Esperanto for histones: CENP-A, not CenH3, is the centromeric histone H3 variant.

The first centromeric protein identified in any species was CENP-A, a divergent member of the histone H3 family that was recognised by autoantibodies from patients with scleroderma-spectrum disease. It has recently been suggested to rename this protein CenH3. Here, we argue that the original name should be maintained both because it is the basis of a long established nomenclature for centromere proteins and because it avoids confusion due to the presence of canonical histone H3 at centromeres.

Evidence for a susceptibility locus for schizophrenia on chromosome 6pter-p22.

We have performed linkage analysis in 186 multiplex families to search for genes that predispose to schizophrenia. Under a model with partially dominant inheritance, moderately broad disease definition and assuming locus homogeneity, a lod score of 3.2 was obtained for D6S260 on chromosome 6p23. A multipoint lod score of 3.9 (P = 2.3 x 10(-5)) was achieved when the F13A1 and D6S260 loci were analysed, allowing for locus heterogeneity. Adjusted for testing of multiple models, the multipoint lod score of 3.9 under heterogeneity has a genome wide significance of between 5-8%. The nonparametric affected pedigree member test provided results (P = 3 x 10(-4)) also supporting this finding. Our findings provide supportive evidence for a susceptibility locus for schizophrenia on distal chromosome 6p, and support a model of locus heterogeneity.

Microtubule dynamics and tubulin interacting proteins.

Microtubule dynamics are crucial in generation of the mitotic spindle. During the transition from interphase to mitosis, there is an increase in the frequency of microtubule catastrophes. Recent work has identified two proteins, Op 18/stathmin and XKCM1, which can promote microtubule catastrophes in vitro and in cells or extracts. Although both of these proteins share the ability to bind tubulin dimers, their mechanisms of action in destabilizing microtubules are distinct.

Ran stimulates spindle assembly by altering microtubule dynamics and the balance of motor activities.

The guanosine tri-phosphatase Ran stimulates assembly of microtubule spindles. However, it is not known what aspects of the microtubule cytoskeleton are subject to regulation by Ran in mitosis. Here we show that Ran-GTP stimulates microtubule assembly by increasing the rescue frequency of microtubules three- to eightfold. In addition to changing microtubule dynamics, Ran-GTP also alters the balance of motor activities, partly as a result of an increase in the amount of motile Eg5, a plus-end-directed microtubule motor that is essential for spindle formation. Thus, Ran regulates multiple processes that are involved in spindle assembly.

Control of microtubule dynamics by the antagonistic activities of XMAP215 and XKCM1 in Xenopus egg extracts.

Microtubules are dynamic polymers that move stochastically between periods of growth and shrinkage, a property known as dynamic instability. Here, to investigate the mechanisms regulating microtubule dynamics in Xenopus egg extracts, we have cloned the complementary DNA encoding the microtubule-associated protein XMAP215 and investigated the function of the XMAP215 protein. Immunodepletion of XMAP215 indicated that it is a major microtubule-stabilizing factor in Xenopus egg extracts. During interphase, XMAP215 stabilizes microtubules primarily by opposing the activity of the destabilizing factor XKCM1, a member of the kinesin superfamily. These results indicate that microtubule dynamics in Xenopus egg extracts are regulated by a balance between a stabilizing factor, XMAP215, and a destabilizing factor, XKCM1.

Bolus-tracking MRI with a simultaneous T1- and T2*-measurement.

