Separation of mitochondria from microbodies of Pisum sativum (L. cv. Alaska) cotyledons

A method is described for separating mitochondria from microbodies in cotyledon preparations of Pisum sativum L. cv. Alaska. Pure and intact mitochondria were obtained on a continuous: discontinuous sucrose density gradient as shown by marke-enzyme assay and electron microscopy. Manipulation of sucrose-gradient construction to widen the distance between organelles provided a quick method for the separation of the mitochondria from the microbodies. The shorter time of exposure of mitochondria to centrifugation and osmotic stress produces mitochondria free of contamination.     

Resistance to variable-time schedules produced by spaced-response reinforcement schedules

Two experiments are reported in which rats were exposed to either a tandem fixed-ratio 1 differential-reinforcement-of-low-rate 10 sec schedule (tand FR1 DRL 10) (Experiment 1) or to a DRL 10 sec schedule (Experiment 2) prior to being exposed to a variable-time (VT) schedule. Response decrement was not universally found during the VT phase, one rat emmitting an increase in response rate relative to baseline (Experiment 1), while another showed neither an increase nor decrease during VT relative to baseline (Experiment2). The VT schedule induced less efficient responding in both experiments. Interresponse time (IRT) distributions (Experiment 2) indicated that the VT changed the pattern of responding. These results, when combined with others from our laboratory, indicate that although unsignalled variable delay of reinforcement may be a sufficient condition for producing resistance to response-independent reinforcement, it is not a necessary condition. It was concluded that a response competition model of the kind proposed by Henton and Iversen (1978) might have explanatory merit in this kind of experimental situation.     

Acute and definitive management of traumatic osteocutaneous defects of the lower extremity

Twenty-two lower extremity osteocutaneous defects resulting from high-energy trauma were managed from the onset of injury to rehabilitation by a collaborative effort between orthopedic and plastic surgeons. Emergency debridement of devitalized soft tissue and bone, external fracture stabilization, and serial debridements prepared the wound for closure with predominantly free-muscle transfers performed an average of 17 days (range 3 to 43 days) after injury. Cancellous or vascularized fibula grafting, depending on defect size, was performed an average of 9 weeks (range 6 to 16 weeks) after muscle flap closure. In this group of patients, whose average injury severity score was 18 (range 9 to 45) and whose average segmental bone defect was 8 cm (range 3 to 18 cm), the average time after injury to full weight bearing was 61 weeks (range 39 to 120 weeks). The early infection rate was 14 percent. Two extremities were amputated. There have been no chronic infections. Follow-up has ranged from 9 to 34 months.     

Isolation and characterization of platelet glycoprotein IV (CD36)

Platelet glycoprotein IV (GPIV, Mr 88,000), which is immunologically related to the leukocyte differentiation antigen CD36, has been isolated from both intact and trypsinized platelet membranes by a series of steps involving (i) phase partitioning in Triton X-114, (ii) ion exchange chromatography on DEAE-cellulose, (iii) lectin affinity chromatography on wheat germ agglutinin-Sepharose, and (iv) size exclusion chromatography on Ultrogel AcA-44. The homogenous product contained 26% carbohydrate (sialic acid, Gal, Man, GalNAc, GlcNAc), of which approximately two-thirds were in alkali-labile O-glycosidic linkages. A rabbit polyclonal antibody raised against purified GPIV gave a single band on immunoblot and on immunoprecipitation from solubilized, 3H-labeled platelet membranes indicating its monospecificity. The antibody gave a strongly positive reaction with platelets on flow cytofluorimetry further confirming the surface location of GPIV. Immunoblotting and flow cytometry also identified GPIV-like molecules on the surface of U937, HEL, and C32 cells but not on HT1080 fibroblasts. Amino acid analysis showed values comparable with those deduced from the cloning data for GPIb, GPIIb, and GPIIIa. Automated Edman degradation allowed the identification of the sequence of the first 36 residues of the NH2-terminal domain. G-X-D-R-N-X-G-L-I-A-G-A-V-I-G-A-V- L-A-V-F-G-G-I-L-M-P-V-G-D-L-P-X-Q-K-F. There are no identifiable homologies between the NH2-terminal domain and other known protein sequences. Following a hydrophilic hexapeptide, there is a hydrophobic sequence of 23 amino acids (underlined) that is of the size and composition expected for a transmembrane domain. Since the NH2-terminal domain lacks tyrosine, but GPIV may be readily iodinated in intact platelets, this suggests that GPIV may have a configuration expressing other extramembranous domains.     

