Bub1 Kinase Targets Sgo1 to Ensure Efficient Chromosome Biorientation in Budding Yeast Mitosis

During cell division all chromosomes must be segregated accurately to each daughter cell. Errors in this process give rise to aneuploidy, which leads to birth defects and is implicated in cancer progression. The spindle checkpoint is a surveillance mechanism that ensures high fidelity of chromosome segregation by inhibiting anaphase until all kinetochores have established bipolar attachments to spindle microtubules. Bub1 kinase is a core component of the spindle checkpoint, and cells lacking Bub1 fail to arrest in response to microtubule drugs and precociously segregate their DNA. The mitotic role(s) of Bub1 kinase activity remain elusive, and it is controversial whether this C-terminal domain of Bub1p is required for spindle checkpoint arrest. Here we make a detailed analysis of budding yeast cells lacking the kinase domain (bub1ΔK). We show that despite being able to arrest in response to microtubule depolymerisation and kinetochore-microtubule attachment defects, bub1ΔK cells are sensitive to microtubule drugs. This is because bub1ΔK cells display significant chromosome mis-segregation upon release from nocodazole arrest. bub1ΔK cells mislocalise Sgo1p, and we demonstrate that both the Bub1 kinase domain and Sgo1p are required for accurate chromosome biorientation after nocodazole treatment. We propose that Bub1 kinase and Sgo1p act together to ensure efficient biorientation of sister chromatids during mitosis

Nuc2p, a Subunit of the Anaphase-Promoting Complex, Inhibits Septation Initiation Network Following Cytokinesis in Fission Yeast

In most cell types, mitosis and cytokinesis are tightly coupled such that cytokinesis occurs only once per cell cycle. The fission yeast Schizosaccharomyces pombe divides using an actomyosin-based contractile ring and is an attractive model for the study of the links between mitosis and cytokinesis. In fission yeast, the anaphase-promoting complex/cyclosome (APC/C) and the septation initiation network (SIN), a spindle pole body (SPB)–associated GTPase-driven signaling cascade, function sequentially to ensure proper coordination of mitosis and cytokinesis. Here, we find a novel interplay between the tetratricopeptide repeat (TPR) domain–containing subunit of the APC/C, Nuc2p, and the SIN, that appears to not involve other subunits of the APC/C. Overproduction of Nuc2p led to an increase in the presence of multinucleated cells, which correlated with a defect in actomyosin ring maintenance and localization of the SIN component protein kinases Cdc7p and Sid1p to the SPBs, indicative of defective SIN signaling. Conversely, loss of Nuc2p function led to increased SIN signaling, characterized by the persistent localization of Cdc7p and Sid1p on SPBs and assembly of multiple actomyosin rings and division septa. Nuc2p appears to function independently of the checkpoint with FHA and ring finger (CHFR)–related protein Dma1p, a known inhibitor of the SIN in fission yeast. Genetic and biochemical analyses established that Nuc2p might influence the nucleotide state of Spg1p GTPase, a key regulator of the SIN. We propose that Nuc2p, by inhibiting the SIN after cell division, prevents further deleterious cytokinetic events, thereby contributing to genome stability

Substitution as a Mechanism for Genetic Robustness: The Duplicated Deacetylases Hst1p and Sir2p in Saccharomyces cerevisiae

How duplicate genes provide genetic robustness remains an unresolved question. We have examined the duplicated histone deacetylases Sir2p and Hst1p in Saccharomyces cerevisiae and find that these paralogs with non-overlapping functions can provide genetic robustness against null mutations through a substitution mechanism. Hst1p is an NAD+-dependent histone deacetylase that acts with Sum1p to repress a subset of midsporulation genes. However, hst1Δ mutants show much weaker derepression of target loci than sum1Δ mutants. We show that this modest derepression of target loci in hst1Δ strains occurs in part because Sir2p substitutes for Hst1p. Sir2p contributes to repression of the midsporulation genes only in the absence of Hst1p and is recruited to target promoters by a physical interaction with the Sum1 complex. Furthermore, when Sir2p associates with the Sum1 complex, the complex continues to repress in a promoter-specific manner and does not spread. Our results imply that after the duplication, SIR2 and HST1 subfunctionalized. The single SIR2/HST1 gene from Kluyveromyces lactis, a closely related species that diverged prior to the duplication, can suppress an hst1Δ mutation in S. cerevisiae as well as interact with Sir4p in S. cerevisiae. In addition, the existence of two distinct protein interaction domains for the Sir and Sum1 complexes was revealed through the analysis of a chimeric Sir2–Hst1 molecule. Therefore, the ability of Sir2p to substitute for Hst1p probably results from a retained but reduced affinity for the Sum1 complex that is a consequence of subfunctionalization via the duplication, degeneration, and complementation mechanism. These results suggest that the evolutionary path of duplicate gene preservation may be an important indicator for the ability of duplicated genes to contribute to genetic robustness