The aim of this study was to propose and evaluate a methodology to analyze simultaneously acquired T2*-weighted dynamic susceptibility contrast (DSC) MRI and T(1)-weighted dynamic contrast enhanced (DCE) MRI data. Two generalized models of T2*-relaxation are proposed to account for tracer leakage, and a two-compartment exchange model is used to separate tracer in intra- and extravascular spaces. The methods are evaluated using data extracted from ROIs in three mice with subcutaneously implanted human colorectal tumors. Comparing plasma flow values obtained from DCE-MRI and DSC-MRI data defines a practical experimental paradigm to measure T2*-relaxivities, and reveals a factor of 15 between values in tissue and blood. Comparing mean transit time values obtained from DCE-MRI and DSC-MRI without leakage correction, indicates a significant reduction of susceptibility weighting in DSC-MRI during tracer leakage. A one-parameter gradient correction model provides a good approximation for this susceptibility loss, but redundancy of the parameter limits the practical potential of this model for DSC-MRI. Susceptibility loss is modeled more accurately with a variable T2*-relaxivity, which allows to extract new parameters that cannot be derived from DSC-MRI or DCE-MRI alone. They reflect the cellular and vessel geometry, and thus may lead to a more complete characterization of tissue structure.

XCTK2: a kinesin-related protein that promotes mitotic spindle assembly in Xenopus laevis egg extracts.

We used a peptide antibody to a conserved sequence in the motor domain of kinesins to screen a Xenopus ovary cDNA expression library. Among the clones isolated were two that encoded a protein we named XCTK2 for Xenopus COOH-terminal kinesin 2. XCTK2 contains an NH2-terminal globular domain, a central alpha-helical stalk, and a COOH-terminal motor domain. XCTK2 is similar to CTKs in other organisms and is most homologous to CHO2. Antibodies raised against XCTK2 recognize a 75-kD protein in Xenopus egg extracts that cosediments with microtubules. In Xenopus tissue culture cells, the anti-XCTK2 antibodies stain mitotic spindles as well as a subset of interphase nuclei. To probe the function of XCTK2, we have used an in vitro assay for spindle assembly in Xenopus egg extracts. Addition of antibodies to cytostatic factor-arrested extracts causes a 70% reduction in the percentage of bipolar spindles formed. XCTK2 is not required for maintenance of bipolar spindles, as antibody addition to preformed spindles has no effect. To further evaluate the function of XCTK2, we expressed XCTK2 in insect Sf-9 cells using the baculovirus expression system. When purified (recombinant XCTK2 is added to Xenopus egg extracts at a fivefold excess over endogenous levels) there is a stimulation in both the rate and extent of bipolar spindle formation. XCTK2 exists in a large complex in extracts and can be coimmunoprecipitated with two other proteins from extracts. XCTK2 likely plays an important role in the establishment and structural integrity of mitotic spindles.

The bipolar kinesin, KLP61F, cross-links microtubules within interpolar microtubule bundles of Drosophila embryonic mitotic spindles.

Previous genetic and biochemical studies have led to the hypothesis that the essential mitotic bipolar kinesin, KLP61F, cross-links and slides microtubules (MTs) during spindle assembly and function. Here, we have tested this hypothesis by immunofluorescence and immunoelectron microscopy (immunoEM). We show that Drosophila embryonic spindles at metaphase and anaphase contain abundant bundles of MTs running between the spindle poles. These interpolar MT bundles are parallel near the poles and antiparallel in the midzone. We have observed that KLP61F motors, phosphorylated at a cdk1/cyclin B consensus domain within the BimC box (BCB), localize along the length of these interpolar MT bundles, being concentrated in the midzone region. Nonphosphorylated KLP61F motors, in contrast, are excluded from the spindle and display a cytoplasmic localization. Immunoelectron microscopy further suggested that phospho-KLP61F motors form cross-links between MTs within interpolar MT bundles. These bipolar KLP61F MT-MT cross-links should be capable of organizing parallel MTs into bundles within half spindles and sliding antiparallel MTs apart in the spindle midzone. Thus we propose that bipolar kinesin motors and MTs interact by a "sliding filament mechanism" during the formation and function of the mitotic spindle.

A model for the proposed roles of different microtubule-based motor proteins in establishing spindle bipolarity.

