Distal tubular segments of the rabbit kidney after adaptation to altered Na- and K-intake

Summary The baso-lateral cell-membrane area in kidney tubules appears to be associated with the capacity for electrolyte transport; in the rabbit, it decreases from the distal convoluted tubule (DCT-cells) over the connecting tubule (CNT-cells) to the cortical collecting duct (principal cells).Adaptation to low Na-, high K-intake changes this pattern: CNT-cells at the beginning of the connecting tubule have the highest membrane area, which decreases along the segment, but remains two-fold higher than in controls. Principal cells have a four-fold higher membrane area than in controls. Simultaneous treatment with the antimineralocorticoid canrenoate-K inhibits the structural changes in CNT-cells only in end-portions of the connecting tubule and in principal cells.After prolonged high Na-, low K-intake DCT-cells display a two-fold higher membrane area than controls, while CNT-cells and principal cells are not affected. Simultaneous treatment with DOCA does not affect the DCT-cells but provokes a moderate increase in membrane area in CNT-cells, and a 5.5-fold increase in principal cells.The data provide evidence that DCT-, CNT- and principal cells are functionally different cell types. The baso-lateral cell-membrane area, associated with electrolyte-transport capacity, appears to be influenced in DCT-cells mainly by Na-intake, in CNT-cells mainly by K-intake and in part also by mineralocorticoids, and in principal cells mainly by mineralocorticoids.Supported by SFB90, CARVAS, Heidelberg and Swiss National Science Foundation, Grant No. 3.900-0.79     

Structural adaptation to selective pressure for altered ligand specificity in the Pseudomonas aeruginosa amide receptor, amiC

The AmiC protein in Pseudomonas aeruginosa is the negative regulator and ligand receptor for an amide-inducible aliphatic amidase operon. In the wild-type PAC1 strain, amidase expression is induced by acetamide or lactamide, but not by butyramide. A mutant strain of P. aeruginosa, PAC181, was selected for its sensitivity to induction by butyramide. The molecular basis for the butyramide inducible phenotype of P.aeruginosa PAC181 has now been determined, and results from a Thr-->Asn mutation at position 106 in PAC181-AmiC. In the wild-type PAC1-AmiC protein this residue forms part of the side wall of the amide-binding pocket but does not interact with the acetamide ligand directly. In the crystal structure of PAC181-AmiC complexed with butyramide, the Thr-->Asn mutation increases the size of the ligand binding site such that the mutant protein is able to close into its 'on' configuration even in the presence of butyramide. Although the mutation allows butyramide to be recognized as an inducer of amidase expression, the mutation is structurally sub-optimal, and produces a significant decrease in the stability of the mutant protein.     

Structural adaptation of enzymes to low temperatures

A systematic comparative analysis of 21 psychrophilic enzymes belonging to different structural families from prokaryotic and eukaryotic organisms is reported. The sequences of these enzymes were multiply aligned to 427 homologous proteins from mesophiles and thermophiles. The net flux of amino acid exchanges from meso/thermophilic to psychrophilic enzymes was measured. To assign the observed preferred exchanges to different structural environments, such as secondary structure, solvent accessibility and subunit interfaces, homology modeling was utilized to predict the secondary structure and accessibility of amino acid residues for the psychrophilic enzymes for which no experimental three-dimensional structure is available. Our results show a clear tendency for the charged residues Arg and Glu to be replaced at exposed sites on alpha-helices by Lys and Ala, respectively, in the direction from 'hot' to 'cold' enzymes. Val is replaced by Ala at buried regions in alpha-helices. Compositional analysis of psychrophilic enzymes shows a significant increase in Ala and Asn and a decrease in Arg at exposed sites. Buried sites in beta-strands tend to be depleted of VAL: Possible implications of the observed structural variations for protein stability and engineering are discussed.     

