scholarly journals FINE STRUCTURE OF DIVISION IN CILIATE PROTOZOA

1967 ◽  
Vol 34 (2) ◽  
pp. 463-481 ◽  
Author(s):  
R. A. Jenkins
Keyword(s):  
Electron Microscopy ◽  
Nuclear Envelope ◽  
Osmium Tetroxide ◽  
Distinctive Features ◽  
Chromosome Alignment ◽  
Mass Cultures ◽  
Spindle Microtubules ◽  
Spindle Elongation ◽  
And Function ◽  
Ciliate Protozoa

The mitotic, micronuclear division of the heterotrichous genus Blepharisma has been studied by electron microscopy. Dividing ciliates were selected from clone-derived mass cultures and fixed for electron microscopy by exposure to the vapor of 2% osmium tetroxide; individual Blepharisma were encapsulated and sectioned. Distinctive features of the mitosis are the presence of an intact nuclear envelope during the entire process and the absence of centrioles at the polar ends of the micronuclear figures. Spindle microtubules (SMT) first appear in advance of chromosome alignment, become more numerous and precisely aligned by metaphase, lengthen greatly in anaphase, and persist through telophase. Distinct chromosomal and continuous SMT are present. At telophase, daughter nuclei are separated by a spindle elongation of more than 40 µ, and a new nuclear envelope is formed in close apposition to the chromatin mass of each daughter nucleus and excludes the great amount of spindle material formed during division. The original nuclear envelope which has remained structurally intact then becomes discontinuous and releases the newly formed nucleus into the cytoplasm. The micronuclear envelope seems to lack the conspicuous pores that are typical of nuclear envelopes. The morphology, size, formation, and function of SMT and the nature of micronuclear division are discussed.

scholarly journals The structure of the yeast Ctf3 complex

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Stephen M Hinshaw ◽  
Andrew N Dates ◽  
Stephen C Harrison
Keyword(s):  
Electron Microscopy ◽  
Cell Cycle ◽  
High Resolution ◽  
Atomic Model ◽  
Kinetochore Assembly ◽  
Cryo Electron Microscopy ◽  
Molecular Interpretation ◽  
Spindle Microtubules ◽  
And Function ◽  
Assembly Site

Kinetochores are the chromosomal attachment points for spindle microtubules. They are also signaling hubs that control major cell cycle transitions and coordinate chromosome folding. Most well-studied eukaryotes rely on a conserved set of factors, which are divided among two loosely-defined groups, for these functions. Outer kinetochore proteins contact microtubules or regulate this contact directly. Inner kinetochore proteins designate the kinetochore assembly site by recognizing a specialized nucleosome containing the H3 variant Cse4/CENP-A. We previously determined the structure, resolved by cryo-electron microscopy (cryo-EM), of the yeast Ctf19 complex (Ctf19c, homologous to the vertebrate CCAN), providing a high-resolution view of inner kinetochore architecture (Hinshaw and Harrison, 2019). We now extend these observations by reporting a near-atomic model of the Ctf3 complex, the outermost Ctf19c sub-assembly seen in our original cryo-EM density. The model is sufficiently well-determined by the new data to enable molecular interpretation of Ctf3 recruitment and function.

scholarly journals A unique kinesin-8 surface loop provides specificity for chromosome alignment

2014 ◽  
Vol 25 (21) ◽  
pp. 3319-3329 ◽  
Author(s):  
Haein Kim ◽  
Cindy Fonseca ◽  
Jason Stumpff
Keyword(s):  
Motor Domain ◽  
Common Mechanism ◽  
Surface Loop ◽  
C Terminus ◽  
Chromosome Alignment ◽  
Spindle Microtubules ◽  
And Function ◽  
Positively Charged ◽  
Mitotic Chromatin

Microtubule length control is essential for the assembly and function of the mitotic spindle. Kinesin-like motor proteins that directly attenuate microtubule dynamics make key contributions to this control, but the specificity of these motors for different subpopulations of spindle microtubules is not understood. Kif18A (kinesin-8) localizes to the plus ends of the relatively slowly growing kinetochore fibers (K-fibers) and attenuates their dynamics, whereas Kif4A (kinesin-4) localizes to mitotic chromatin and suppresses the growth of highly dynamic, nonkinetochore microtubules. Although Kif18A and Kif4A similarly suppress microtubule growth in vitro, it remains unclear whether microtubule-attenuating motors control the lengths of K-fibers and nonkinetochore microtubules through a common mechanism. To address this question, we engineered chimeric kinesins that contain the Kif4A, Kif18B (kinesin-8), or Kif5B (kinesin-1) motor domain fused to the C-terminal tail of Kif18A. Each of these chimeric kinesins localizes to K-fibers; however, K-fiber length control requires an activity specific to kinesin-8s. Mutational studies of Kif18A indicate that this control depends on both its C-terminus and a unique, positively charged surface loop, called loop2, within the motor domain. These data support a model in which microtubule-attenuating kinesins are molecularly “tuned” to control the dynamics of specific subsets of spindle microtubules.

