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Division site selection in Escherichia coli involves dynamic redistribution of Min proteins within coiled structures that extend between the two cell poles.

The MinCDE proteins of Escherichia coli are required for proper placement of the division septum at midcell. The site selection process requires the rapid oscillatory redistribution of the proteins from pole to pole. We report that the three Min proteins are organized into extended membrane-associated coiled structures that wind around the cell between the two poles. The pole-to-pole oscillation of the proteins reflects oscillatory changes in their distribution within the coiled structure. We also report that the E. coli MreB protein, which is required for maintaining the rod shape of the cell, also forms extended coiled structures, which are similar to the MreB structures that have previously been reported in Bacillus subtilis. The MreB and MinCDE coiled arrays do not appear identical. The results suggest that at least two functionally distinct cytoskeletal-like elements are present in E. coli and that structures of this type can undergo dynamic changes that play important roles in division site placement and possibly other aspects of the life of the cell.

Adenosine Triphosphatases↗

Assessment of wood utility pole climbability using psychophysical and mechanical measurements.

The issue of climbability has been raised on several occasions for more than a decade in North America. Presently, climbability is estimated from the pole hardness measured by the Pilodyn measurements (6 J). However, the use of Pilodyn measurements to discriminate the pole hardness value is criticized by climbers, who claim that the Pilodyn hardness measurement is affected by species-treatment combinations and that it does not reflect gaff penetration or climbability. Furthermore, climbability evaluations have been conducted in which test poles were climbed by linemen, and corresponding subjective ratings were recorded. However, the ability of psychophysical measurements to accurately discriminate close hardness pole values and to differentiate species-treatment combinations at specific hardness levels have not yet been fully documented. The aim of this study is to evaluate the psychophysical perception of linemen and the mechanical measurements of gaff penetration and gaff impact during the climbing of different wood species and treatment combinations in order to compare these results with Pilodyn measurements within a precise range of pole hardnesses, to study the relationships between these variables, and, finally, to propose various design guidelines for the development of a better tool for the evaluation of climbability.

Adult↗

The Cdc31p-binding protein Kar1p is a component of the half bridge of the yeast spindle pole body.

KAR1 has been identified as an essential gene which is involved in karyogamy of mating yeast cells and in spindle pole body duplication of mitotic cells (Rose, M. D., and G. R. Fink. 1987. Cell. 48:1047-1060). We investigated the cell cycle-dependent localization of the Kar1 protein (Kar1p) and its interaction with other SPB components. Kar1p is associated with the spindle pole body during the entire cell cycle of yeast. Immunoelectron microscopic studies with anti-Kar1p antibodies or with the monoclonal antibody 12CA5 using an epitope-tagged, functional Kar1p revealed that Kar1p is associated with the half bridge or the bridge of the spindle pole body. Cdc31p, a Ca(2+)-binding protein, was previously identified as the first component of the half bridge of the spindle pole body (Spang, A., I. Courtney, U. Fackler, M. Matzner, and E. Schiebel. 1993. J. Cell Biol. 123:405-416). Using an in vitro assay we demonstrate that Cdc31p specifically interacts with a short sequence within the carboxyl terminal half of Kar1p. The potential Cdc31p-binding sequence of Kar1p contains three acidic amino acids which are not found in calmodulin-binding peptides, explaining the different substrate specificities of Cdc31p and calmodulin. Cdc31p was also able to bind to the carboxy terminus of Nuflp/Spc110p, another component of the SPB (Kilmartin, J. V., S. L. Dyos, D. Kershaw, and J. T. Finch. 1993. J. Cell Biol. 123:1175-1184). The association of Kar1p with the spindle pole body was independent of Cdc31p. Cdc31p, on the other hand, was not associated with SPBs of kar1 cells.

Base Sequence↗

The absence of centrioles from spindle poles of rat kangaroo (PtK2) cells undergoing meiotic-like reduction division in vitro.

Light and electron microscopy were used to study somatic cell reduction division occurring spontaneously in tetraploid populations of rat kangaroo Potorous tridactylis (PtK2) cells in vitro. Light microscopy coupled with time-lapse photography documented the pattern of reduction division which includes an anaphase-like movement of double chromatid chromosomes to opposite spindle poles followed by the organization of two separate metaphase plates and synchronous anaphase division to form four poles and four daughter nuclei. The resulting daughter cells were isolated and cloned, showing their viability, and karyotyped to determine their ploidy. Ultrastructural analysis of cells undergoing reduction consistently revealed two duplexes of centrioles (one at each of two spindle poles) and two spindle poles in each cell that lacked centrioles but with microtubules terminating in a pericentriolar-like cloud of material. These results suggest that the centriole is not essential for spindle pole formation and division and implicate the could region as a necessary component of the spindle apparatus.