Effect of proliferating cell nuclear antigen ubiquitination and chromatin structure on the dynamic properties of the Y-family DNA polymerases

Y-family DNA polymerases carry out translesion synthesis past damaged DNA. DNA polymerases (pol) η and ι are usually uniformly distributed through the nucleus but accumulate in replication foci during S phase. DNA-damaging treatments result in an increase in S phase cells containing polymerase foci. Using photobleaching techniques, we show that polη is highly mobile in human fibroblasts. Even when localized in replication foci, it is only transiently immobilized. Although ubiquitination of proliferating cell nuclear antigen (PCNA) is not required for the localization of polη in foci, it results in an increased residence time in foci. polι is even more mobile than polη, both when uniformly distributed and when localized in foci. Kinetic modeling suggests that both polη and polι diffuse through the cell but that they are transiently immobilized for ∼150 ms, with a larger proportion of polη than polι immobilized at any time. Treatment of cells with DRAQ5, which results in temporary opening of the chromatin structure, causes a dramatic immobilization of polη but not polι. Our data are consistent with a model in which the polymerases are transiently probing the DNA/chromatin. When DNA is exposed at replication forks, the polymerase residence times increase, and this is further facilitated by the ubiquitination of PCNA

Relativistic Calculations for Incoherent Photoproduction of Eta Mesons

We develop a relativistic model for incoherent eta-photoproduction on nuclei. The elementary process is described using an effective Lagrangian containing photons, nucleons, the S11(1535) and D13(1520) nucleon resonances, and rho, omega, and eta mesons. The nucleon and eta wavefunctions are obtained from relativistic wave equations. Final-state interactions of the outgoing particles are included via optical potentials. The effects of these interactions are found to be large and lead to reduced cross sections.The incoherent cross sections for isovector transitions are much larger than those for isoscalar ones. The dominant contributions are those from the S11 and D13 resonances. We find important interference effects between the contributions of these two resonances. We give some detailed calculations for the cross sections for incoherent eta-photoproduction on C-12. We find that the incoherent cross section for a subset of states in the excitation energy region below 17 MeV are significantly larger than those of the coherent process. These cross sections may thus be accessible experimentally.Comment: 24 pages, 7 figures, to be published in Nuclear Physics

Securin Is Not Required for Chromosomal Stability in Human Cells

Abnormalities of chromosome number are frequently observed in cancers. The mechanisms regulating chromosome segregation in human cells are therefore of great interest. Recently it has been reported that human cells without an hSecurin gene lose chromosomes at a high frequency. Here we show that, after hSecurin knockout through homologous recombination, chromosome losses are only a short, transient effect. After a few passages hSecurin(−/−) cells became chromosomally stable and executed mitoses normally. This was unexpected, as the securin loss resulted in a persisting reduction of the sister-separating protease separase and inefficient cleavage of the cohesin subunit Scc1. Our data demonstrate that securin is dispensable for chromosomal stability in human cells. We propose that human cells possess efficient mechanisms to compensate for the loss of genes involved in chromosome segregation

