Entire Rod Research Articles

Functional phycobilisome core structures in a phycocyanin-less mutant of cyanobacterium Synechococcus sp. PCC 7942

We have constructed a mutant Synechococcus sp. PCC 7942, termed R2HECAT, in which the entire phycobilisome rod operon has been deleted. In the whole cell absorption spectra of R2HECAT, the peak corresponding to phycocyanin (PC), λmax≈620 nm, could not be detected. However, a single pigment-protein fraction with λmax=654 nm could be isolated on sucrose gradients from R2HECAT. Analysis of this pigment-protein fraction by non-denaturing PAGE indicates an apparent molecular mass of about 1200-1300 kDa. On exposure to low temperature, the isolated pigment-protein complex dissociated to a protein complex with a molecular mass of about 560 kDa. When analysed by SDS-PAGE, the pigment-protein fraction was found to consist of the core polypeptides but lacked PC, 27, 33, 30, and the 9 kDa polypeptides which are a part of the rods. All the chromophore bearing polypeptides of the core were found to be chromophorylated. CD as well as absorption spectra showed the expected maxima around 652 and 675 nm from allophycocyanin (APC) and allophycocyanin B (APC-B) chromophores. Low temperature fluorescence and excitation spectra also showed that the core particles were fully functional with respect to the energy transfer between the APC chromophores. We conclude that PC and therefore the rods are dispensable for the survival of Synechococcus sp. PCC 7942. The results indicate that stable and functional core can assemble in absence of the rods. These rod-less phycobilisome core is able to transfer energy to Photosystem II.

On the use of a segmental shooting technique for multiple solutions of planar elastica problems

The solution of the non-linear equation of equilibrium for plane elastica problems is avoided by using a segmental shooting technique. In this procedure, the rod is divided into a series of segments, with each one undergoing only small deformations. The segments can be loaded in various manners and solved individually. Geometric and force compatibility is used to assemble the segments to allow analysis of the entire rod. The original boundary value problem is then solved by considering a sequence of initial value problems which converge to the required boundary conditions using a shooting technique. Since only one segment is considered at a time, the numerical calculations can be done accurately and efficiently on a PC type computer. When multiple solutions exist, the procedure outlined is useful in determining all of the solutions and in giving a partial understanding of the stability of each of the particular solutions. The procedure is applied to cantilevers and semi-circular arches to illustrate the technique.

Dp71, the nonmuscle product of the Duchenne muscular dystrophy gene is associated with the cell membrane

The 70.8 kDa protein, Dp71, is the major Duchenne muscular dystrophy (DMD) gene product in many nonmuscle tissues including the brain. Dp71 shares most of the C-terminal and cysteine-rich domains with the dystrophins but lacks the entire large rod shaped domain of spectrin-like repeats, and the N-terminal actin-binding domain. The function of Dp71 is unknown. Using subcellular fractionation and immunostaining we show that Dp71 is associated with the plasma membrane. Dp71 is also associated with the plasma membrane in mdx myogenic cells transfected with a vector expressing Dp71.

Tunicamycin does not inhibit transport of phosphatidylinositol toXenopus rod outer segments

Tunicamycin inhibits the dolichol pathway for N-linked glycosylation of proteins, including photoreceptor opsin, and causes a buildup of tubulo-vesicular profiles in the intersegmental space between photoreceptor rod inner and outer segments associated with disruption of new disc assembly. We tested the hypothesis that a tunicamycin lesion in photoreceptors would block lipid transport into the outer segment. Adult Xenopus retinas were preincubated in dim red light with 20 micrograms ml-1 of tunicamycin for one hour followed by incubation in the light for 2-6 h with tunicamycin plus either [3H]mannose, [3H]leucine, [2-(3)H]glycerol or [3H]myo-inositol. Tunicamycin caused accumulation of tubulo-vesicular membranes in the intersegmental space and significantly reduced both [3H]leucine and [3H]mannose incorporation into the basal region of rod outer segments. However, tunicamycin had no effect on [3H]glycerol incorporation into the rod outer segment phospholipids. After 5 h incubation with [3H]glycerol, radiolabel in outer segment fractions was associated primarily with phosphatidylinositol in both control and tunicamycin treated retinas. Quantitative light microscope autoradiography of both [3H]glycerol and [3H]inositol labelled retinas showed diffuse labelling over the entire rod outer segment in both control and tunicamycin treated retinas with no accumulation of radioactivity in the basal discs of control retinas or in the tubulo-vesicular structures in the intersegmental space of tunicamycin treated retinas. Our results indicate that despite the morphological disruption and inhibition of glycoprotein transport to outer segments after tunicamycin treatment, transport of labelled phosphatidylinositol occurs normally. These data add to a growing body of evidence separating the lipid and protein transport pathways to the outer segment.

