Detecting spatial patterns in biological array experiments

Chemical genetic screening and DNA and protein microarrays are among a number of increasingly important and widely used biological research tools that involve large numbers of parallel experiments arranged in a spatial array. It is often difficult to ensure that uniform experimental conditions are present throughout the entire array, and as a result, one often observes systematic spatially correlated errors, especially when array experiments are performed using robots. Here, the authors apply techniques based on the discrete Fourier transform to identify and quantify spatially correlated errors superimposed on a spatially random background. They demonstrate that these techniques are effective in identifying common spatially systematic errors in high-throughput 384-well microplate assay data. In addition, the authors employ a statistical test to allow for automatic detection of such errors. Software tools for using this approach are provided.     

High flexibility of the actomyosin crossbridge resides in skeletal muscle myosin subfragment-2 as demonstrated by a new single molecule assay

Popular views of force generation in muscle indicate that a lever arm in the myosin head initiates displacement of the thin filament. However, this lever arm is attached to the thick filament backbone by a flexible combination of coiled coils and hinges in the myosin subfragment-2 (S2); therefore, efficient force generation depends on tension development in this linking structure. Herein, a single molecule assay is developed to examine the flexibility of the intact S2 relative to that of the myosin head. Fluorescently labeled myosin rod is polymerized onto a single myosin molecule that is bound to actin, and the resulting Brownian motion of the rod is analyzed at video rates by digital image processing. Complete rotations of the rod suggest significant amounts of random coil in the linking structure. The close similarity of twist rates for double-headed and single-headed myosin indicates that most of the flexibility originates at or beyond the first pitch of coiled coil in S2 and most likely at the hinge connecting S2 and the light meromyosin. The myosin head has a smaller but still detectable impact on this flexibility, since the addition of ADP to the rigor crossbridge produces differential effects on the torsional characteristics of double-headed versus single-headed myosin.     

Crystal structure of methyl 1,2,3,4-tetra-O-acetyl-beta-D-glucopyranuronate

The identity of the crystalline product formed by the acetylation of a mixture of methyl alpha- and beta-D-glucopyranuronates has been confirmed as being methyl 1,2,3,4-tetra-O-acetyl-beta-D-glucopyranuronate (3), which agrees with the assignment from 1H NMR. The absolute configuration of compound 3 was assigned to agree with the known chirality of the precursor sugar, D-glucono-6,3-lactone.     

Epizootiology of Sin Nombre and El Moro Canyon hantaviruses, southeastern Colorado, 1995-2000

Sin Nombre virus (SNV) is an etiologic agent of hantavirus pulmonary syndrome. To better understand the natural history of this virus we studied population dynamics and temporal pattern of infection of its rodent hosts in southeastern Colorado (USA) from 1995 to 2000. We present evidence for the presence of two hantaviruses, SNV in deer mice (Peromyscus maniculatus) and El Moro Canyon virus in western harvest mice (Reithrodontomys megalotis), at our study sites. Sin Nombre virus appeared only sporadically in deer mouse populations; overall prevalence of antibody to SNV was 2.6%. El Moro Canyon virus was enzootic: seroconversions occurred throughout the year; antibody prevalence (11.9% overall) showed a delayed-density-dependent pattern, peaking as relative abundance of mice was declining. Males of both host species were more frequently infected than were females. An apparently lower mean survivorship (persistence at the trapping site) for SNV antibody-positive deer mice could indicate a detrimental effect of SNV on its host, but might also be explained by the fact that antibody-positive mice were older when first captured.     

Pregnancy diagnosis and abnormalities of pregnancy in the dog

New and traditional techniques for pregnancy diagnosis in the dog are reviewed. Imaging techniques yield the most information, with ultrasonography the preferred technique for assessment of fetal viability and radiography the preferred technique for assessment of litter size. Diabetes mellitus and pregnancy toxemia are uncommon complications of pregnancy. Pathogenesis, diagnosis and treatment of infectious and non-infectious causes of pregnancy loss are reviewed.     

A computational comparison of the atomic models of the actomyosin interface

Several atomic models of the actomyosin interface have been proposed based on the docking together of their component structures using electron microscopy and resonance energy-transfer measurements. Although these models are in approximate agreement in the location of the binding interfaces when myosin is tightly bound to actin, their relationships to molecular docking simulations based on computational free-energy calculations are investigated here. Both rigid-docking and flexible-docking conformational search strategies were used to identify free-energy minima at the interfaces between atomic models of myosin and actin. These results suggest that the docking model produced by resonance energy-transfer data is closer to a free-energy minimum at the interface than are the available atomic models based on electron microscopy. The conformational searches were performed using both scallop and chicken skeletal muscle myosins and identified similarly oriented actin-binding interfaces that serve to validate that these models are at the global minimum. These results indicate that the existing docking models are close to but not precisely at the lowest-energy initial contact site for strong binding between myosin and actin that should represent an initial contact between the two proteins; therefore, conformational changes are likely to be important during the transition to a strongly bound complex.     