Cell Line↗

Nucleation of microtubules in vitro by isolated spindle pole bodies of the yeast Saccharomyces cerevisiae.

Spindle pole bodies (SPBs) were isolated from the yeast Saccharomyces cerevisiae by an adaptation of the Kleinschmidt monolayer technique. Spheroplasts prepared from the cells were lysed on an air-water interface. Spread preparations were picked up on grids, transferred to experimental test solutions, and prepared for whole-mount electron microscopy. Using purified exogenous tubulin from porcine brain tissue, the isolated SPBs were shown to nucleate the assembly of microtubules in vitro. Microtubule growth was directional and primarily onto the intranuclear face of the SPB. Neither the morphology nor the microtubule-initiating capacity of the SPB was affected by treatment with the enzymes DNase, RNase, or phospholipase although both properties were sensitive to trypsin. Analysis of SPBs at various stages of the cell cycle showed that newly replicated SPBs had the capacity to nucleate microtubules. SPBs isolated from exponentially growing cells initiated a subset of the yeast spindle microtubules equivalent to the number of pole-to-pole microtubules seen in vivo. However, SPBs isolated from cells in stationary phase and therefore arrested in G1 nucleated a number of microtubules equal to the total chromosomal and pole-to-pole tubules in the yeast spindle. This may mean that in G1-arrested cells, the SPB is associated with microtubule attachment sites of the yeast chromatin.

Cell Cycle↗

Direct percutaneous approach to the upper pole of the kidney: MRI anatomy with assessment of the visceral risk.

PURPOSE: In an attempt to determine the visceral risk secondary to a direct percutaneous puncture of the upper renal calix, the anatomic relations of the upper pole of the kidney were studied by magnetic resonance imaging. METHODS: Examination was performed on 25 normal volunteers placed successively in the right and left prone oblique position. The kidney axis and minimal distances from the cutaneous plane at the level of the upper and lower poles were measured. Axial and tangential simulated percutaneous approaches to the upper renal calix were compared in term of risk of damage to the pulmonary, splenic, and hepatic parenchyma. RESULTS: The transversal anteversion angle was statistically comparable for right and left kidneys, but the sagittal anteversion angle was significantly higher for right kidneys (p = 0.05). The minimal distance from the cutaneous plane was statistically comparable for the upper and lower poles. The lower pole was significantly deeper for left than right kidneys (p = 0.01). The visceral risk was statistically comparable for left and right kidneys and was significantly higher in case of an approach in the axis of the upper renal calix or through the 10th intercostal space compared to a puncture via the l1th space (p = 0.0001). CONCLUSION: A percutaneous puncture of the upper pole of the kidney above the 11th rib increases the risk of visceral damage. Preoperative evaluation, with the aid of CT scan or MRI, of the risk of pulmonary, splenic, or hepatic injury could be carried out in these cases.

Adult↗

Upper-pole access for percutaneous nephrolithotomy: comparison of supracostal and infracostal approaches.

BACKGROUND AND PURPOSE: The advantage of upper-pole access for nephrolithotomy is direct access to most of the intrarenal collecting system and upper ureter. Upper-pole access can be achieved either supracostally and subcostally. Because of the anatomic location of the kidneys, the supracostal approach is associated with a higher rate of pulmonary complications. We compared the efficacy and safety of the supracostal and infracostal upper-pole approaches. PATIENTS AND METHODS: A total 464 patients were treated with percutaneous nephrolithotomy (PCNL) via the upper pole, of which 170 punctures (group I) were performed supracostally and 294 (group II) subcostally. In both groups, PCNL was done by the standard technique with fluoroscopic guidance. The operative time, success rate, hospital stay, and complications in the two groups were compared. RESULTS: Patients were stone free in 82.2% and 77.1% of the cases in groups I and II, respectively, and had stone fragments <4 mm in 10.7% and 14.7%, respectively. The operative time, success rate, and septic and hemorrhagic complications were not significantly different in the two groups. Hydrothorax was found in 26 patients (15.3%) of group I and 4 (1.4%) of group II. Only 9 patients (5.3%) in group I needed intercostal drainage. CONCLUSION: Percutaneous nephrolithotomy via the upper pole is effective using both supracostal and infracostal approaches, with acceptable rates of complications. The rate of pulmonary complications is higher with the supracostal approach. If the supracostal approach is indicated, it should be used with caution.

Adult↗

Lower pole calicostomy for the management of iatrogenic ureteropelvic junction obstruction.