BACKGROUND: In eukaryotes, assembly of the mitotic spindle requires the interaction of chromosomes with microtubules. During this process, several motor proteins that move along microtubules promote formation of a bipolar microtubule array, but the precise mechanism is unclear. In order to examine the roles of different motor proteins in building a bipolar spindle, we have used a simplified system in which spindles assemble around beads coated with plasmid DNA and incubated in extracts from Xenopus eggs. Using this system, we can study spindle assembly in the absence of paired cues, such as centrosomes and kinetochores, whose microtubule-organizing properties might mask the action of motor proteins. RESULTS: We blocked the function of individual motor proteins in the Xenopus extracts using specific antibodies. Inhibition of Xenopus kinesin-like protein 1 (Xklp1) led either to the dissociation of chromatin beads from microtubule arrays, or to collapsed microtubule bundles on beads. Inhibition of Eg5 resulted in monopolar microtubule arrays emanating from chromatin beads. Addition of antibodies against dynein inhibited the focusing of microtubule ends into spindle poles in a dose-dependent manner. Inhibition of Xenopus carboxy-terminal kinesin 2 (XCTK2) affected both pole formation and spindle stability. Co-inhibition of XCTK2 and dynein dramatically increased the severity of spindle pole defects. Inhibition of Xklp2 caused only minor spindle pole defects. CONCLUSIONS: Multiple microtubule-based motor activities are required for the bipolar organization of microtubules around chromatin beads, and we propose a model for the roles of the individual motor proteins in this process.

Microtubule-based motor function in mitosis.

Microtubule-based motors are essential both for the proper assembly of the mitotic spindle and for chromosome segregation. Mitotic motors in the yeast Saccharomyces cerevisiae exhibit either overlapping or opposing activities in order to achieve proper spindle function, whereas the analysis of motors using vertebrate cytoplasmic extracts has revealed less functional redundancy. In several systems, biochemical, genetic and two-hybrid approaches have been used both to identify associated nonmotor proteins and to address the molecular mechanisms behind kinetochore movements during chromosome alignment and segregation.

Partial tetrasomy 12pter-12p12.3 in a girl with Pallister-Killian syndrome: extraordinary finding of an analphoid, inverted duplicated marker.

Cytogenetic analysis in a girl with multiple congenital anomalies indicating Pallister-Killian syndrome (PKS) showed a supernumerary marker chromosome in 1/76 lymphocytes and 34/75 fibroblast metaphases. GTG-banding pattern was consistent with the chromosomal region 12pter-12q11. While fluorescence-in-situ hybridisation (FISH) with a whole chromosome 12 painting probe confirmed the origin of the marker, a chromosome 12 specific alpha-satellite probe did not hybridise to it. FISH analysis with a specific subtelomeric probe 12p showed hybridisation to both ends of the marker chromosome. High-resolution multicolour-banding (MCB) studies revealed the marker to be a der(12)(pter-->p12.3::p12.3-->pter). Summarising the FISH information, we defined the marker as an inverted duplication of 12pter-12p12.3 leading to partial tetrasomy of chromosome 12p. In skin fibroblasts, cultured at the patient's age of 1 year and 9 years, the marker chromosome was found in similar frequencies, even after several culture passages. Therefore, we consider the marker to have a functional centromere although it lacks detectable centromeric alpha-satellite sequences. To the best of our knowledge, this is the first proven analphoid marker of chromosome 12. Molecular genetic studies indicated that this marker is of paternal origin. The finding of partial tetrasomy 12pter-12p12.3 in our PKS patient allows to narrow down the critical region for PKS.

Microtubule dynamics in Xenopus egg extracts.

The organization and function of microtubules change dramatically during the cell cycle. At the onset of mitosis, a radial array of microtubules is broken down and reorganized into a bipolar spindle. This event requires changes in the dynamic behavior of individual microtubules. Through the use of Xenopus laevis egg extracts, a number of proteins affecting microtubule behavior have been identified. Recently, progress has also been made towards understanding how the activities of such microtubule-affecting proteins are regulated in a cell cycle-dependent manner. It is hoped that understanding how microtubule behavior is controlled during the cell cycle in vitro may illuminate the role of microtubule dynamics in various cellular processes.