The dance of actin and myosin: a structural and spectroscopic perspective

Actin and myosin interact in a cyclic series of steps linked to the hydrolysis of ATP that are representative of an ancient and widespread molecular mechanism. Spectroscopic findings are related to the analysis of the actin and myosin structures and results from kinetics, fibers, single molecules, electron microscopy, genetics, and a variety of other biophysical and biochemical studies on actin and myosin to provide an overview of the steps in this molecular process. The synthesis of the key findings from these fields reveals a highly efficient engine that amplifies subtle changes in the active site into unsurpassed molecular displacements. Recent developments in resonance energy-transfer spectroscopy and X-ray crystallography are enabling a detailed elucidation of the stages of a large power stroke that concurs with evidences from diverse lines of structural and kinetic inquiry. A complete image of actin and myosin motility appears to include twists, tilts, steps, and dynamics from both partners that could be described as a molecular dance.     

Force transmission via axial tendons in undulating fish: a dynamic analysis

Sonomicrometrics of in vivo axial strain of muscle has shown that the swimming fish body bends like a homogenous, continuous beam in all species except tuna. This simple beam-like behavior is surprising because the underlying tendon structure, muscle structure and behavior are complex. Given this incongruence, our goal was to understand the mechanical role of various myoseptal tendons. We modeled a pumpkinseed sunfish, Lepomis gibbosus, using experimentally-derived physical and mechanical attributes, swimming from rest with steady muscle activity. Axially oriented muscle-tendons, transverse and axial myoseptal tendons, as suggested by current morphological knowledge, interacted to replicate the force and moment distribution. Dynamic stiffness and damping associated with muscle activation, realistic muscle force generation, and force distribution following tendon geometry were incorporated. The vertebral column consisted of 11 rigid vertebrae connected by joints that restricted bending to the lateral plane and endowed the body with its passive viscoelasticity. In reaction to the acceleration of the body in an inviscid fluid and its internal transmission of moment via the vertebral column, the model predicted the kinematic response. Varying only tendon geometry and stiffness, four different simulations were run. Simulations with only intrasegmental tendons produced unstable axial and lateral tail forces and body motions. Only the simulation that included both intra- and intersegmental tendons, muscle-enhanced segment stiffness, and a stiffened caudal joint produced stable and large lateral and axial forces at the tail. Thus this model predicts that axial tendons function within a myomere to (1) convert axial force to moment (moment transduction), (2) transmit axial forces between adjacent myosepta (segment coupling), and, intersegmentally, to (3) distribute axial forces (force entrainment), and (4) stiffen joints in bending (flexural stiffening). The fact that all four functions are needed to produce the most realistic swimming motions suggests that axial tendons are essential to the simple beam-like behavior of fish.     

Combining richness and abundance into a single diversity index using matrix analogues of Shannon's and Simpson's indices

Shannon's and Simpson's indices have been the most widely accepted measures of ecological diversity for the past fifty years, even though neither statistic accounts for species abundances across geographic locales ("patches"). An abundant species that is endemic to a single patch can be as much of a conservation concern as a rare cosmopolitan species. I extend Shannon's and Simpson's indices to simultaneously account for species richness and relative abundances – i.e. extend them to multispecies metacommunities – by making the inputs to each index a matrix, rather than a vector. The Shannon's index analogue of diversity is mutual entropy of species and patches divided by marginal entropy of the individual geographic patches. The Simpson's index analogue of diversity is a modification of mutual entropy, with the logarithm moved to the outside of the summation, divided by Simpson's index of the patches. Both indices are normalized for number of patches, with the result being inversely proportional to biodiversity. These methods can be extended to account for time-series of such matrices and average age-classes of each species within each patch, as well as provide a measure of spatial coherence of communities.     

Characterization ofBradyrhizobium sp. (Astragalus sinicus L.) using serological agglutination,intrinsic antibiotic resistance,plasmid visualization,and field performance

Eleven isolates ofBradyrhizobium sp. (Astragalus sinicus L.), originating from plants ofAstragalus sinicus (Chinese milkvetch) in China and Japan, were characterized using serological agglutination, intrinsic antibiotic resistance, and plasmid visualization. Serological agglutination, using antisera against three isolates effective onA. sinicus and two isolates effective on otherAstragalus spp., showed that there may be two serogroups among the eleven isolates effective onA. sinicus, and that the antigenic components are heterogeneous. Results of both intrinsic antibiotic resistance and plasmid visualization demonstrated similarities between four of the isolates.Ten isolates were further evaluated for effectiveness on the host,A. sinicus, in a field study in Puyallup, Washington (47°N, 122°W). This study showed that the Chinese commercial inoculum and one field isolate were about ten times more effective than the control based on plant dry weight and seed yield. Eight other isolates were less effective.