It is often possible to use endoscopic techniques for the management of iatrogenic upper ureteral or ureteropelvic junction obstruction. However, in some cases with severe stricture disease or significant ischemic injury open surgical reconstruction is necessary. We report our experience with ileal ureter-lower pole calicostomy for the management of these complex urological injuries. During the last 3 years we treated 3 patients with severe ureteral/ureteropelvic junction obstruction secondary to iatrogenic injuries, including ureteral avulsion during ureteroscopic stone extraction, ureteral laceration during dilation for diagnostic ureteropyeloscopy, and ureteral ligation with ureteropelvic junction disruption and large peri-pelvic urinoma. In all cases unsuccessful attempts at endoscopic management necessitated open repair. Lower pole heminephrectomy was performed in all patients to expose the lower pole calix and ileal ureter-lower pole calicostomy was created due to the injury of large segments of the ureter. Satisfactory results were demonstrated on postoperative excretory urography and by a lack of symptoms. Followup averaged 23 months (range 20 to 26) with stable renal function in all patients. We believe that ileal ureter-lower pole calicostomy represents an attractive alternative for the management of severe ischemic, iatrogenic upper ureteral or ureteropelvic junction obstruction when endoscopic maneuvers are not possible or ineffective.

Adult↗

Muscular and metabolic costs of uphill backpacking: are hiking poles beneficial?

PURPOSE: The purpose of the present study was to compare pole and no-pole conditions during uphill backpacking, which was simulated on an inclined treadmill with a moderately heavy (22.4 kg, 30% body mass) backpack. METHODS: Physiological measurements of oxygen consumption, heart rate, and RPE were taken during 1 h of backpacking in each condition, along with joint kinematic and electromyographic comparisons from data collected during a third test session. RESULTS: The results showed that although imposing no metabolic consequence, pole use elicited a longer stride length (1.27 vs 1.19 m), kinematics that were more similar to those of unloaded walking, and reduced activity in several lower extremity muscles. Although pole use evoked a greater heart rate (113.5 vs 107 bpm), subjects were backpacking more comfortably as indicated by their ratings of perceived exertion (10.8 vs 11.6). The increased cardiovascular demand was likely to support the greater muscular activity in the upper extremity, as was observed in triceps brachii. CONCLUSION: By redistributing some of the backpack effort, pole use alleviated some stress from the lower extremities and allowed a partial reversal of typical load-bearing strategies.

Adult↗

Increased radiodensity of the proximal pole of the scaphoid: a common finding in computed tomography imaging of the wrist.

OBJECTIVES: To determine whether the proximal pole (PP) of the scaphoid is denser than the distal pole (DP) in a diverse population of patients undergoing computed tomography (CT) imaging of the wrist. METHODS: Sixty-nine patients who had CT of the wrist were retrospective reviewed. We measured CT density of the medullary bone of PP and DP of the scaphoid and calculated PP/DP ratio. To evaluate the variability of PP/DP ratio, we compared the ratios of these 3 groups. These patients were separated into 3 groups based on their diagnoses: group 1, healed scaphoids treated by casting; group 2, scaphoid nonunions; group 3, intact scaphoids. RESULTS: Proximal pole was denser than DP in most subjects. Proximal pole/distal pole ratio was similar among groups (P < 0.05). Fracture healing, sex, and age did not affect this ratio. However, we did not study scaphoids with avascular necrosis. CONCLUSIONS: Proximal pole was denser than DP in most of our subjects, including those with intact scaphoids, healing scaphoid fractures, and nonunions.

Adult↗

Normal pole formation during total inhibition of wall synthesis of Bacillus subtilis.

Previous work has shown that the side wall of a Gram-positive rod is initially laid down as a compact layer inside the older wall. It is then stretched as it comes to bear tension due to the osmotic pressure inside the cell. If the polar wall is likewise capable of a degree of expansion, then no new murein need be added while the planar cross-wall splits and converts into two poles. In Bacillus subtilis mutant strain FJ6, which is deficient in autolytic enzymes, pole formation can be caused by addition of exogenous muramidase (10 micrograms hen egg white lysozyme ml-1 for 10 min at 35 degrees C). This strain grows as long filaments with many completed cross-walls, but enzymic treatment caused the formation of many new poles of normal morphology as judged by thin section electron microscopy. Fully separated poles of normal appearance were also found when more than 100 times the MIC (1 microgram ml-1) of vancomycin was added to block wall growth totally and rapidly 10 min before the addition of lysozyme. We conclude, therefore, that no new murein is needed in the conversion of the flat septum into poles and that the unstressed cross-wall is capable of the necessary expansion.