scholarly journals The structure of the yeast Ctf3 complex

2019 ◽  
Author(s):  
Stephen M. Hinshaw ◽  
Andrew N. Dates ◽  
Stephen C. Harrison
Keyword(s):  
Electron Microscopy ◽  
Cell Cycle ◽  
High Resolution ◽  
Atomic Model ◽  
Kinetochore Assembly ◽  
Cryo Electron Microscopy ◽  
Molecular Interpretation ◽  
Spindle Microtubules ◽  
And Function ◽  
Assembly Site

ABSTRACTKinetochores are the chromosomal attachment points for spindle microtubules. They are also signaling hubs that control major cell cycle transitions and coordinate chromosome folding. Most well-studied eukaryotes rely on a conserved set of factors, which are divided among two loosely-defined groups, for these functions. Outer kinetochore proteins contact microtubules or regulate this contact directly. Inner kinetochore proteins designate the kinetochore assembly site by recognizing a specialized nucleosome containing the H3 variant Cse4/CENP-A. We previously determined the structure, resolved by cryo-electron microscopy (cryo-EM), of the yeast Ctf19 complex (Ctf19c, homologous to the vertebrate CCAN), providing a high-resolution view of inner kinetochore architecture. We now extend these observations by reporting a near-atomic model of the Ctf3 complex, the outermost Ctf19c sub-assembly seen in our original cryo-EM density. The model is sufficiently well-determined by the new data to enable molecular interpretation of Ctf3 recruitment and function.

Filaments in Fibroblast in Pulmonary Alveolar Wall

1972 ◽  
Vol 30 ◽  
pp. 248-249
Author(s):  
Kuen-Shan Hung ◽  
M. Sue Hertweck ◽  
John D. Hardy ◽  
Clayton G. Loosli
Keyword(s):  
Electron Microscopy ◽  
Connective Tissue ◽  
Cell Body ◽  
Osmium Tetroxide ◽  
Mesenchymal Cells ◽  
Structure And Function ◽  
Alveolar Wall ◽  
And Function ◽  
Mouse Lungs ◽  
Interalveolar Septum

A small number of fibroblasts (alveolar septal cells, pulmonary interstitial cells, mesenchymal cells) are present in the alveolar areas of the adult lungs. These cells, like fibroblasts elsewhere, may be important in production and maintenance of connective tissue fibers found throughout the lung. Very little attention has been given to these cells in studies of the lungs. Therefore their precise structure and function are not clear. This report demonstrates filaments in the fibroblasts in the alveolar ducts and alveolar walls of the mouse lungs.The lungs were inflated with 2% glutaraldehyde, post-fixed with 1% osmium tetroxide and processed for electron microscopy. The cell body and processes of the alveolar fibroblast are located in the interalveolar septum (Fig. I).

scholarly journals Spindle microtubule differentiation and deployment during micronuclear mitosis in Paramecium.

1985 ◽  
Vol 101 (5) ◽  
pp. 1966-1976 ◽  
Author(s):  
J B Tucker ◽  
S A Mathews ◽  
K A Hendry ◽  
J B Mackie ◽  
D L Roche
Keyword(s):  
Nuclear Envelope ◽  
Early Stage ◽  
Microtubule Assembly ◽  
Membrane Association ◽  
Paramecium Tetraurelia ◽  
Spindle Microtubule ◽  
Spatial Control ◽  
Spindle Microtubules ◽  
Spindle Elongation ◽  
Anaphase B

