scholarly journals LOCAL REDUCTION OF SPINDLE FIBER BIREFRINGENCE IN LIVING NEPHROTOMA SUTURALIS (LOEW) SPERMATOCYTES INDUCED BY ULTRAVIOLET MICROBEAM IRRADIATION

1965 ◽  
Vol 25 (1) ◽  
pp. 95-117 ◽  
Author(s):  
Arthur Forer
Keyword(s):  
Meiotic Division ◽  
Mitotic Spindle ◽  
Constant Velocity ◽  
Irradiation Effects ◽  
Spindle Fiber ◽  
Ultraviolet Microbeam ◽  
Local Reduction ◽  
Fiber Birefringence ◽  
Oriented Material ◽  
Spindle Fibers

Irradiation of the mitotic spindle in living Nephrotoma suturalis (Loew) spermatocytes with an ultraviolet microbeam of controlled dose produced a localized area of reduced birefringence in the spindle fibers. The birefringence was reduced only at the site irradiated, and only on the spindle fibers irradiated. Areas of reduced birefringence, whether produced during metaphase or during anaphase, immediately began to move toward the pole in the direction of the chromosomal fiber, even though the associated chromosomes did not necessarily move poleward. Both the poleward and the chromosomal sides of the area of reduced birefringence on each chromosomal fiber moved poleward with about the same, constant, velocity. On the average, the areas of reduced birefringence moved poleward with about the same velocities as did the chromosomes during anaphase. The area of reduced birefringence was interpreted as a region in which most, though not necessarily all, of the previously oriented material was disoriented by the irradiation. The poleward movement of the areas of reduced birefringence indicates that the spindle fibers are not static, nonchangeable structures. The poleward movement possibly represents the manner in which the birefringent spindle fibers normally become organized. All the experiments reported were on primary spermatocytes which completed the second meiotic division subsequent to the experimentation. Since both the irradiated and the control cells completed the two meiotic divisions, the movement and irradiation effects studied in the first division were nondegenerative.

scholarly journals UV microbeam irradiations of the mitotic spindle. II. Spindle fiber dynamics and force production.

1990 ◽  
Vol 111 (4) ◽  
pp. 1505-1518 ◽  
Author(s):  
T P Spurck ◽  
O G Stonington ◽  
J A Snyder ◽  
J D Pickett-Heaps ◽  
A Bajer ◽  
...  
Keyword(s):  
Mitotic Spindle ◽  
Force Production ◽  
The Other ◽  
Force Generation ◽  
Spindle Fiber ◽  
Spindle Microtubule ◽  
Microtubule Stability ◽  
Fiber Dynamics ◽  
Spindle Fibers

Metaphase and anaphase spindles in cultured newt and PtK1 cells were irradiated with a UV microbeam (285 nM), creating areas of reduced birefringence (ARBs) in 3 s that selectively either severed a few fibers or cut across the half spindle. In either case, the birefringence at the polewards edge of the ARB rapidly faded polewards, while it remained fairly constant at the other, kinetochore edge. Shorter astral fibers, however, remained present in the enlarged ARB; presumably these had not been cut by the irradiation. After this enlargement of the ARB, metaphase spindles recovered rapidly as the detached pole moved back towards the chromosomes, reestablishing spindle fibers as the ARB closed; this happened when the ARB cut a few fibers or across the entire half spindle. We never detected elongation of the cut kinetochore fibers. Rather, astral fibers growing from the pole appeared to bridge and then close the ARB, just before the movement of the pole toward the chromosomes. When a second irradiation was directed into the closing ARB, the polewards movement again stopped before it restarted. In all metaphase cells, once the pole had reestablished connection with the chromosomes, the unirradiated half spindle then also shortened to create a smaller symmetrical spindle capable of normal anaphase later. Anaphase cells did not recover this way; the severed pole remained detached but the chromosomes continued a modified form of movement, clumping into a telophase-like group. The results are discussed in terms of controls operating on spindle microtubule stability and mechanisms of mitotic force generation.

scholarly journals Motile kinetochores and polar ejection forces dictate chromosome position on the vertebrate mitotic spindle

1994 ◽  
Vol 124 (3) ◽  
pp. 223-233 ◽  
Author(s):  
CL Rieder ◽  
ED Salmon
Keyword(s):  
Mitotic Spindle ◽  
Constant Velocity ◽  
Molecular Mechanisms ◽  
Dynamic Instability ◽  
Microtubule Dynamic ◽  
Pole Motion ◽  
Microtubule Motors ◽  
Chromosome Position ◽  
Pulling Forces