Attentionally splitting the mass distribution of hand-held rods

Two experiments on the length-perception capabilities of effortful or dynamic touch differed only in terms of what the subject intended to perceive, while experimental conditions and apparatus were held constant. In each trial, a visually occluded rod was held as still as possible by the subject at an intermediate position. For two thirds of the trials, a weight was attached to the rod above or below the hand. In Experiment 1, in which the subject's task was to perceive the distance reachable with the portion of the rod forward of the hand, perceived extent was a function of the first moment of the mass distribution associated with the forward portion of the rod, and indifferent to the first moment of the entire rod. In Experiment 2, in which the task was to perceive the distance reachable with the entire rod if it was held at an end, the pattern of results was reversed. These results indicate the capability of selective sensitivity to different aspects of a hand-held object's mass distribution, without the possibility of differential exploration specific to these two tasks. Results are discussed in relation to possible roles of differential information, intention, and self-organization in the explanations of selective perceptual abilities.

Molecular architecture of the neurofilament: I. Subunit arrangement of neurofilament L protein in the intermediate-sized filament

Using the smallest subunit (NF-L) of a neurofilament and a glial fibrillary acidic protein, the subunit arrangement in intermediate filaments was studied by low-angle rotary shadowing. NF-L formed a pair of 70 to 80 nm rods in a low ionic strength solution at pH 6.8. Two 70 to 80 nm rods appeared to associate in an antiparallel manner with an overlap of about 55 nm, almost the same length as the α-helix-rich central rod domain of intermediate filament proteins. The overlap extended for three-beaded segments, present at 22 nm intervals along the pairs of rods. The observations that (1) 70 to 80 nm rods were a predominant structure in a low ionic strength solution at pH 8.5, (2) the molecular weights of the rod and the pair were measured by sedimentation equilibrium as 190,000 and 37,000 respectively, and (3) the rods formed from the trypsin-digested NF-L had a length of about 47 nm, indicated that the 70 to 80 nm rod is the four-chain complex and the pair of rods is the eight-chain complex. Similar structures were observed with glial fibrillary acidic protein, indicating that these oligomeric structures are common to other intermediate filament proteins. NF-L assembled into short intermediate-sized filaments upon dialysis against a low-salt solution containing 1 to 2 mm-MgCl2 at 4 °C. The majority of these short filaments possessed four or five-beaded segments, suggesting that the pair of rods were arranged in a half-staggered fashion in neurofilaments. On the basis of these observations, we propose the following model for the intermediate filament subunit arrangement. (1) The four-chain complex is the 70 to 80 nm rod, in which two coiled-coil molecules align in parallel and in register. (2) Two four-chain complexes form the eight-chain complex by associating in an antiparallel fashion with the overlap of the entire central rod domain. (3) The eight-chain complex is the building block of the intermediate filament. The eight-chain complexes are arranged in a half-staggered fashion within the intermediate filament.

Assembly properties of two CNBr fragments of avian desmin that correspond to the headpiece domain and Helix 1B

To study how different domains of the muscle-specific intermediate filament protein, desmin, contribute to its polymerization, two of its CNBr fragments were examined as to their oligomeric structure under assembly conditions. One of these, D88, covers residues 1–88 and represents almost the entire headpiece; the other, D109, covers residues 145–254, and includes the entire Helix 1B and part of linker L 12 of the intact molecule. Chemical cross-linking followed by SDS-PAGE, and analytical gel filtration, revealed that in 10 mM Tris-HCl, pH 8.5, conditions that favor tetramerization of intact desmin D88 formed only dimers. D109, on the other hand, formed primarily a dimeric species but low levels of trimeric and tetrameric species were also detectable. These data are consistent with the proposal that, during assembly of intact protein molecules into IF, the headpiece and Helix 1 contribute to dimerization of two polypeptides into a parallel, in-register coiled-coil. However, additional interactions, including headpiece-to-rod binding and hydrophobic interaction along the entire rod domain, are required to stabilize the tetramers and full-size IF.