Effects of pyrethroid insecticides on operant responding maintained by food.

The pyrethroids are potent insecticides with low concomitant mammalian lethality when compared with other major insecticides. While high doses can lead to hyperactivity, tremors, convulsion and death, low doses have not been as well studied. Since operant behavior can be a sensitive measure of CNS function, male Holtzman rats were trained on a VR25 schedule maintained by 45 mg food pellets. Rats were injected IP with one of four different technical grade pyrethroids: permethrin, allethrin, deltamethrin and fenvalerate. All agents were effective in reducing operant responding and did so in a dose-dependent manner at levels 10 to 100 times below their LD50 values. Time course studies indicated a relatively short duration of action for the Type I agents of less than 60 min for permethrin and 15 min for allethrin. Type II agents were generally effective for greater than 60 min. Results of these studies indicate that operant responding maintained by food is a sensitive measure of the behaviorally disruptive effects of subconvulsive doses of pyrethroids.

The CODATA/IUIS Hybridoma Data Bank: development of a hybrid system to handle complex data relationships.

System design for the Hybridoma Data Bank, a database of comprehensive information on immunoreagents for use by scientists in diverse disciplines, is described. Unique problems include: use of nomenclature from diverse fields that is neither static nor standard; the need for two representations of the database--textual for readability and numeric for complex search capabilities, analysis and data compression; and a method of translating between the two representations of the database.

Cooperative vendor selection saves money, helps improve care.

A cooperative effort between physicians and facility administrators with respect to purchase of materials and services can reduce costs dramatically and provide clinical benefits, as well. At Hamot Medical Center in Erie, Pa., orthopaedic physicians and hospital administrators developed a process for selecting a single source of implants used in total joint replacement procedures. As a result, the medical center was able to enter into a volume purchasing contract anticipated to save the medical center about $630,000 over the two-year period of the contract.

Meningioma progression in mice triggered by Nf2 and Cdkn2ab inactivation.

Aggressive variants of meningiomas (WHO grade II and III) represent up to 30% of those tumors that are among the most common primary central nervous system tumors in adults. Currently, there is no effective treatment for grade-II and -III meningiomas, the main treatment remaining surgical excision. Genetic studies have highlighted two main events associated with meningioma progression: an increase of chromosomal instability in tumors with NF2 inactivation and homozygous deletions or point mutations of the CDKN2AB locus. In this study we demonstrated that in mice, in addition to bi-allelic Nf2 inactivation, homozygous and heterozygous Adenovirus Cre-mediated Cdkn2ab deletions lead to increased meningioma frequency (72% and 50%, respectively) with a shorter latency (3.5 and 7.8 months, respectively) compared with control cohorts and induce grade II/III meningioma progression with an incidence of 34% and 28%, respectively. Moreover, Cdkn2ab inactivation in arachnoidal cells was associated with decreased senescence compared with Nf2(-/-) and wild-type arachnoidal cells in vitro. We have established three mouse meningioma cell lines and generated a syngenic orthotopic meningioma mouse model with 50-100% grade-II/III meningiomas after reimplantation. Comparative genomic hybridization of four meningiomas from Cdkn2ab homozygous mice and three cell cultures revealed the absence of unbalanced chromosomal segments in tumors and several chromosome imbalances in cell cultures. In addition, we were able to detect meningiomas by using bioluminescence and to evaluate tumor vascular permeability by dynamic magnetic resonance imaging. These results show that Nf2 and Cdkn2ab cooperate to promote meningioma progression in mice. The short latency of tumor development and the ability to derive grade II/III meningioma cell cultures are key aspects of this model to promote its use in pre-clinical drug testing.