We argue that hypotheses for how chromosomes achieve a metaphase alignment, that are based solely on a tug-of-war between poleward pulling forces produced along the length of opposing kinetochore fibers, are no longer tenable for vertebrates. Instead, kinetochores move themselves and their attached chromosomes, poleward and away from the pole, on the ends of relatively stationary but shortening/elongating kinetochore fiber microtubules. Kinetochores are also "smart" in that they switch between persistent constant-velocity phases of poleward and away from the pole motion, both autonomously and in response to information within the spindle. Several molecular mechanisms may contribute to this directional instability including kinetochore-associated microtubule motors and kinetochore microtubule dynamic instability. The control of kinetochore directional instability, to allow for congression and anaphase, is likely mediated by a vectorial mechanism whose magnitude and orientation depend on the density and orientation or growth of polar microtubules. Polar microtubule arrays have been shown to resist chromosome poleward motion and to push chromosomes away from the pole. These "polar ejection forces" appear to play a key role in regulating kinetochore directional instability, and hence, positions achieved by chromosomes on the spindle.

scholarly journals The role of the ameiotic1 gene in the initiation of meiosis and in subsequent meiotic events in maize.

Genetics ◽  
1993 ◽  
Vol 135 (4) ◽  
pp. 1151-1166 ◽  
Author(s):  
I Golubovskaya ◽  
Z K Grebennikova ◽  
N A Avalkina ◽  
W F Sheridan
Keyword(s):  
Meiotic Division ◽  
Higher Plants ◽  
Gene Interaction ◽  
Microtubule Organizing Center ◽  
Spindle Fiber ◽  
Chromosome Synapsis ◽  
Organizing Center ◽  
Relationship Of ◽  
The Relationship

Abstract Understanding the initiation of meiosis and the relationship of this event with other key cytogenetic processes are major goals in studying the genetic control of meiosis in higher plants. Our genetic and structural analysis of two mutant alleles of the ameiotic1 gene (am1 and am1-praI) suggest that this locus plays an essential role in the initiation of meiosis in maize. The product of the ameiotic1 gene affects an earlier stage in the meiotic sequence than any other known gene in maize and is important for the irreversible commitment of cells to meiosis and for crucial events marking the passage from premeiotic interphase into prophase I including chromosome synapsis. It appears that the period of ameiotic1 gene function in meiosis at a minimum covers the interval from some point during premeiotic interphase until the early zygotene stage of meiosis. To study the interaction of genes in the progression of meiosis, several double meiotic mutants were constructed. In these double mutants (i) the ameiotic1 mutant allele was brought together with the meiotic mutation (afd1) responsible for the fixation of centromeres in meiosis; and with the mutant alleles of the three meiotic genes that control homologous chromosome segregation (dv1, ms43 and ms28), which impair microtubule organizing center organization, the orientation of the spindle fiber apparatus, and the depolymerization of spindle filaments after the first meiotic division, respectively; (ii) the afd1 mutation was combined with two mutations (dsy1 and as1) affecting homologous pairing; (iii) the ms43 mutation was combined with the as1, the ms28 and the dv1 mutations; and (iv) the ms28 mutation was combined with the dv1 mutation and the ms4 (polymitotic1) mutations. An analysis of gene interaction in the double mutants led us to conclude that the ameiotic1 gene is epistatic over the afd1, the dv1, the ms43 and the ms28 genes but the significance of this relationship requires further analysis. The afd gene appears to function from premeiotic interphase throughout the first meiotic division, but it is likely that its function begins after the start of the ameiotic1 gene expression. The afd1 gene is epistatic over the two synaptic mutations dsy1 and as1 and also over the dv1 mutation. The new ameiotic*-485 and leptotene arrest*-487 mutations isolated from an active Robertson's Mutator stocks take part in the control of the initiation of meiosis.

scholarly journals Visualization of Mad2 Dynamics at Kinetochores, along Spindle Fibers, and at Spindle Poles in Living Cells

2000 ◽  
Vol 150 (6) ◽  
pp. 1233-1250 ◽  
Author(s):  
B.J. Howell ◽  
D.B. Hoffman ◽  
G. Fang ◽  
A.W. Murray ◽  
E.D. Salmon
Keyword(s):  
Chromosome Segregation ◽  
Binding Sites ◽  
Metaphase Plate ◽  
Atp Depletion ◽  
Anaphase Promoting Complex ◽  
Spindle Fiber ◽  
Spindle Poles ◽  
Anaphase Onset ◽  
Kinetochore Function ◽  
Spindle Fibers