Intraflagellar transport delivers tubulin isotypes to sensory cilium middle and distal segments

Sensory cilia are assembled and maintained by kinesin-2-dependent intraflagellar transport (IFT). We investigated whether two Caenorhabditis elegans α- and β-tubulin isotypes, identified through mutants that lack their cilium distal segments, are delivered to their assembly sites by IFT. Mutations in conserved residues in both tubulins destabilize distal singlet microtubules. One isotype, TBB-4, assembles into microtubules at the tips of the axoneme core and distal segments, where the microtubule tip tracker EB1 is found, and localizes all along the cilium, whereas the other, TBA-5, concentrates in distal singlets. IFT assays, fluorescence recovery after photobleaching analysis and modelling indicate that the continual transport of sub-stoichiometric numbers of these tubulin subunits by the IFT machinery can maintain sensory cilia at their steady-state length.     

Structural basis for adaptation of lactobacilli to gastrointestinal mucus

The mucus layer covering the gastrointestinal (GI) epithelium is critical in selecting and maintaining homeostatic interactions with our gut bacteria. However, the underpinning mechanisms of these interactions are not understood. Here, we provide structural and functional insights into the canonical mucus‐binding protein (MUB), a multi‐repeat cell‐surface adhesin found in Lactobacillus inhabitants of the GI tract. X‐ray crystallography together with small‐angle X‐ray scattering demonstrated a ‘beads on a string’ arrangement of repeats, generating 174 nm long protein fibrils, as shown by atomic force microscopy. Each repeat consists of tandemly arranged Ig‐ and mucin‐binding protein (MucBP) modules. The binding of full‐length MUB was confined to mucus via multiple interactions involving terminal sialylated mucin glycans. While individual MUB domains showed structural similarity to fimbrial proteins from Gram‐positive pathogens, the particular organization of MUB provides a structural explanation for the mechanisms in which lactobacilli have adapted to their host niche by maximizing interactions with the mucus receptors, potentiating the retention of bacteria within the mucus layer. Together, this study reveals functional and structural features which may affect tropism of microbes across mucus and along the GI tract, providing unique insights into the mechanisms adopted by commensals and probiotics to adapt to the mucosal environment.     

Structural and mechanical adaptation of immature bone to strenuous exercise

To investigate the adaptive responses of immature bone to increased loads, young (3-wk-old) White Leghorn roosters were subjected to moderately intense treadmill running for 5 or 9 wk. The training program induced significant increases in maximal O2 consumption and muscle fumarase activity in the 12-wk-old birds, demonstrating that growing chickens have the ability to enhance their aerobic capacity. The structural and mechanical properties of the runners' tarsometatarsus bones were compared with sedentary age-matched controls at 8 and 12 wk of age. Suppression of circumferential growth occurred with exercise at both ages, whereas exercise enhanced middiaphysial cortical thickening, especially on the bones' concave surfaces. Although cross-sectional area moments of inertia did not change with exercise, significant decreases in bending stiffness, energy to yield, and energy to fracture were observed. It was concluded that strenuous exercise may retard long-bone maturation, resulting in more compliant bones.     

Structural aspects of the adaptation of Nostoc muscorum to salt

The physiological and biochemical changes during the adaptation of Nostoc muscorum to salt are accompanied by specific structural changes. Cells of Nostoc muscorum exposed to saline medium vary in size and envelope organization. There are also drastic changes in the intracellular organization of the thylakoidal assembly. The heterocysts exhibit a preferential tolerance to NaCl rather than mannitol. These findings suggest that Nostoc muscorum is equipped with a specific physiological capacity for NaCl tolerance.     

Carbohydrate and lipid metabolism in rats during adaptation to cold

Cold adapted rats are shown to have glucose and fatty acids concentration in blood inchanged, lactate concentration increased and triglyceride concentration decreased against the control level. Glucose utilization rate in the tolerance test grows. Glycogen content falls, hexokinase and succinate dehydrogenase activity increases, glucose-6-phosphatase and NAD+-isocytratedehydrogenase activity decreases in the liver of experimental animals. The results indicate that utilization of carbohydrate and lipid substrates for thermogenesis is intensified in cold-adapted rats. The hypothesis is supported by the data of tests dealing with IPNA injection or with bringing the animals back under thermocomfortable conditions.     

