Disruption of CENP antigen function perturbs dynein anchoring to the mitotic kinetochore

Injection of purified autoantibodies against human centromeric proteins into HeLa cells during interphase disrupts the organization of the kinetochore and interferes with chromosomal movements during the subsequent mitosis even though the chromosomes retain the ability to bind microtubules. We have investigated the hypothesis that this phenotype arises from effects on cytoplasmic dynein, the microtubule motor protein. In previous experiments we found that introduction of anticentromere antibodies into cell nuclei during the G₁- or S-phases causes a prometaphase-like arrest, while injections during G₂-phase cause a metaphase arrest. We show here that, in both cases, the level of detectable cytoplasmic dynein at kinetochores is significantly decreased. In contrast, when injected cells were permitted to enter mitosis in the absence of microtubules (conditions where trilaminar kinetochores could be detected by electron microscopy), the intensity of dynein labeling on the kinetochores was identical to that seen in uninjected control cells exposed to colcemid. Therefore, the loss of dynein label on mitotic kinetochores was correlated both with the injection of anticentromere antibodies and with the presence of intact spindle microtubules. We suggest that the injection of anticentromere antibodies somehow weakens the association of dynein with the kinetochore, so that when microtubules are present, these motor molecules are pulled away from the kinetochores as they generate force. This model offers an explanation for the failure of chromosomes of injected cells to move normally in mitosis even though they have attached microtubules.     

Expressed Peromyscus maniculatus (Pema) MHC class I genes: evolutionary implications and the identification of a gene encoding a Qa1-like antigen

To gain insight into the evolution of rodent major histocompatibility complex (MHC) class I genes and identify important (conserved) nonclassical class I (class Ib) gene products and residues in these proteins, sixPeromyscus maniculatus MHC (Pema) class I cDNA clones were isolated and sequenced. FivePema class I cDNAs appeared most similar to mouse and rat classical class I (class Ia) genes. One exhibited highest similarity to anH2 class Ib gene,H2-T23 (encoding the Qa1 antigen). Phylogenetic trees constructed withPema, RT1, andH2 class I sequences suggested that the lineages of some rodent class Ib genes (e.g.,T23 andT24) originated prior toMus andPeromyscus speciation [>50 million years (My) ago]. Sequences of four Qa1-like proteins from three species permitted the identification of ten Qa1-specific amino acids. On the basis of molecular modeling, three residues showed the potential to interact with T-cell receptors and three residues (all corresponding to polymorphic positions among H2 class Ia proteins) were predicted to influence antigen binding. The recognition of mouse Qa1 proteins by a subset of T-cells in influenced by a locus,Qdm, which encodes the H2-D leader peptide. One of thePema class I cDNA clones classified asH2-K, D/L-like (class Ia) is predicted to encode an identical peptide, implying that an antigen binding protein (Qa1) and the antigen to which it binds (the product ofQdm) has been conserved for over 50 My. The nucleotide sequence data reported in this paper have been submitted to the GenBank nucleotide sequence database and have been assigned the accession numbers U12822 (Pm13), U12885 (Pm41), U12886 (Pm52), U12887 (Pm62), U16846 (Pm11), and U16847 (Pm53)     

Age-Dependent Impairment of Mitochondrial Function in Primate Brain

Abstract: It has been hypothesized that some of the functional impairments associated with aging are the result of increasing oxidative damage to mitochondrial DNA that produces defects in oxidative phosphorylation. To test this hypothesis, we examined the enzymes that catalyze oxidative phosphorylation in crude mitochondrial preparations from frontoparietal cortex of 20 rhesus monkeys (5-34 years old). Samples were assayed for complex I, complex II-III, complex IV, complex V, and citrate synthase activities. When enzyme activities were corrected for citrate synthase activities (to account for variable degrees of mitochondrial enrichment), linear regression analysis demonstrated a significant negative correlation of the activities of complex I (p < 0.002) and complex IV (p < 0.03) with age but no significant change in complex II-III or complex V activities. Relative to animals 6.9 ± 0.9 years old (n = 7), the citrate synthase-corrected activity of complex I was reduced by 17% in animals 22.5 ± 0.9 years old (n = 6) (p < 0.05) and by 22% in animals 30.7 ± 0.9 years old (n = 7) (p < 0.01). Similar age-related reductions in the activities of complexes I and IV were obtained when enzyme activities were corrected for complex II-III activity. These findings show an age-associated progressive impairment of mitochondrial complex I and complex IV activities in cerebral cortices of primates.     

