Reduced cross-bridge dependent stiffness of skinned myocardium from mice lacking cardiac myosin binding protein-C

The role of cardiac myosin binding protein-C (MyBP-C) on myocardial stiffness was examined in skinned papillary muscles of wild-type (WT^+/+) and homozygous truncated cardiac MyBP-C (MyBP-C^t/t) male mice. No MyBP-C was detected by gel electrophoresis or by Western blots in the MyBP-C^t/t myocardium. Rigor-bridge dependent myofilament stiffness, i.e., rigor minus relaxed stiffness, in the MyBP-C^t/t myocardium (281 ± 44 kN/m²) was 44% that in WT^+/+ (633 ± 141 kN/m²). The center-to-center spacing between thick filaments as determined by X-ray diffraction in MyBP-C^t/t (45.0 ± 1.2 nm) was not significantly different from that in WT^+/+ (43.2 ± 0.9 nm). The fraction of cross-sectional area comprised of myofibrils, as determined by electron microscopy, was reduced in the MyBP-C^t/t (39.9%) by 10% compared to WT^+/+ (44.5%). These data suggest that the 56% reduction in rigor-bridge dependent stiffness of the skinned MyBP-C^t/t myocardium could not be due solely to a 10% reduction in the number of thick filaments per cross-sectional area and must also be due to approximately 50% reduction in the stiffness of the rigor-bridge attached thick filaments lacking MyBP-C. (Mol Cell Biochem 263: 73–80, 2004)     

The effect of wind, season and latitude on the migration speed of white storks Ciconia ciconia, along the eastern migration route

The relation between wind, latitude and daily migration speed along the entire migration route of white storks was analysed. Mean daily migration speed was calculated using satellite telemetry data for autumn and spring migration of white storks from their breeding grounds in Germany and Poland to wintering grounds in Africa and back. The National Center for Environmental Prediction (NCEP) reanalysis data were used to systematically fit 850 mb wind vectors to daily migration speed along the migration route. White storks migrated significantly faster and had a shorter migration season in autumn (10 km/h) compared to spring (6.4 km/h). In autumn mean daily migration speed was significantly slower in Europe (8.0 km/h) than in the Middle East (11.1 km/h) and Africa (11.0 km/h). In spring mean daily migration speed was significantly faster in Africa (10.5 km/h) as birds left their wintering grounds than in the Middle East (4.3 km/h). Migration speed then increased in Europe (6.5 km/h) as birds approached their breeding grounds. In both spring and autumn tailwind (at 850mb) and latitude were found to be significant variables related to daily migration speed.     

Coupling of Mitochondrial Fatty Acid Uptake to Oxidative Flux in the Intact Heart

The coordination of long chain fatty acid (LCFA) transport across the mitochondrial membrane (V_PAL) with subsequent oxidation rate through β-oxidation and the tricarboxylic acid (TCA) cycle (V_tca) has been difficult to characterize in the intact heart. Kinetic analysis of dynamic ¹³C-NMR distinguished these flux rates in isolated rabbit hearts. Hearts were perfused in a 9.4 T magnet with either 0.5 mM [2,4,6,8,10,12,14,16-¹³C₈] palmitate (n = 4), or 0.5 mM ¹³C-labeled palmitate plus 0.08 mM unlabeled butyrate (n = 4). Butyrate is a short chain fatty acid (SCFA) that bypasses the LCFA transporters of mitochondria. In hearts oxidizing palmitate alone, the ratio of V_TCA to V_PAL was 8:1. This is consistent with one molecule of palmitate yielding eight molecules of acetyl-CoA for the subsequent oxidation through the TCA cycle. Addition of butyrate elevated this ratio; V_TCA/V_PAL = 12:1 due to an SCFA-induced increase in V_TCA of 43% (p < 0.05). However, SCFA oxidation did not significantly reduce palmitate transport into the mitochondria: V_PAL = 1.0 ± 0.2 μmol/min/g dw with palmitate alone versus 0.9 ± 0.1 with palmitate plus butyrate. Thus, the products of β-oxidation are preferentially channeled to the TCA cycle, away from mitochondrial efflux via carnitine acetyltransferase.     

