Spindle assembly

The activated CDKl-cyclin B kinase finally phosphor-ylates a large number of proteins most of which are not well characterized, leading to chromosome condensation, nuclear envelope breakdown, spindle assembly, and chromosome segregation. 

A signal transduction pathway required for proper chromosome alignment during mitosis. The spindle assembly checkpoint is activated during mitosis in response to the presence of chromosomes that are not attached to spindle microtubules or that are not properly aligned at the metaphase plate. The spindle checkpoint... 

Wadsworth, P. (1993). Mitosis spindle assembly and chromosome motion. Curr. Opin. Cell Biol. 5, 123-128. 

The duration of the M phase is largely determined by the time necessary for the formation of a functional metaphase spindle and the correct alignment of all chromosomes in the metaphase plate. The spindle assembly checkpoint prevents the exit from the M phase before the proper alignment of all chromosomes into a metaphase plate in many cell types. This kind of control is already operational... 

Goncpy In the normally fertilized egg there is a centrosome. However, you are saying that the spindle is assembled using a non-centrosomal pathway in the onecell stage mouse embryo. This seems surprising to me in light of Rebecca Heald s work, which shows that the centrosomal pathway of spindle assembly is dominant over the non-centrosomal one (Heald et al 1997). 

Kalab P, Pu RT, Dasso M 1999 The ran GTPase regulates mitotic spindle assembly. Curr Biol 9 481-484... 

Goncyy What Bill showed is that Bazooka, Inscuteable and Pins are required for a 90° spindle rotation in Drosophila. In the one cell stage C. elegans embryo, it appears that PAR-3, the Bazooka homologue, is not required for the 90° rotation of centrosomes that precedes spindle assembly (Cheng et al 1995). 

Goncyy No. There is a 90° rotation of the centrosome pair and associated pronuclei that precedes spindle assembly. 

Sawin, K.E., and Mitchison, T.J. (1991) Mitotic spindle assembly by two different pathways in vitro. J. Cell Biol. 112, 925. 

C. During operational qualification the analyst or vendor would assess if the equipment works as specified, generating appropriately documented data. The procedures will verify that the instrument s individual operational units are functioning within a given range or tolerance, reproducibly. For the dissolution apparatus, the water bath temperature and spindle assembly and shaft rpm speed would be obvious operational parameters. 

If the equipment passes the calibration for basket and paddles, then it can be assumed that the spindle assemblies, motor, and drive belt are functioning properly. The analyst may be able to test the wobble using equipment that assesses the run out measurement for the basket. 

Major components of dissolution equipment are the tester (including typically, but not limited to, spindle assemblies, belt, motor, tension adjuster, and circulator pump and hoses), water bath, paddles, baskets and shafts, vessels, samplers, and analyzers. Mechanical aspects, such as media temperature, paddle or basket speed, shaft centering and wobble, and vibration can all have a significant impact on the dissolution of the product. Mechanical and chemical calibration should therefore be conducted periodically, usually every 6 months, to ensure that the equipment is working properly. 

Vibration interference is a common problem with dissolution equipment (23). Careful leveling of the top plate and lids is critical. Within the spindle assembly, the bearings can become worn and cause vibration and wobble of the shaft. The drive belts should be checked for wear and dirt. The tension adjustments for the belt should be optimized for smooth operation. Surging of spindles, though difficult to detect without closely scrutinizing the tester operation, can cause spurious results. Vessels need to be locked in place so that they are not moving with the flow of water in the bath. 

Burlde A (2005) Poly(ADP-ribose). The most elaborate metabolite of NAD. Febs J 272 4576-4589 Burlde A, Brabeck C, Diefenbach J, Beneke S (2005) The emerging role of poly(ADP-ribose) polymerase-1 in longevity. Int J Biochem Cell Biol 37 1043—1053 Cardenas-Corona M, Jacobson E, Jacobson M (1987) Endogenous polymers of ADP-ribose are associated with the nuclear matrix. J Biol Chem 262 14863—14866 Chang P, Jacobson MK, Mitchison TJ (2004) Poly(ADP-ribose) is required for spindle assembly and structure. Nature 432 645-649... 

In higher eukaryotes, at the onset of S phase cyclin A accumulates which stimulates DNA synthesis. The amount of cyclin A continues to be high after the S phase because of its role in chromosome condensation. Cyclin A is degraded when cells enter prometaphase. The level of another cyclin called cyclin B rises during G2 phase, which helps to complete the chromosome condensation and spindle assembly, which allow transition to metaphase. Cyclin B is degraded by APC during metaphase. ... 

Metaphase Cyclin B Chromosome condensation and spindle assembly 349... 

Exit from mitosis Plk1 Spindle assembly 354... 

The cell cycle contains built-in control mechanisms that register defects in the course of the cell cycle and bring about a halt in the cell cycle to enable the fault to be repaired or to lead the cell to programmed cell death. These control mechanisms are also known as checkpoints. These are biochemical pathways that are activated when a fault occurs and can influence other critical steps of the cell cycle. Of particular importance for the cell cycle is the DNA damage checkpoint. Another important checkpoint is the spindle assembly checkpoint, which is not well biochemically characterized, however. 

Destruction of cohesin allows the spindle microtubules to pull the separated chromatids to opposite poles of the cell. Failure of spindle attachment to a single kinetochore activates the SAC (spindle assembly checkpoint), which arrests cells at metaphase until corrections are effected and equal distribution of chromosomes has been ensured. A sensory mechanism initiates the wait anaphase signal from an imattached kinetochore and triggers the accu mulation of the checkpoint components that comprise the Bub (budding uninhibited by benomyl)-Mad (mitotic arrest deficient) families of proteins. 

The spindle assembly checkpoint induced by paclitaxel is partially impaired when BRCAl expression is repressed in MCF-7 cells. BRCAl up-regulates BubRl transcription, and BubRl transcription and expression are significantly down-regulated in MCF-7... 

Mondal G, Baral RN, Roychoudhury S. A new Mad2-interacting domain of Cdc20 is critical for the function of Mad2-Cdc20 complex in the spindle assembly checkpoint. BiochemJ2006-,396 243-253. 

Weitzel DH, Vandre DD. Differential spindle assembly checkpoint response in human lung adenocarcinoma cells. Cell Tissue Res 2000 300 57-65. 

Anand S, Penrhyn-Lowe S, Venkitaraman AR. AURORA-A amplification overrides the mitotic spindle assembly checkpoint, inducing resistance to Taxol. Cancer Cell 2003 3 51-62. 

Sudo T, Nitta M, Saya H et al. Dependence of paclitaxel sensitivity on a functional spindle assembly checkpoint. Cancer Res 2004 64 2502-2508. 

Cyclic uptake and release of Ca2+ from the extracellular medium occur during mitosis in Physarum pofycephalum, and correlate with specific structural and kinetic events in the mitotic nuclei.442 The membrane system in the mitotic apparatus in Haemantkus endosperm cells functions in the localized release of Ca2+, so regulating the events of mitosis.443 It is known that calcium exerts effects on the stability of spindle microtubules. An alternative view is that free magnesium concentration acts as the fundamental regulator of the cell cycle.444 Tubulin polymerization depends on the presence of magnesium and the absence of calcium, and control of the Ca2+/Mg2+ ratio is relevant to spindle assembly. 

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