The hydraulic conductance of the angiosperm leaf lamina: a comparison of three measurement methods

A comparison was made of three methods for measuring the leaf lamina hydraulic conductance (K(lamina)) for detached mature leaves of six woody temperate angiosperm species. The high-pressure method, the evaporative flux method and the vacuum pump method involve, respectively, pushing, evaporating and pulling water out of the lamina while determining the flow rate into the petiole and the water potential drop across the leaf. Tests were made of whether the high-pressure method and vacuum pump method measurements of K(lamina) on single leaves were affected by irradiance. In Quercus rubra, the high pressure method was sensitive to irradiance; K(lamina) measured under high irradiance (>1200 micro mol m(-2) s(-1 )photosynthetically active radiation) was 4.6-8.8 times larger than under ambient laboratory lighting (approximately 6 micro mol m(-2) s(-1 )photosynthetically active radiation). By constrast, the vacuum pump method was theoretically expected to be insensitive to irradiance, and this expectation was confirmed in experiments on Hedera helix. When used in the ways recommended here, the three methods produced measurements that agreed typically within 10%. There were significant differences in species' K(lamina); values ranged from 1.24x10(-4) kg s(-1) m(-2) MPa(-1) for Acer saccharum to 2.89x10(-4) kg s(-1) m(-2) MPa(-1) for Vitis labrusca. Accurate, rapid determination of K(lamina) will allow testing of the links between K(lamina), water-use, drought tolerance, and the enormous diversity of leaf form, structure and composition.     

Structural analysis of ligand‐bound states of the Salmonella type III secretion system ATPase InvC

Translocation of virulence effector proteins through the type III secretion system (T3SS) is essential for the virulence of many medically relevant Gram‐negative bacteria. The T3SS ATPases are conserved components that specifically recognize chaperone–effector complexes and energize effector secretion through the system. It is thought that functional T3SS ATPases assemble into a cylindrical structure maintained by their N‐terminal domains. Using size‐exclusion chromatography coupled to multi‐angle light scattering and native mass spectrometry, we show that in the absence of the N‐terminal oligomerization domain the Salmonella T3SS ATPase InvC can form monomers and dimers in solution. We also present for the first time a 2.05 å resolution crystal structure of InvC lacking the oligomerization domain (InvCΔ79) and map the amino acids suggested for ATPase intersubunit interaction, binding to other T3SS proteins and chaperone–effector recognition. Furthermore, we validate the InvC ATP‐binding site by co‐crystallization of InvCΔ79 with ATPγS (2.65 å) and ADP (2.80 å). Upon ATP‐analogue recognition, these structures reveal remodeling of the ATP‐binding site and conformational changes of two loops located outside of the catalytic site. Both loops face the central pore of the predicted InvC cylinder and are essential for the function of the T3SS ATPase. Our results present a fine functional and structural correlation of InvC and provide further details of the homo‐oligomerization process and ATP‐dependent conformational changes underlying the T3SS ATPase activity.     

Poly(ADP-ribose) polymerase 1 (PARP-1) binds to 8-oxoguanine-DNA glycosylase (OGG1)

Human 8-oxoguanine-DNA glycosylase (OGG1) plays a major role in the base excision repair pathway by removing 8-oxoguanine base lesions generated by reactive oxygen species. Here we report a novel interaction between OGG1 and Poly(ADP-ribose) polymerase 1 (PARP-1), a DNA-damage sensor protein involved in DNA repair and many other cellular processes. We found that OGG1 binds directly to PARP-1 through the N-terminal region of OGG1, and this interaction is enhanced by oxidative stress. Furthermore, OGG1 binds to PARP-1 through its BRCA1 C-terminal (BRCT) domain. OGG1 stimulated the poly(ADP-ribosyl)ation activity of PARP-1, whereas decreased poly(ADP-ribose) levels were observed in OGG1(-/-) cells compared with wild-type cells in response to DNA damage. Importantly, activated PARP-1 inhibits OGG1. Although the OGG1 polymorphic variant proteins R229Q and S326C bind to PARP-1, these proteins were defective in activating PARP-1. Furthermore, OGG1(-/-) cells were more sensitive to PARP inhibitors alone or in combination with a DNA-damaging agent. These findings indicate that OGG1 binding to PARP-1 plays a functional role in the repair of oxidative DNA damage.     