Perceptions of effort and heaviness during fatigue and during the size-weight illusion

Previous work has shown that force perception and the sense of motor effort are different attributes of sensorimotor function. This study explores the hypothesis that one reason force and effort perceptions are distinct is to inform an individual of impaired motor function when muscular force lags effort. This hypothesis predicts that effort and force perceptions will dissociate when motor function is impaired by fatigue but not during the size-weight illusion. All subjects reported a distinct increase in effort when lifting a standard test weight as fatigue developed. When fatigue was sufficiently marked so that they could barely lift the test weight, they rated their effort as similar to that required to lift a maximal weight in the unfatigued state. The perceived heaviness of the test weight also increased as fatigue developed, but this fatigue-weight illusion was smaller than the increase in effort for all subjects and displayed greater variability. In contrast, both the perceived weight of a small object and the effort required to lift it increased in parallel when small and large objects were lifted sequentially. The size-weight and size-effort illusions appear to be examples of a common phenomenon in which perceptual experience is rescaled to maintain acuity under different working conditions. The fatigue-weight illusion also has the effect of increasing perceptual acuity as the subject's weight lifting range decreases due to fatigue.     

Studies on the growth and development of protoplasts of the moss,Physcomitrella patens,and its control by light

Protoplasts prepared from protonemal cultures of the moss Physcomitrella patens begin to regenerate a new cell wall within 1 h of removal from cellulase. The wall is seen as a gradually thickening mat of fibres when examined by scanning electron microscopy. Development of filaments from protoplasts takes place in the majority of cases only after one or more cell divisions have occurred. The direction of emergence of filaments is random in uniform light, but strongly negatively phototropic in bright unidirectional horizotal light. Filament growth is also strongly negatively phototropic. The influence of unidirectional light can be destroyed by incubating protoplasts in the presence of colchicine. Filaments growing in unidirectional light have cytoplasmic microtubules running along their long axes and in close association with large organelles. These results are discussed in terms of the potential for this system for the study of polarity in plants.     

Fluorine-19 chemical shifts as probes of the structure and reactivity of the iron-molybdenum cofactor of nitrogenase

The reaction of the iron-molybdenum cofactor with thiolate and the redox behavior of the iron-molybdenum cofactor-thiolate complex have been studied by 19F NMR using p-CF3C6H4S- as the reporter ligand. These experiments give results different from those produced by other methods which have been performed near 4 K rather than at ambient temperature. Specifically, these data show that the iron-molybdenum cofactor-thiolate complex is not the product of an irreversible reaction. Rather, the complex is in dynamic equilibrium with the free iron-molybdenum cofactor and free thiolate. Models of the reactions of nitrogenase may need to take this temperature-dependent difference into account because the lability of the iron-molybdenum thiolate bond means its making and breaking could be involved in substrate binding or reduction. The 19F NMR results reported here also show that the S = 3/2 state of the iron-molybdenum cofactor-thiolate complex can be easily and reversibly oxidized by one electron. However, electron exchange between the oxidized and reduced states of the complex is quite slow at approximately 1 mM. Based on low temperature spectroscopic studies, the oxidized iron-molybdenum cofactor-thiolate complex was expected to be diamagnetic. Isotropically shifted NMR spectra of the oxidized cofactor samples at 240-320 K, however, indicate at least partial population of a paramagnetic state, possibly with S = 1.     

Introns and reading frames: correlation between splicing sites and their codon positions

Computer analyses of the entire GenBank database were conducted to examine correlation between splicing sites and codon positions in reading frames. Intron insertion patterns (i.e., splicing site locations with respect to codon positions) have been analyzed for all of the 74 codons of all the eukaryote taxonomic groups: primates, rodents mammals, vertebrates, invertebrates, and plants. We found that reading frames are interrupted by an intron at a codon boundary (as opposed to the middle of a codon) significantly more often than expected. This observation is consistent with the exon shuffling hypothesis, because exons that end at codon boundaries can be concatenated without causing a frame shift and thus are evolutionarily advantageous. On the other hand, when introns interrupt at the middles of codons, they exist in between the first and second bases much more frequently than between the second and third bases, despite the fact that boundaries between the first and second bases of codons are generally far more important than those between the second and third bases. The reason for this is not clear and yet to be explained. We also show that the length of an exon is a multiple of 3 more frequently than expected. Furthermore, the total length of two consecutive exons is also more frequently a multiple of 3. All the observations above are consistent with results recently published by Long, Rosenberg, and Gilbert (1995).      

Can hydraulic redistribution put bread on our table?

Hydraulic redistribution is the process where soil water is translocated by plant roots from wet to dry areas as it is drawn through xylem pathways by a water potential gradient. Hydraulic redistribution places soil water resources where they would otherwise not be, which results in a range of ecological and hydrological consequences. Although deep-rooted plants can transfer water up from depth into shallow soil layers, any localised ??irrigation?? of neighbouring plants tends to be obscured by recovery of the very same water by the donor plants during daytime transpiration. A new intercropping system was recently trialled which eliminates transpiration by the donor plant through complete shoot removal in order to maximise hydraulic redistribution. In the absence of any transpiring shoots, the donor plants are left to wick water up from depth 24 hours a day via their root systems, to the benefit of neighbouring shallow-rooted crops. This system allows deeper-rooted ??nurse plants?? to capture water that is out of reach of crops in a ??water safety-net?? role, which may be of considerable benefit in water-scarce environments.