 2 accepted) but did not differ between species. The amount of occupied space per respiratory and transpiratory costs did not differ between species or between sun and shade branches. In beech and spruce, the proportion of foliage investment in the annual C balance of sun and shade branches remained rather stable, whereas respiratory costs distinctly increased in shade foliage. Hence, shade branches were costly structures to occupy space, achieving only low and even negative C balances (rejection of hypothesis 3), which conflicts with the claimed C autonomy of branches. Our findings suggest that competitiveness is determined by the standing foliage mass and the annual branch volume increment rather than annual investments in foliage. Expressing competitiveness in terms of space-related resource investments versus returns, as demonstrated here, has the potential of promoting mechanistic understanding of plant–plant interactions.