Molecular and phenotypic effects of heterozygous, homozygous, and compound heterozygote myosin heavy-chain mutations

Autosomal dominant familial hypertrophic cardiomyopathy (FHC) has variable penetrance and phenotype. Heterozygous mutations in MYH7 encoding beta-myosin heavy chain are the most common causes of FHC, and we proposed that "enhanced" mutant actin-myosin function is the causative molecular abnormality. We have studied individuals from families in which members have two, one, or no mutant MYH7 alleles to examine for dose effects. In one family, a member homozygous for Lys207Gln had cardiomyopathy complicated by left ventricular dilatation, systolic impairment, atrial fibrillation, and defibrillator interventions. Only one of five heterozygous relatives had FHC. Leu908Val and Asp906Gly mutations were detected in a second family in which penetrance for Leu908Val heterozygotes was 46% (21/46) and 25% (3/12) for Asp906Gly. Despite the low penetrance, hypertrophy was severe in several heterozygotes. Two individuals with both mutations developed severe FHC. The velocities of actin translocation (V(actin)) by mutant and wild-type (WT) myosins were compared in the in vitro motility assay. Compared with WT/WT, V(actin) was 34% faster for WT/D906G and 21% for WT/L908V. Surprisingly V(actin) for Leu908Val/Asp906Gly and Lys207Gln/Lys207Gln mutants were similar to WT. The apparent enhancement of mechanical performance with mutant/WT myosin was not observed for mutant/mutant myosin. This suggests that V(actin) may be a poor predictor of disease penetrance or severity and that power production may be more appropriate, or that the limited availability of double mutant patients prohibits any definitive conclusions. Finally, severe FHC in heterozygous individuals can occur despite very low penetrance, suggesting these mutations alone are insufficient to cause FHC and that uncharacterized modifying mechanisms exert powerful influences.     

Root cooperation in a clonal plant: connected strawberries segregate roots

The ability to selectively avoid competition with members of the same clone should be highly advantageous but has not been demonstrated in plants. We found that physical connection between plants in a clone of the wild strawberry Fragaria chiloensis induced them to segregate their roots, significantly increasing clonal performance. Such increase in performance was not found when plants were grown in containers that artificially divided their rooting zones. There was no effect of connection in a different clone of F. chiloensis with a lower degree of carbon transport between connected plants, suggesting that the mechanism for root segregation depended upon transport of a signal through the strawberry runners. We suggest that clonal integration allows some clones to coordinate below-ground resource foraging with other clone members, thus exhibiting a type of root cooperation.     

COMPARATIVE PATTERNS OF NET PHOTOSYNTHESIS IN AN ASSEMBLAGE OF MOSSES WITH CONTRASTING MICRODISTRIBUTIONS

Effects of plant water content, previous desiccation, photon flux density, and temperature on rate of net photosynthesis were measured in the common species in the assemblage of desiccationtolerant mosses that dominates the flora on granitic boulders at a semiarid site in the inland chaparral of San Diego County, California. Species differed significantly in their responses to water content, desiccation, and light. Species previously found to occur in microsites of lower water availability attained maximum net photosynthesis at lower water contents and showed greater ability to recover from prolonged desiccation. Species found in microsites of lower light availability had higher rates of net photosynthesis at low photon flux density. Results suggest that differences in potential rates of net photosynthesis partly determine relative ability of these species to colonize xeric or dark microsites, but that other factors also determine microdistribution.     