Bacillus subtilis↗

Characterization and spontaneous mutation of a novel gene, polE, involved in pellicle formation in Acetobacter tropicalis SKU1100.

Acetobacter tropicalis SKU1100 produces a pellicle polysaccharide, consisting of galactose, glucose and rhamnose, which attaches to the cell surface. This strain forms two types of colony on agar plates: a rough-surfaced colony (R strain) and a mucoid smooth-surfaced colony (S strain). The R strain forms a pellicle, allowing it to float on the medium surface in static culture, while the S strain does not. The pellicle is an assemblage of cells which are tightly associated with capsular polysaccharides (CPS) on the cell surface. In this study, a gene required for pellicle formation by the R strain was investigated by transposon mutagenesis using Tn10. The resulting mutant, designated Pel-, has a smooth-surfaced colony and a defect in pellicle formation, as for the S strain. The mutant produced polysaccharide which was instead secreted into the culture medium as extracellular polysaccharide (EPS). An ORF was identified at the Tn10 insertion site, designated polE, upstream of which polABCD genes were also found. The deduced amino acid sequences of polABCD showed a high level of homology to those of rfbBACD which are involved in dTDP-rhamnose synthesis, whereas polE had a relatively low level of homology to glycosyltransferase. In this study a polB (rfbA) disruptant was also prepared, which lacked both CPS and EPS production. A plasmid harbouring the polE or polB genes could restore pellicle formation in the Pel(-) mutant and S strains, and in the DeltapolB mutant, respectively. Thus both polE and polB are evidently involved in pellicle formation, most likely by anchoring polysaccharide to the cell surface and through the production of dTDP-rhamnose, respectively. The Pel- and DeltapolB mutants were unable to grow in static culture and became more sensitive to acetic acid due to the loss of pellicle formation. Additionally, this study identified the mutation sites of several S strains which were spontaneously isolated from the original culture and found them to be concentrated in a sequence of 7 C residues in the coding sequence of polE, with the deletion or addition of a single C nucleotide.

Acetobacter↗

The type IV secretion apparatus protein VirB6 of Agrobacterium tumefaciens localizes to a cell pole.

Agrobacterium tumefaciens VirB proteins assemble a type IV secretion apparatus for the transfer of DNA and proteins to plant cells. To study the role of the VirB6 protein in the assembly and function of the type IV apparatus, we determined its subcellular location by immunofluorescence microscopy. In wild-type bacteria VirB6 localized to the cell poles but in the absence of the tumour-inducing plasmid it localized to random sites on the cell membranes. Five of the 11 VirB proteins, VirB7-VirB11, are required for the polar localization of VirB6. We identified two regions of VirB6, a conserved tryptophan residue at position 197 and the extreme C-terminus, that are essential for its polar localization. Topology determination by PhoA fusion analysis placed both regions in the cell cytoplasm. Alteration of tryptophan 197 or the deletion of the extreme C-terminus led to the mislocalization of the mutant protein. The mutations abolished the DNA transfer function of the protein as well. The C-terminus of VirB6, in silico, can form an amphipathic helix that may encode a protein-protein interaction domain essential for targeting the protein to a cell pole. We previously reported that another DNA transfer protein, VirD4, localizes to a cell pole. To determine whether VirB6 and VirD4 localize to the same pole, we performed colocalization experiments. Both proteins localized to the same pole indicating that VirB6 and VirD4 are in close proximity and VirB6 is probably a component of the transport apparatus.

Agrobacterium tumefaciens↗

Theoretical examination of ultrasonic pole figures via comparison with the results analyzed by finite element polycrystal model.

The ultrasonic wave velocities in a polycrystalline aggregate are sensitively influenced by texture development due to plastic deformation. According to Sayer's model, it is possible to construct ultrasonic pole figures via the crystallite orientation distribution function (CODF), which can be calculated by using ultrasonic wave velocities. In the previous papers, the theoretical modeling to simulate ultrasonic wave velocities propagating in solid materials under plastic deformation has been proposed by the authors and proved to be a good agreement with experimental results. Generally, wave velocities are dependent upon the propagating wave frequency; hence to evaluate texture development via ultrasonic pole figures it is necessary to examine an influence of frequency dependence on the ultrasonic wave velocities. In the present paper, the proposed theoretical modeling is applied to the texture characterization in polycrystalline aggregates of FCC metals under various plastic strain histories via ultrasonic pole figures, and also the frequency dependence is examined by using Granato-Lücke's dislocation strings model. Then the simulated ultrasonic pole figures are compared with the pole figures analyzed by the finite element polycrystal model (FEPM). The good qualitative agreement between both results suggests the sufficient accuracy of our proposed theoretical modeling.