THE RELATIONSHIP BETWEEN MATING-SYSTEM CHARACTERS AND INBREEDING DEPRESSION IN MIMULUS GUTTATUS

Several recent theoretical considerations of mating-system evolution predict within-population covariation between levels of inbreeding depression and genetically controlled mating-system characters. If inbreeding depression is caused by deleterious recessive alleles, families with characters that promote self-pollination should show lower levels of inbreeding depression than families with characters that promote outcrossing. The converse is expected if inbreeding depression is due to overdominant allelic interactions. Whether these associations between mating-system and viability loci evolve will have important consequences for mating-system evolution. The evolution of selfing within the genus Mimulus is associated with a reduction in stigma-anther separation (i.e., a loss of herkogamy) and high autogamous seed set. In this study we compared families from two M. guttatus populations that differed genetically in their degree of stigma-anther separation. In one of these populations we also compared families that differed genetically in the degree to which they autogamously set seed in a pollinator-free greenhouse. Dams often differed significantly in levels of inbreeding depression for aboveground biomass and flower production, but variation in inbreeding depression was never explained by herkogamy class or autogamy class. Several factors might account for why families with traits associated with selfing did not show lower inbreeding depression, and these are discussed. Our study also demonstrated significant variation among self progeny from a given female likely due to differences in pollination date and position of fruit maturation. The detection of significant dam × sire interactions suggests biparental inbreeding or differences in combining ability for specific pairs of parents.     

Validity of Gō models: comparison with a solvent-shielded empirical energy decomposition

Do Gō-type model potentials provide a valid approach for studying protein folding? They have been widely used for this purpose because of their simplicity and the speed of simulations based on their use. The essential assumption in such models is that only contact interactions existing in the native state determine the energy surface of a polypeptide chain, even for non-native configurations sampled along folding trajectories. Here we use an all-atom molecular mechanics energy function to investigate the adequacy of Gō-type potentials. We show that, although the contact approximation is accurate, non-native contributions to the energy can be significant. The assumed relation between residue-residue interaction energies and the number of contacts between them is found to be only approximate. By contrast, individual residue energies correlate very well with the number of contacts. The results demonstrate that models based on the latter should give meaningful results (e.g., as used to interpret phi values), whereas those that depend on the former are only qualitative, at best.     

Basolateral Ca2+-dependent K+-channels play a key role in Cl- secretion induced by taurodeoxycholate from colon mucosa

The diarrhea associated with malabsorption of bile salts such as the secondary hydrophobic taurodeoxycholate (TDC) may be partly explained by the TDC-induced increase in colon Cl(-) secretion. We, therefore, investigated the effects of TDC (0.5-8 mM) on electrical parameters and electrolyte transport of rat proximal colon mucosa mounted in Ussing chambers. Colonic secretion, measured as short circuit current (I(SC)), progressively increased on mucosal incubation with TDC ranging 0.5-2 mM; up to TDC 2 mM, a spontaneous recovery toward control values with no changes in epithelial resistance (Rt), and lactate dehydrogenase (LDH) release was observed. In contrast, for TDC > 2 mM, I(SC) increased further and the effect was progressive and associated with a significant decrease in the Rt and increased LDH release, implying a cytolytic effect. Mucosal preincubation with the Cl(-) channel inhibitor 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB), fully prevented the precytolytic effect of TDC on I(SC). Serosal preincubation with furosemide, a Na(+)/K(+)/2Cl(-) cotransporter inhibitor, significantly reduced TDC-induced increase in I(SC). Inhibition of the basolateral Ca(2+)-dependent K(+) channel-rSK4-with serosal clotrimazole or incubation with mucosal Ca(2+)-free (EGTA) buffer completely prevented precytolytic TDC-induced increase in I(SC). In conclusion, Cl(-) secretion is activated in colon mucosa by TDC low concentrations; while at higher concentrations, a detergent cytotoxic effect intervenes. Activation of the Ca(2+)-dependent basolateral K(+) pathway, through TDC-induced apical Ca(2+) influx, provides the Na(+)/K(+)/2Cl(-) basolateral activation, thereby the driving force for the apical exit of Cl(-) ions. These findings further enhance the knowledge of the pathogenic mechanisms of diarrhea associated with bile salt malabsorption.     