The limits and frontiers of desiccation-tolerant life

Drying to equilibrium with the air is lethal to most speciesof animals and plants, making drought (i.e., low external waterpotential) a central problem for terrestrial life and a majorcause of agronomic failure and human famine. Surprisingly, awide taxonomic variety of animals, microbes, and plants do toleratecomplete desiccation, defined as water content below 0.1 g H₂Og^–1 dry mass. Species in five phyla of animals and fourdivisions of plants contain desiccation-tolerant adults, juveniles,seeds, or spores. There seem to be few inherent limits on desiccationtolerance, since tolerant organisms can survive extremely intenseand prolonged desiccation. There seems to be little phylogeneticlimitation of tolerance in plants but may be more in animals.Physical constraints may restrict tolerance of animals withoutrigid skeletons and to plants shorter than 3 m. Physiologicalconstraints on tolerance in plants may include control by hormoneswith multiple effects that could link tolerance to slow growth.Tolerance tends to be lower in organisms from wetter habitats,and there may be selection against tolerance when water availabilityis high. Our current knowledge of limits to tolerance suggeststhat they pose few obstacles to engineering tolerance in prokaryotesand in isolated cells and tissues, and there has already beenmuch success on this scientific frontier of desiccation tolerance.However, physical and physiological constraints and perhapsother limits may explain the lack of success in extending toleranceto whole, desiccation-sensitive, multicellular animals and plants.Deeper understanding of the limits to desiccation tolerancein living things may be needed to cross this next frontier.     

Reduced cross-bridge dependent stiffness of skinned myocardium from mice lacking cardiac myosin binding protein-C

The role of cardiac myosin binding protein-C (MyBP-C) on myocardial stiffness was examined in skinned papillary muscles of wild-type (WT(+/+)) and homozygous truncated cardiac MyBP-C (MyBP-C(t/t) male mice. No MyBP-C was detected by gel electrophoresis or by Western blots in the MyBP-C(t/t) myocardium. Rigor-bridge dependent myofilament stiffness, i.e., rigor minus relaxed stiffness, in the MyBP-C(t/t) myocardium (281 +/- 44 kN/m2) was 44% that in WT(+/+) (633 +/- 141 kN/m2). The center-to-center spacing between thick filaments as determined by X-ray diffraction in MyBP-C(t/t) (45.0 +/- 1.2 nm) was not significantly different from that in WT(+/+) (43.2 +/- 0.9 nm). The fraction of cross-sectional area comprised of myofibrils, as determined by electron microscopy, was reduced in the MyBP-C(t/t) (39.9%) by 10% compared to WT(+/+) (44.5%). These data suggest that the 56% reduction in rigor-bridge dependent stiffness of the skinned MyBP-C(t/t) myocardium could not be due solely to a 10% reduction in the number of thick filaments per cross-sectional area and must also be due to approximately 50% reduction in the stiffness of the rigor-bridge attached thick filaments lacking MyBP-C.     

Differential cross-bridge kinetics of FHC myosin mutations R403Q and R453C in heterozygous mouse myocardium

The kinetic effects of the cardiac myosin point mutations R403Q and R453C, which underlie lethal forms of familial hypertrophic cardiomyopathy (FHC), were assessed using isolated myosin and skinned strips taken from heterozygous (R403Q/+ and R453C/+) male mouse hearts. Compared with wild-type (WT) mice, actin-activated ATPase was increased by 38% in R403Q/+ and reduced by 45% in R453C/+, maximal velocity of regulated thin filament (V(RTF)) in the in vitro motility assay was increased by 8% in R403Q/+ and was not different in R453C/+, myosin concentration at half-maximal V(RTF) was reduced by 30% in R403Q/+ and not different in R453C/+, and the characteristic frequency for oscillatory work production (b frequency), determined by sinusoidal analysis in the skinned strip at maximal calcium activation, was 27% lower in R403Q/+ and 18% higher in R453C/+. The calcium sensitivity for isometric tension in the skinned strip was not different in R403Q/+ (pCa(50) 5.64 +/- 0.02) and significantly enhanced in R453C/+ (5.82 +/- 0.03) compared with WT (5.58 +/- 0.02). We conclude that isolated myosin and skinned strips of R403Q/+ and R453C/+ myocardium show marked differences in cross-bridge kinetic parameters and in calcium sensitivity of force production that indicate different functional roles associated with the location of each point mutation at the molecular level.