Human embryonic myosin heavy chain cDNA: Interspecies sequence conservation of the myosin rod, chromosomal locus and isoform specific transcription of the gene

A 3.6 kilobase cDNA clone coding for the human embryonic myosin heavy chain has been isolated and characterized from an expression library prepared from human fetal skeletal muscle. The derived amino acid sequence for the entire rod part of myosin shows 97% sequence homology between human and rat and a striking interspecies sequence conservation among the charged amino acid residues. The single copy gene is localized to human chromosome 17 and its expression in fetal skeletal muscle is developmentally regulated. The sequence information permits the design of isoform-specific probes for studies on the structure of the gene and its role in normal and defective human myogenesis. Myosin heavy chain cDNA; Nucleotide sequence; Amino acid sequence; Myosin rod; Chromosomal mapping; Gene transcription; (Human embryo)

Perceiving extents of rods by wielding: Haptic diagonalization and decomposition of the inertia tensor.

We report two experiments on the length-perception capabilities of the hand-related haptic subsystem. On each trial, a visually occluded rod was wielded by the subject at a position intermediate between its two ends. The position was either 1/2 or 3/4 of the rod's length. On two-thirds of the trials, a weight was attached to the rod at a point either above or below its center of gravity and not coincident with the hand's position. In Experiment 1, the subject's task was to perceive the distance reachable with the portion of the rod extending beyond the position of the grasp. In the second experiment, the subject's task was to perceive the distance reachable with the entire rod if it were held at its proximal end. In Experiment 1, perceived reaching distance was a function of the moment of inertia of the amount of rod forward of the grasp about an axis through the proximal end of the rod segment. In Experiment 2, perceived reaching distance was a function of the moment of inertia of the entire rod about the given axis of rotation intermediate between the rod's ends. The results are discussed in terms of (a) the notion of smart perceptual instruments capitalizing on invariant properties of the inertia tensor and (b) how the haptic decomposition of moments of inertia follows the principle of equivalence of forces.

Synthesis of polyphosphoinositides in vertebrate photoreceptor membranes

Rod outer segments isolated from bovine retinas incorporated 32P into phospholipids after incubation with [γ- 32p]atp in a Mg 2+-containing medium. Only phosphatidylinositol 4-phosphate, phosphatidylinositol 4,5-bisphosphate and phosphatidate were labelled. The incorporation of label into lipids was detected as early as 20 s after the start of incubation and the products were stable for at least 10 min. The reactions were time, protein and ATP-concentration dependent. Entire rod outer segments showed higher diacylglycerol kinase and lower phosphatidylinositol and phosphatidylinositol 4-phosphate kinase activities than the disc membranes obtained from them. Exogenously added phosphatidylinositol (up to 1 inM) in the presence of Triton X-100 increased phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate labelling in rod outer segments (8- and 6-fold, respectively). Triton X-100 at a concentration of 0.4% stimulated phosphorylation of endogenous phosphoinositides. Diacylglycerol kinase activity was largely suppressed by the detergent, but this effect was partially reversed by addition of phosphatidylinositol. It is suggested that the rod outer segments contain phosphatidylinositol kinase and phosphatidylinositol 4-phosphate kinase bound to disc membranes, as well as an active diacylglycerol kinase occurring either as a soluble or a peripherally bound protein in disc membranes.

Phosphatidate phosphatase activity in isolated rod outer segment from bovine retina

Phosphatidate phosphohydrolase (EC 3.1.3.4) was detected in isolated bovine rod outer segments and its properties investigated. The enzyme activity was assayed using aqueously dispersed 1,2-diacyl-sn-[2- 3H]glycerol 3-phosphate as substrate. The phosphatidic acid concentration was optimal at 1 mM and the estimated K m value was 6.7 · 10 −4 M. The activity was linear for 60 min with a protein concentration of 0.3 mg. A clear pH optimum was observed at 7.5. When the enzyme activity was measured using as substrate phosphatidic acid containing 15% lyso compound, the production of diacylglycerols was inhibited by about 70–75% at all concentrations studied. In rod outer segment preparations containing 0.2 mM Mg 2+, further additions of the ion (0.2–2.5 mM) only produced a slight inhibition of the activity at 2.5 mM. F − (50 mM), Ca 2+ (1 mM) and EDTA (50 mM) inhibited the dephosphorylation of the substrate by 80, 10 and 70%, respectively. The aqueously dispersed phosphatidic acid-dependent activity present in rod outer segments was stimulated by Triton X-100 and taurocholate. Similar values were observed in the enzyme activity of entire rod outer segments and in that of disks obtained from them, showing the enzyme to be associated with disk membrane. The [ 3H]diacyIglycerol] production from [ 3H]phosphatidic acid was analyzed in synaptosomal-mitochondrial and microsomal fractions and in a crude pigment epithelium preparation. The degree of dephosphorylation of phosphatidic acid in these subcellular fractions was as follows: microsomes > synaptosomal-mitochondrial fraction > rod outer segment > crude pigment epithelium. The present report is the first evidence of phosphatidic acid phosphohydrolase activity associated with rod outer segment membranes.