ST7 is a novel low-density lipoprotein receptor-related protein (LRP) with a cytoplasmic tail that interacts with proteins related to signal transduction pathways

In 1997, McCormick and co-workers identified a novel putative tumor suppressor gene, designated ST7, encoding a unique protein with transmembrane receptor characteristics [Qing et al. (1999) Oncogene 18, 335-342]. Using degenerate primers corresponding to the highly conserved region of the ligand-binding domains of members of the low-density lipoprotein receptor (LDLR) superfamily, Ishii et al. [Genomics (1998) 51, 132-135] discovered a low-density lipoprotein receptor-related protein (LRP) that closely resembles ST7. Later, another LRP closely resembling ST7 and LRP3 was found (murine LRP9) [Sugiyama et al. (2000) Biochemistry 39, 15817-15825]. These results strongly suggested that ST7 was also a novel member of the low-density lipoprotein receptor superfamily. Proteins of this superfamily have been shown to function in endocytosis and/or signal transduction. To evaluate the relationship of ST7 to the LDLR superfamily proteins and to determine whether ST7 may function in endocytosis and/or signal transduction, we used proteomic tools to analyze the functional motifs present in the protein. Our results indicate that ST7 is a member of a subfamily of the LDLR superfamily and that its cytoplasmic domain contains several motifs implicated in endocytosis and signal transduction. Use of the yeast two-hybrid system to identify proteins that associate with ST7's cytoplasmic domain revealed that this domain interacts with three proteins involved in signal transduction and/or endocytosis, viz., receptor for activated protein C kinase 1 (RACK1), muscle integrin binding protein (MIBP), and SMAD anchor for receptor activation (SARA), suggesting that ST7, like other proteins in the LDLR superfamily, functions in these two pathways. Clearly, ST7 is an LRP, and therefore, it should now be referred to as LRP12.     

The response of cortical neurons to in vivo-like input current: theory and experiment: II. Time-varying and spatially distributed inputs

The response of a population of neurons to time-varying synaptic inputs can show a rich phenomenology, hardly predictable from the dynamical properties of the membrane’s inherent time constants. For example, a network of neurons in a state of spontaneous activity can respond significantly more rapidly than each single neuron taken individually. Under the assumption that the statistics of the synaptic input is the same for a population of similarly behaving neurons (mean field approximation), it is possible to greatly simplify the study of neural circuits, both in the case in which the statistics of the input are stationary (reviewed in La Camera et al. in Biol Cybern, 2008) and in the case in which they are time varying and unevenly distributed over the dendritic tree. Here, we review theoretical and experimental results on the single-neuron properties that are relevant for the dynamical collective behavior of a population of neurons. We focus on the response of integrate-and-fire neurons and real cortical neurons to long-lasting, noisy, in vivo-like stationary inputs and show how the theory can predict the observed rhythmic activity of cultures of neurons. We then show how cortical neurons adapt on multiple time scales in response to input with stationary statistics in vitro. Next, we review how it is possible to study the general response properties of a neural circuit to time-varying inputs by estimating the response of single neurons to noisy sinusoidal currents. Finally, we address the dendrite–soma interactions in cortical neurons leading to gain modulation and spike bursts, and show how these effects can be captured by a two-compartment integrate-and-fire neuron. Most of the experimental results reviewed in this article have been successfully reproduced by simple integrate-and-fire model neurons.