Fine-scale mapping of physicochemical and microbial landscapes of the coral skeleton

The coral skeleton harbours a diverse community of bacteria and microeukaryotes exposed to light, O₂ and pH gradients, but how such physicochemical gradients affect the coral skeleton microbiome remains unclear. In this study, we employed chemical imaging of O₂ and pH, hyperspectral reflectance imaging and spatially resolved taxonomic and inferred functional microbiome characterization to explore links between the skeleton microenvironment and microbiome in the reef-building corals Porites lutea and Paragoniastrea benhami. The physicochemical environment was more stable in the deep skeleton, and the diversity and evenness of the bacterial community increased with skeletal depth, suggesting that the microbiome was stratified along the physicochemical gradients. The bulk of the coral skeleton was in a low O₂ habitat, whereas pH varied from pH 6–9 with depth. Physicochemical gradients of O₂ and pH of the coral skeleton explained the β-diversity of the bacterial communities, and skeletal layers that showed O₂ peaks had a higher relative abundance of endolithic algae, reflecting a link between the abiotic environment and the microbiome composition. Our study links the physicochemical, microbial and functional landscapes of the coral skeleton and provides new insights into the involvement of skeletal microbes in the coral holobiont metabolism.     

Sodium and potassium relations of Spergularia marina following N and P deprivation: results of short-term growth studies

In this, we consider the coordination of plant growth and ion acquisition, reporting the short-term adjustments of growth and K⁺ and Na⁺ relations which follow when plants are subject to a sudden deprivation of N and P. The plant used for the experiments, Spergularia marina (L.) Grieseb., is a small coastal halophyte, and the growth medium was 0.2 × modified seawater. By considering nutrients whose availability has not been changed, we report on an aspect of organismal integration which has received little attention either experimentally or in mathematical models. The studies are limited to the first 60 h after N and P deprivation in order to consider changes that, if they are not primary responses, are not temporally remote, passive adjustments. For growth analyses, plants were used approximately 30 days after germination and 16 days after transfer to solution culture. Random harvests were made at hourly invervals, and after 12 h, one-half of the plants were transferred to cultures without N or P. Tissue analyses were used to calculate relative growth rates, relative accumulation rates and net uptake rates. For comparison, isotope uptake studies using ⁴²K⁺ and ²²Na⁺ were conducted at 12, 36 and 60 h after deprivation. The effects on growth and biomass allocation were very rapid, detectable within 13 h. K⁺ transport also responded quickly, and from the beginning of the study, there was essentially no net translocation of K⁺ to the shoot. Isotope studies confirmed the responsiveness, with translocation reduced 33 and 90% after 12 and 36 h, respectively. Though Na⁺ adjustments were slower, they were coordinated with growth such that tissue concentrations in the N and P-deprived plants were comparable to those in the controls. We conclude that N and C are insufficient elements on which to build mathematical models useful to environmental physiologists. At a minimum, the incorporation of K⁺ relations in growth models would both allow the development of the osmotic potential needed to drive cell expansion, and provide a means to probe –experimentally as well as mathematically – the coordinating mechanisms of plant growth and resource management.     

SPERMATOGENESIS IN COLEOCHAETE PULVINATA (CHAROPHYCEAE): EARLY BLEPHAROPLAST DEVELOPMENT

Transmission electron microscopy of serial thin sections was used to reconstruct several early developmental stages of the blepharoplast in Coleochaete pulvinata spermatids. These were compared to published studies of blepharoplast development in Charales and the closest relatives of charophycean green algae among embryophytes, i.e., hornworts and liverworts. Bicentriolar centrosomes such as occur in bryophytes and fern allies were not observed in Coleochaete. Centriole replication in C. pulvinata was orthogonal as in Charales. The resulting two daughter centrioles were oriented perpendicularly and joined proximally by electron-dense material. Their orthogonal relationship was maintained throughout blepharoplast development by a massive, banded connective which appeared early. In spermatids of hornworts and liverworts, a multilayered structure (MLS) develops in association with two centrioles destined to become flagellar basal bodies. When the MLS of these lower land plants is sectioned at right angles to the long axis of the microtubular layer, the MLS is observed to lie beneath cross sections of both centrioles. In contrast, when developing MLSs of C. pulvinata and Charales are similarly sectioned, they occur beside a cross section of just one of the two centrioles. In C. pulvinata (as in other charophytes), MLS lamellae are oriented at a 90-degree angle to the long axis of the S₁ microtubules from the beginning. This contrasts with the 40–45 degree angle between the MLS lamellae and S₁ microtubules universally reported for archegoniates. In early C. pulvinata spermatids, spline microtubules are closely associated with an anterior mitochondrion having a low stromal density and few cristae. An anterior mitochondrion is typically associated with blepharoplast development in hornworts and liverworts, but has not previously been reported to occur in Coleochaete or any other charophycean alga. In Coleochaete, as in hornworts and liverworts, but unlike Charales, structure of mature blepharoplasts reflects early blepharoplast ontogeny. Very little change in positional relationships among blepharoplast components (flagella, connective, MLS) occurs during development. These character-state differences are of importance in cladistic analyses of charophycean algae and lower land plants.