Differential regulation of ROMK (Kir1.1) in distal nephron segments by dietary potassium

ROMK channels are well-known to play a central role in renal K secretion, but the absence of highly specific and avid-ROMK antibodies has presented significant roadblocks toward mapping the extent of expression along the entire distal nephron and determining whether surface density of these channels is regulated in response to physiological stimuli. Here, we prepared new ROMK antibodies verified to be highly specific, using ROMK knockout mice as a control. Characterization with segmental markers revealed a more extensive pattern of ROMK expression along the entire distal nephron than previously thought, localizing to distal convoluted tubule regions, DCT1 and DCT2; the connecting tubule (CNT); and cortical collecting duct (CD). ROMK was diffusely distributed in intracellular compartments and at the apical membrane of each tubular region. Apical labeling was significantly increased by high-K diet in DCT2, CNT1, CNT2, and CD (P < 0.05) but not in DCT1. Consistent with the large increase in apical ROMK, dramatically increased mature glycosylation was observed following dietary potassium augmentation. We conclude 1) our new antibody provides a unique tool to characterize ROMK channel localization and expression and 2) high-K diet causes a large increase in apical expression of ROMK in DCT2, CNT, and CD but not in DCT1, indicating that different regulatory mechanisms are involved in K diet-regulated ROMK channel functions in the distal nephron.     

The metabolism of social subterranean rodents: adaptation to aridity

Summary The social Damara mole-rat Cryptomys damarensis (124 g), has a mean (±SD) resting metabolic rate of 0.57±0.09 cm³ O₂ g^-1 h^-1, within a thermoneutral zone of 27–31° C. This rate of metabolism is 43% lower than that predicted by the curve for rodents, and 29% lower than that predicted by the subterranean rodent curve. These data support the hypothesis that the resting metabolic rates of social and solitary subterranean rodents are lower than those of solitary species inhabiting mesic habitats. These low resting metabolic rates may represent an energy-saving adaptation to aridity. The energetic cost of burrowing, in relation to the dispersion patterns of food in arid habitats, may explain these low metabolic rates.     

Contributions of working muscle to whole body lipid metabolism are altered by exercise intensity and training

To evaluate the contribution of working muscle to whole body lipid oxidation, we examined the effects of exercise intensity and endurance training (9 wk, 5 days/wk, 1 h, 75% Vo(2 peak)) on whole body and leg free fatty acid (FFA) kinetics in eight male subjects (26 +/- 1 yr, means +/- SE). Two pretraining trials [45 and 65% Vo(2 max) (45UT, 65UT)] and two posttraining trials [65% of pretraining Vo(2 peak) (ABT), and 65% of posttraining Vo(2 peak) (RLT)] were performed using [1-(13)C]palmitate infusion and femoral arteriovenous sampling. Training increased Vo(2 peak) by 15% (45.2 +/- 1.2 to 52.0 +/- 1.8 ml.kg(-1).min(-1), P < 0.05). Muscle FFA fractional extraction was lower during exercise (EX) compared with rest regardless of workload or training status ( approximately 20 vs. 48%, P < 0.05). Two-leg net FFA balance increased from net release at rest ( approximately -36 micromol/min) to net uptake during EX for 45UT (179 +/- 75), ABT (236 +/- 63), and RLT (136 +/- 110) (P < 0.05), but not 65UT (51 +/- 127). Leg FFA tracer measured uptake was higher during EX than rest for all trials and greater during posttraining in RLT (716 +/- 173 micromol/min) compared with pretraining (45UT 450 +/- 80, 65UT 461 +/- 72, P < 0.05). Leg muscle lipid oxidation increased with training in ABT (730 +/- 163 micromol/min) vs. 65UT (187 +/- 94, P < 0.05). Leg muscle lipid oxidation represented approximately 62 and 30% of whole body lipid oxidation at lower and higher relative intensities, respectively. In summary, training can increase working muscle tracer measured FFA uptake and lipid oxidation for a given power output, but both before and after training the association between whole body and leg lipid metabolism is reduced as exercise intensity increases.