Journal Article↗

Conservative duplication of spindle poles during meiosis in Saccharomyces cerevisiae.

During sporulation in diploid Saccharomyces cerevisiae, spindle pole bodies acquire the so-called meiotic plaque, a prerequisite for spore formation. Mpc70p is a component of the meiotic plaque and is thus essential for spore formation. We show here that MPC70/mpc70 heterozygous strains most often produce two spores instead of four and that these spores are always nonsisters. In wild-type strains, Mpc70p localizes to all four spindle pole bodies, whereas in MPC70/mpc70 strains Mpc70p localizes to only two of the four spindle pole bodies, and these are always nonsisters. Our data can be explained by conservative spindle pole body distribution in which the two newly synthesized meiosis II spindle pole bodies of MPC70/mpc70 strains lack Mpc70p.

Fungal Proteins↗

A yeast gene essential for regulation of spindle pole duplication.

In eucaryotic cells, duplication of spindle poles must be coordinated with other cell cycle functions. We report here the identification in Saccharomyces cerevisiae of a temperature-sensitive lethal mutation, esp1, that deregulates spindle pole duplication. Mutant cells transferred to the nonpermissive temperature became unable to continue DNA synthesis and cell division but displayed repeated duplication of their spindle pole bodies. Although entry into this state after transient challenge by the nonpermissive temperature was largely lethal, rare survivors were recovered and found to have become increased in ploidy. If the mutant cells were held in G0 or G1 during exposure to the elevated temperature, they remained viable and maintained normal numbers of spindle poles. These results suggest dual regulation of spindle pole duplication, including a mechanism that promotes duplication as cells enter the division cycle and a negative regulatory mechanism, controlled by ESP1, that limits duplication to a single occurrence in each cell division cycle. Tetrad analysis has revealed that ESP1 resides at a previously undescribed locus on the right arm of chromosome VII.

Cell Cycle↗

Deficiency in POLE Exonuclease Causes Synthetic Lethality in Highly Aneuploid Cancer Cells.

UNLABELLED: Aneuploidy is a hallmark of cancer and is associated with drug resistance and poor clinical outcomes across diverse cancer types. However, no therapies have been clinically established to target highly aneuploid tumors. By analyzing nearly half a million tumor samples subjected to comprehensive genomic profiling, we identified a striking mutual exclusivity between POLE exonuclease domain mutations and high aneuploidy burden. This observation was independently validated using data from The Cancer Genome Atlas (TCGA) and the Cancer Cell Line Encyclopedia (CCLE). Probabilistic modeling revealed that the elevated quantity and unique spectrum of mutations induced by POLE exonuclease deficiency increase the likelihood of inactivating essential genes on chromosome arms harboring losses, leading to a synthetic lethal phenotype in highly aneuploid cells. Functional experiments demonstrated that POLE exonuclease activity is essential for the viability of highly aneuploid cancer cell lines but dispensable in diploid cells. These findings suggest that selective inhibition of POLE exonuclease activity may represent a promising therapeutic strategy for targeting highly aneuploid tumors. SIGNIFICANCE: An integrated approach using large-scale genomic analyses, probabilistic modeling and functional validation identified POLE exonuclease as a potential synthetic lethal target to overcome cancer aneuploidy.

Humans↗

Clearance of lower-pole stones following shock wave lithotripsy: effect of the infundibulopelvic angle.

OBJECTIVE: To assess the effect of anatomic factors, especially the angle of the lower-pole infundibulum, on stone clearance following shock wave lithotripsy (SWL) in order to determine selection criteria for percutaneous nephrolithotomy. METHODS: We retrospectively analyzed 116 patients with single lower-pole stones measuring 11-20 mm treated with SWL. Intravenous urograms were reviewed to measure the infundibulopelvic angle, the angle of the infundibulum to the vertical, and the anatomy of lower-pole calyces. RESULTS: The overall stone-free rate was 52%. Factors most closely associated with a stone-free status were obtuse infundibulopelvic angle, lack of calyceal distortion, and a large infundibular diameter. The infundibulopelvic angle was the only factor to attain significance in predicting stone-free status (p = 0.012). The size of the stone did not predict eventual stone-free status (p = 0.911), but larger stones were more likely to require intervention after SWL. CONCLUSION: For solitary lower-pole stones 11-20 mm in size, the angle of the lower-pole infundibulum as it relates to the pelvis plays a role in eventual stone clearance and should be taken into account before choosing a mode of treatment.

Adolescent↗