Spindles underwent a 12-fold elongation before anaphase B was completed during the closed mitoses of micronuclei in Paramecium tetraurelia. Two main classes of spindle microtubules have been identified. A peripheral sheath of microtubules with diameters of 27-32 nm was found to be associated with the nuclear envelope and confined to the midportion of each spindle. Most of the other microtubules had diameters of approximately 24 nm and were present along the entire lengths of spindles. Nearly all of the 24-nm microtubules were eliminated from spindle midportions (largely because of microtubule disassembly) at a relatively early stage of spindle elongation. Disassembly of some of these microtubules also occurred at the ends of spindles. About 60% of the total microtubule content of spindles was lost at this stage. Most, perhaps all, peripheral sheath microtubules remained intact. Many of them detached from the nuclear envelope and regrouped to form a compact microtubule bundle in the spindle midportion. There was little, if any, further polymerization of 24-nm microtubules after the disassembly phase. Polymerization of microtubules with diameters of 27-32 nm continued as spindle elongation progressed. Most microtubules in the midportions of well-elongated spindles were constructed from 14-16 protofilaments. A few 24-nm microtubules with 13 protofilaments were also present. The implications of these findings for spatial control of microtubule assembly, disassembly, positioning, and membrane association, that apparently discriminate between microtubules with different protofilament numbers have been explored. The possibility that microtubule sliding occurs during spindle elongation has also been considered.

scholarly journals The role of the integral membrane nucleoporins Ndc1p and Pom152p in nuclear pore complex assembly and function

2006 ◽  
Vol 173 (3) ◽  
pp. 361-371 ◽  
Author(s):  
Alexis S. Madrid ◽  
Joel Mancuso ◽  
W. Zacheus Cande ◽  
Karsten Weis
Keyword(s):  
Electron Microscopy ◽  
Saccharomyces Cerevisiae ◽  
Nuclear Envelope ◽  
Lipid Bilayers ◽  
Nuclear Pore Complex ◽  
Transmembrane Proteins ◽  
Nuclear Pore ◽  
Large Channel ◽  
And Function

The nuclear pore complex (NPC) is a large channel that spans the two lipid bilayers of the nuclear envelope and mediates transport events between the cytoplasm and the nucleus. Only a few NPC components are transmembrane proteins, and the role of these proteins in NPC function and assembly remains poorly understood. We investigate the function of the three integral membrane nucleoporins, which are Ndc1p, Pom152p, and Pom34p, in NPC assembly and transport in Saccharomyces cerevisiae. We find that Ndc1p is important for the correct localization of nuclear transport cargoes and of components of the NPC. However, the role of Ndc1p in NPC assembly is partially redundant with Pom152p, as cells lacking both of these proteins show enhanced NPC disruption. Electron microscopy studies reveal that the absence of Ndc1p and Pom152p results in aberrant pores that have enlarged diameters and lack proteinaceous material, leading to an increased diffusion between the cytoplasm and the nucleus.

scholarly journals Direct experimental evidence for the existence, structural basis and function of astral forces during anaphase B in vivo

1991 ◽  
Vol 100 (2) ◽  
pp. 279-288 ◽  
Author(s):  
J.R. Aist ◽  
C.J. Bayles ◽  
W. Tao ◽  
M.W. Berns
Keyword(s):  
Critical Mass ◽  
Spindle Pole Body ◽  
Spindle Pole ◽  
Structural Basis ◽  
Mitotic Apparatus ◽  
Pole Body ◽  
Spindle Microtubules ◽  
Spindle Elongation ◽  
And Function ◽  
Anaphase B

The existence, structural basis and function of astral forces that are active during anaphase B in the fungus, Nectria haematococca, were revealed by experiments performed on living cells. When one of the two asters of a mitotic apparatus was damaged, the entire mitotic apparatus migrated rapidly in the direction of the opposing astral forces, showing that the force that accelerated spindle pole body separation in earlier experiments is located in the asters. When a strong solution of the antimicrotubule drug, MBC, was applied at anaphase A, tubulin immunocytochemistry showed that both astral and spindle microtubules were destroyed completely in less than a minute. As a result, separation of the spindle pole bodies during anaphase B almost stopped. By contrast, disrupting only the spindle microtubules with a laser microbeam increased the rate of spindle pole body separation more than fourfold. Taken together, these two experiments show that the astral forces are microtubule-dependent. When only one of the two or three bundles of spindle microtubules was broken at very early anaphase B, most such diminished spindles elongated at a normal rate, whereas others elongated at an increased rate. This result suggests that only a critical mass or number of spindle microtubules needs be present for the rate of spindle elongation to be fully governed, and that astral forces can accelerate the elongation of a weakened or diminished spindle.

scholarly journals The pseudo GTPase CENP-M drives human kinetochore assembly

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Federica Basilico ◽  
Stefano Maffini ◽  
John R Weir ◽  
Daniel Prumbaum ◽  
Ana M Rojas ◽  
...  
Keyword(s):  
Cell Division ◽  
Small Gtpases ◽  
Conformational Switching ◽  
Point Mutant ◽  
Kinetochore Assembly ◽  
Gtp Binding ◽  
Chromosome Alignment ◽  
Spindle Microtubules ◽  
And Function