The spindle checkpoint prevents errors in chromosome segregation by inhibiting anaphase onset until all chromosomes have aligned at the spindle equator through attachment of their sister kinetochores to microtubules from opposite spindle poles. A key checkpoint component is the mitotic arrest–deficient protein 2 (Mad2), which localizes to unattached kinetochores and inhibits activation of the anaphase-promoting complex (APC) through an interaction with Cdc20. Recent studies have suggested a catalytic model for kinetochore function where unattached kinetochores provide sites for assembling and releasing Mad2–Cdc20 complexes, which sequester Cdc20 and prevent it from activating the APC. To test this model, we examined Mad2 dynamics in living PtK1 cells that were either injected with fluorescently labeled Alexa 488-XMad2 or transfected with GFP-hMAD2. Real-time, digital imaging revealed fluorescent Mad2 localized to unattached kinetochores, spindle poles, and spindle fibers depending on the stage of mitosis. FRAP measurements showed that Mad2 is a transient component of unattached kinetochores, as predicted by the catalytic model, with a t1/2 of ∼24–28 s. Cells entered anaphase ∼10 min after Mad2 was no longer detectable on the kinetochores of the last chromosome to congress to the metaphase plate. Several observations indicate that Mad2 binding sites are translocated from kinetochores to spindle poles along microtubules. First, Mad2 that bound to sites on a kinetochore was dynamically stretched in both directions upon microtubule interactions, and Mad2 particles moved from kinetochores toward the poles. Second, spindle fiber and pole fluorescence disappeared upon Mad2 disappearance at the kinetochores. Third, ATP depletion resulted in microtubule-dependent depletion of Mad2 fluorescence at kinetochores and increased fluorescence at spindle poles. Finally, in normal cells, the half-life of Mad2 turnover at poles, 23 s, was similar to kinetochores. Thus, kinetochore-derived sites along spindle fibers and at spindle poles may also catalyze Mad2 inhibitory complex formation.

scholarly journals THE FINE STRUCTURE OF THE MID-BODY OF THE RAT ERYTHROBLAST

1962 ◽  
Vol 13 (1) ◽  
pp. 109-115 ◽  
Author(s):  
Robert C. Buck ◽  
James M. Tisdale
Keyword(s):  
Bone Marrow ◽  
Plasma Membrane ◽  
Fine Structure ◽  
Electron Microscope ◽  
Cleavage Furrow ◽  
Intercellular Bridge ◽  
Spindle Fiber ◽  
Thin Sections ◽  
Rat Bone ◽  
Spindle Fibers

The development of the mid-body has been studied in mitotic erythroblasts of the rat bone marrow by means of thin sections examined with the electron microscope. A differentiated region on the continuous spindle fibers, consisting of a localized increase in density, is observed at the equatorial plane. The mid-body seems to develop by the aggregation of such denser lengths of spindle fiber. Its appearance precedes that of the cleavage furrow. A plate-like arrangement of fibrillary material lies transversely across the telophase intercellular bridge. Later, this material becomes amorphous and assumes the form of a dense ring closely applied to a ridge in the plasma membrane encircling the middle of the bridge. Although the mid-body forms in association with the spindle fibers, it is a structurally distinct part, and the changes which it undergoes are not shared by the rest of the bundle of continuous fibers.

scholarly journals Distribution of tyrosinated and nontyrosinated alpha-tubulin during mitosis.

1986 ◽  
Vol 102 (3) ◽  
pp. 1118-1126 ◽  
Author(s):  
G G Gundersen ◽  
J C Bulinski
Keyword(s):  
Mitotic Spindle ◽  
Cultured Cells ◽  
Alpha Tubulin ◽  
Restricted Distribution ◽  
C Terminus ◽  
Major Species ◽  
A Minor ◽  
Spindle Function ◽  
Mitotic Cells ◽  
Spindle Fibers

The C-terminus of alpha-tubulin undergoes a reversible posttranslational tyrosination/detyrosination. The distributions of the tyrosinated (Tyr) and nontyrosinated (Glu) species during mitosis of cultured cells have been investigated by immunofluorescence using antibodies directed against the C-terminus of either Tyr or Glu tubulin. The distribution of Tyr tubulin differed from that of Glu tubulin at each stage of mitosis; in general, the distribution of Tyr tubulin was similar to that of total tubulin, whereas Glu tubulin had a more restricted distribution. The Glu species was found in half-spindle fibers but was not detected in astral fibers at any stage and was seen in the interzone only during telophase. These results were confirmed by a direct comparison of the distributions of Tyr and Glu tubulin in cells double-labeled with the two antibodies. Evidence for the occurrence of Tyr and Glu tubulin in each class of half-spindle fibers (kinetochore and polar) was obtained from the staining patterns of the two antibodies in cold-treated cells. Immunoblots of extracts prepared from synchronous mitotic cells showed that Glu tubulin was a minor species of the total tubulin in the spindle; no changes in the amount of either Tyr or Glu tubulin were detected at any stage of mitosis. These results show that Tyr tubulin is the major species in the mitotic spindle and is found in all classes of spindle fibers, whereas Glu tubulin is present in small amounts and shows a more restricted distribution. The presence of two biochemically distinct forms of alpha-tubulin in the spindle may be important for spindle function.

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