Conformational stability of the myosin rod.

Chymotryptic cleavage patterns of myosin rods from pig stomach, chicken gizzard, and rabbit skeletal muscle indicate that short (approximately 45 nm) heavy meromyosin subfragment 2 (SF2) is a consistent product of all three rods, whereas long (approximately 60 nm) SF2 is derived only from skeletal muscle myosin. Differential scanning calorimetry was used to follow the thermally induced melting transition of the rods and certain of their subfragments. In 0.12 M KCl, sodium phosphate buffer, pH 6.2-7.6, the light meromyosin (LMM) and SF2 domains of each rod had essentially identical conformational stabilities. Temperature midpoints for the melting transitions were 54-56 degrees C for the two smooth muscle myosin rods and 50-53 degrees C for the skeletal muscle myosin rod. In 0.6 M K Cl buffer, melting transitions for the smooth muscle myosin rods were essentially unchanged, but skeletal muscle myosin rods showed multiphase melting, with major transitions at 43 degrees C and 52 degrees C. The first of these was tentatively attributed to LMM, and the second to SF2. In 0.12 M K Cl buffer, the LMM transition was stabilised so that it superimposed on that of SF2. No melting was observed in any of the rods at physiological temperature. These results indicate that, excluding a possible but only narrow hinge region, the entire myosin rod has essentially uniform conformational stability at physiological pH and ionic strength, and thus that the contractile and elastic properties of the cross-bridge exist in the heavy meromyosin subfragment 1 (SF1) domains of the molecule.

Is the SII portion of the cross-bridge in glycerinated rabbit psoas fibers compliant in the rigor state?

To see whether the SII portion of the cross-bridge in rigor fibers is longitudinally compliant, we chemically cross-linked with dimethyl suberimidate the entire rod portion (including the SII portion) of myosin onto the surface of thick filaments in glycerinated rabbit psoas fibers, and studied the effect of the SII fixation on the stiffness of the rigor fibers. The cross-linking of fiber segments with full filament overlap increased the rigor stiffness by approximately 25%. Almost the same absolute amount of the stiffness increase was also observed in rigor fibers with half- or no filament overlap after the cross-linking, and a similar but somewhat larger increment of stiffness was observed in fiber segments cross-linked in relaxing solution. These results indicate that the stiffness increase is not produced by the fixation of the SII portion onto the thick filament surface, but is caused instead by the cross-linking of some parallel elastic elements in muscle, and therefore indicate that the SII portion of the cross-bridge is hardly longitudinally compliant in rigor fibers.

A new histo- and cytochemical method for demonstration of cyclic 3',5'-nucleotide phosphodiesterase activity in retinal rod photoreceptor cells of the rat.

Cyclic 3',5'-mononucleotide phosphodiesterase (cyclic nucleotide PDEase) activity was studied histo- and cytochemically in the retinal rod photoreceptor cells of the rat by means of a newly developed technique utilizing the intrinsic 5' nucleotidase activity instead of an exogenous 5' nucleotidase source (snake venom). Cyclic GMP and was used as a substrate, the intense activity of phosphodiesterase (PDEase) was distributed over the entire rod outer segments; reaction product was observed on the plasmalemma and on the disk membranes of the outer segments. A slight reaction was also observed on the plasmalemma of the inner segments. However, no precipitate was found in the perinuclear and synaptic regions of the rod photoreceptors. In contrast, when cyclic AMP was utilized as a substrate, a moderate reaction was seen in the synaptic region of the plexiform layer. The intensity of the reaction in the outer segments was much reduced in comparison to the results with cyclic GMP. The enzyme activity was almost completely inhibited by 2 mM 3-isobutyl-1-methylxanthine (IBMX) or 2 mM theophylline, which were potent inhibitors of PDEase. To confirm the propriety of our new cytochemical method, the localization of 5' nucleotidase was also studied utilizing 5' AMP or 5' GMP as substrates. In contrast to the activity of cyclic nucleotide PDEase, the activity of 5' nucleotidase was distributed on all membranes of the photoreceptors from the synaptic outer plexiform layer to the tip of outer segments.(ABSTRACT TRUNCATED AT 250 WORDS)

Matching molecules in the catch mechanism.

A new structural model is advanced to account for the specialized "catch" contraction of molluscan smooth muscles. The myosin of the thick filaments of these muscles is pictured as comprising a single layer of molecules whose assembly and activity are controlled by the underlying core of paramyosin. This organization differs from that in other muscles in which the myosin is grouped into bundles. The catch state--in which the lifetime of the actin-myosin linkage is extended--would depend directly on this special organization that allows an interaction of the entire rod region of myosin with the paramyosin core. This interaction might be modulated by phosphorylation of paramyosin.