Kinetochores, multi-subunit complexes that assemble at the interface with centromeres, bind spindle microtubules to ensure faithful delivery of chromosomes during cell division. The configuration and function of the kinetochore–centromere interface is poorly understood. We report that a protein at this interface, CENP-M, is structurally and evolutionarily related to small GTPases but is incapable of GTP-binding and conformational switching. We show that CENP-M is crucially required for the assembly and stability of a tetramer also comprising CENP-I, CENP-H, and CENP-K, the HIKM complex, which we extensively characterize through a combination of structural, biochemical, and cell biological approaches. A point mutant affecting the CENP-M/CENP-I interaction hampers kinetochore assembly and chromosome alignment and prevents kinetochore recruitment of the CENP-T/W complex, questioning a role of CENP-T/W as founder of an independent axis of kinetochore assembly. Our studies identify a single pathway having CENP-C as founder, and CENP-H/I/K/M and CENP-T/W as CENP-C-dependent followers.

scholarly journals The ultrastructure and timing of events in the nuclear cycle of the fungus Entomophaga aulicae

1988 ◽  
Vol 90 (3) ◽  
pp. 501-516
Author(s):  
FAYE MURRIN ◽  
WILLIAM NEWCOMB ◽  
I. BRENT HEATH
Keyword(s):  
Nuclear Envelope ◽  
Linear Array ◽  
Nuclear Division ◽  
Metaphase Plate ◽  
Serial Sections ◽  
Free Zone ◽  
Nuclear Cycle ◽  
Full Complement ◽  
Spindle Microtubules ◽  
Spindle Elongation

The ultrastructure of the mitotic nuclear division cycle of the fungus Entomophaga aulicae was studied from serial sections of hyphal tips and protoplasts. The extranuclear bar-shaped nucleus- associated organelle (NAO) remained associated with the persistent nuclear envelope throughout. Prior to spindle formation, a patch of intranuclear NAO-associated chromatin detached from the nuclear envelope to yield a chromatin free zone containing fine filaments and a linear array of presumptive kinetochores. Early metaphase spindles less than 1μm in length were characterized by a ‘fused’ metaphase plate consisting of kinetochore-associated chromatin and a full complement of at least 15 kinetochore microtubules per half-spindle, while most of the chromatin was remote from the intranuclear spindle. Analysis of the distribution of antiparallel spindle microtubules indicated that polar separation and concomitant spindle elongation through metaphase were not accompanied by intermicrotubule sliding. Anaphase exhibited extensive decondensation of the large patches of condensed chromatin characteristic of all other stages. In a logarithmically growing protoplast population all nuclei contained spindle microtubules, with metaphase occupying approximately 66% of the nuclear cycle time. The calculated genome size of 4.3 pg, and average DNA content per chromosome of 0.3 pg, are extremely high for fungi.

scholarly journals Functional Coordination of Three Mitotic Motors inDrosophila Embryos

2000 ◽  
Vol 11 (1) ◽  
pp. 241-253 ◽  
Author(s):  
David J. Sharp ◽  
Heather M. Brown ◽  
Mijung Kwon ◽  
Gregory C. Rogers ◽  
Gina Holland ◽  
...  
Keyword(s):  
Nuclear Envelope ◽  
Cytoplasmic Dynein ◽  
Spindle Pole ◽  
Nuclear Envelope Breakdown ◽  
Mitotic Motors ◽  
Balance Of Forces ◽  
Spindle Elongation ◽  
And Function ◽  
Anaphase B ◽  
Drosophila Embryos

It is well established that multiple microtubule-based motors contribute to the formation and function of the mitotic spindle, but how the activities of these motors interrelate remains unclear. Here we visualize spindle formation in living Drosophila embryos to show that spindle pole movements are directed by a temporally coordinated balance of forces generated by three mitotic motors, cytoplasmic dynein, KLP61F, and Ncd. Specifically, our findings suggest that dynein acts to move the poles apart throughout mitosis and that this activity is augmented by KLP61F after the fenestration of the nuclear envelope, a process analogous to nuclear envelope breakdown, which occurs at the onset of prometaphase. Conversely, we find that Ncd generates forces that pull the poles together between interphase and metaphase, antagonizing the activity of both dynein and KLP61F and serving as a brake for spindle assembly. During anaphase, however, Ncd appears to have no effect on spindle pole movements, suggesting that its activity is down-regulated at this time, allowing dynein and KLP61F to drive spindle elongation during anaphase B.

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