Polarization properties of OH masers in AGB and post-AGB stars

Context: Ground-state OH maser emission from late-type stars is usually polarized and remains a powerful probe of the magnetic field structure in the outer regions of circumstellar envelopes if observed with high angular and spectral resolutions. Observations in all four Stokes parameters are quite sparse and this is the most thorough, systematic study published to date. Aims: We aim to determine polarization properties of OH masers in an extensive sample of stars that show copious mass loss and search for candidate objects that are well-suited for high angular resolution studies. Methods: Full-polarization observations of the OH 1612 and 1667 MHz maser transitions were carried out for a sample of 117 AGB and post-AGB stars. Several targets were also observed in the 1665 MHz line. Results: Polarized features occur in more than 75% of the sources in the complete sample and there is no intrinsic difference in the occurrence of polarized emission between the three classes of objects of different infrared characteristics. The highest fractional polarization occurs for the post-AGB+PN and the Mira+SR classes at 1612 and 1667 MHz, respectively. Differences in the fractional polarization between the sources at different evolutionary stages appear to be related to depolarization caused by blending. The alignment of the polarization angles at the extreme sides of the shell implies a regular structure of the magnetic field of a strength of 0.3-2.3 mG. Conclusions: Polarized OH maser features are widespread in AGB and post-AGB stars. The relationship between the circular and linear fractional polarizations for a representative sample are consistent with the standard models of polarization for the Zeeman splitting higher than the Doppler line width, whereas the polarized features are the sigma components.Comment: Accepted for publication in Astronomy and Astrophysics. A version with appendices (Tab. A.1 and Fig. B.1) can be downloaded from http://paulo.astro.uni.torun.pl/~pw/arXiv_

MAD3 Encodes a Novel Component of the Spindle Checkpoint Which Interacts with Bub3p, Cdc20p, and Mad2p

We show that MAD3 encodes a novel 58-kD nuclear protein which is not essential for viability, but is an integral component of the spindle checkpoint in budding yeast. Sequence analysis reveals two regions of Mad3p that are 46 and 47% identical to sequences in the NH2-terminal region of the budding yeast Bub1 protein kinase. Bub1p is known to bind Bub3p (Roberts et al. 1994) and we use two-hybrid assays and coimmunoprecipitation experiments to show that Mad3p can also bind to Bub3p. In addition, we find that Mad3p interacts with Mad2p and the cell cycle regulator Cdc20p. We show that the two regions of homology between Mad3p and Bub1p are crucial for these interactions and identify loss of function mutations within each domain of Mad3p. We discuss roles for Mad3p and its interactions with other spindle checkpoint proteins and with Cdc20p, the target of the checkpoint

Vector transition form factors of the NK∗→Θ+N K^*\to\Theta^+ and NKˉ∗→Σ10ˉ∗−N \bar{K}^*\to \Sigma_{\bar{10}}^{*-} in the SU(3) chiral quark-soliton model

We investigate the vector transition form factors of the nucleon and vector meson K^* to the pentaquark baryon Theta^+ within the framework of the SU(3) chiral quark-soliton model. We take into account the rotational 1/N_c and linear $m_{\rm s}$ corrections, assuming isospin symmetry and employing the symmetry-conserving quantization. It turns out that the leading-order contributions to the form factors are almost cancelled by the rotational corrections. Because of this, the flavor SU(3) symmetry-breaking terms yield sizeable effects on the vector transition form factors. In particular, the main contribution to the electric-like transition form factor comes from the wave-function corrections, which is a consequence of the generalized Ademollo-Gatto theorem derived in the present work. We estimate with the help of the vector meson dominance the K^* vector and tensor coupling constants for the Theta^+: $g_{K^{*}N\Theta}=0.74 - 0.87$ and $f_{K^{*}N\Theta}=0.53 - 1.16$. We argue that the outcome of the present work is consistent with the null results of the CLAS experiments in the reactions gamma n -> K^- Theta^+ and gamma p ->bar{K}^0 Theta^+. The results of the present work are also consistent with the recent experiments at KEK. In addition, we present the results of the $\Sigma_{\bar{10}}\to N\bar{K}^*$ transition form factors and its $\bar{K}^*N\Sigma_{\bar{10}}$ coupling constants.Comment: 26 pages, 10 figure

Diffuse-interface model for rapid phase transformations in nonequilibrium systems

A thermodynamic approach to rapid phase transformations within a diffuse interface in a binary system is developed. Assuming an extended set of independent thermodynamic variables formed by the union of the classic set of slow variables and the space of fast variables, we introduce finiteness of the heat and solute diffusive propagation at the finite speed of the interface advancing. To describe the transformation within the diffuse interface, we use the phase-field model which allows us to follow the steep but smooth change of phases within the width of diffuse interface. The governing equations of the phase-field model are derived for the hyperbolic model, model with memory, and for a model of nonlinear evolution of transformation within the diffuse-interface. The consistency of the model is proved by the condition of positive entropy production and by the outcomes of the fluctuation-dissipation theorem. A comparison with the existing sharp-interface and diffuse-interface versions of the model is given.Comment: 15 pages, regular article submitted to Physical Review