Three-Dimensional Thermal-Hydraulic Analysis of Wire-Wrapped Rods in Liquid-Metal Fast Breeder Reactor Core Assemblies

A study is presented to determine the detailed coolant velocity and temperature profile around the entire rod circumference in liquid-metal fast breeder reactor (LMFBR) core assemblies as well as the detailed radial and circumferential temperature profile in the rod. The digital computer code FATHOM-360 developed to perform the above calculations is described. Fuel, radial blanket, and control assembly rods (both wire-trapped and bare) can be analyzed. Coolant, cladding, and fuel (or absorber) temperature profiles are calculated for uniform and nonuniform heat generation (i.e., accounting for power skew across the pellet) in the rod. Temperature distributions can be calculated for both concentric and eccentric positions of the pellet with respect to the fuel rod cladding. Typical examples of the calculational capabilities of the code are presented. Such capabilities are needed for a reliable design of LMFBR core assemblies and rods to provide detailed cladding temperature profiles and accurately calculate the cladding strain on which the fuel rod lifetime and allowable burnup depend. Overall, a more realistic core thermofluids design is possible by implementing the study presented here.

Rhodopsin in the rod outer segment plasma membrane.

Isolated frog retinas were incubated in vitro with a 4-h pulse of [3H]leucine, then chased for 32 h with a nonradioactive amino acid mixture. At the end of the incubation, light and electron microscope autoradiograms were prepared from some of the retinas. The autoradiograms revealed: (a) intense radioactivity in the basal disks of the rod outer segments, (b) diffuse label evenly distributed throughout the rod outer segments, and (c) a high concentration of label in the entire rod outer segment plasma membrane. Incubation under identical conditions, but with puromycin added, significantly inhibited the labeling of all of these components. To identify the labeled proteins, purified outer segments from the remaining retinas were analyzed biochemically by SDS disc gel electrophoresis and gel filtration chromatography. SDS gel electrophoresis showed that about 90% of the total rod outer segment radioactivity chromatographed coincident with visual pigment, suggesting that the radiolabeled protein in the plasma membrane is visual pigment. Gel filtration chromatography demonstrated that the radiolabeled protein co-chromatographed with rhodopsin rather than opsin, and that the newly synthesized visual pigment is both the basal disks and the plasma membrane is present in the native configuration.

Modes of Resonance Absorptions of Magnetostatic Waves in Axially Magnetized YIG Rods

The magnetostatic resonance absorptions for an axially magnetized single crystal of YIG rod, [110] oriented, are observed in a waveguide and a cavity at 9.45 GHz. For the applied magnetic field ranging from 2800 to 3400 Oe resonance absorptions due to the dominant and third modes are observed but the second mode is not excited. The separation of these modes is conducted by comparing the transmitted power with the reflected microwave power pattern. For the dominant and third modes the relations between the field deviation, ΔH, from the cutoff field and the standing wave number n for each absorption are in good agreement between the theoretical and experimental values in a wide range of dc magnetic field. When the entire rod is inserted into the center of a waveguide or a cavity only the absorptions due to modes associated with an odd number, n, are observed for both dominant and third modes

One dimensional elastic wave propagation in a non-homogeneous conical rod

A one dimensional problem concerning the wave propagation in a non-homogeneous conical rod having its cross-section directly proportional to the square of the distance from a chosen point is considered in this paper. One end of the rod is subjected to a pressure step while the other end is stress free and the pulse shape is examined when the propagation speed of the wave has a particular continuous distribution over the entire rod.

Effects of a hallucinogenic agent in totally blind subjects.

A previous study demonstrated that lysergic acid diethylamide (LSD) induced measurable changes in human retinal function while visual hallucinations and illusions were being experienced.1The changes were evident in both dark-adaptation and electroretinographic studies and were interpreted as mildly hypoxic or toxic retinal effects of LSD. In the dark-adaptation curve, LSD delayed the rod-cone break and elevated the entire rod threshold. In the electroretinogram (ERG), the drug increased the scotopicbwave amplitudes and in some subjects also increased the scotopica-wave amplitudes. Because hallucinations and illusions were reported only when measurable ERG and dark-adaptation changes occurred, a possible retinal role in the induction of hallucinations was considered. Consequently, the present study was undertaken to clarify the role of a functioning retina in the induction of LSD-induced visual changes by studying subjects with total blindness (no light perception) and, insofar as could be determined, normal central nervous system