Difference between revisions of "Fast Microtubule Dynamics in Meiotic Spindles Measured by Single Molecule Imaging: Evidence that the Spindle Environment does not Stabilize Microtubules"

From Soft-Matter
Jump to: navigation, search
Line 9: Line 9:
 
'''Publication''': Needleman et al. Fast microtubule dynamics in meiotic spindles measured by single molecule imaging: Evidence that the spindle environment does not stabilize microtubules. Molecular Biology of the Cell (2010) vol. 21 (2) pp. 323
 
'''Publication''': Needleman et al. Fast microtubule dynamics in meiotic spindles measured by single molecule imaging: Evidence that the spindle environment does not stabilize microtubules. Molecular Biology of the Cell (2010) vol. 21 (2) pp. 323
 
==Summary==
 
==Summary==
 +
The meiotic spindle is a piece of cellular machinery composed of microtubules (which are in turn composed of tubulin subunits) that serves to separate chromosomes during cell division. The spindle is incredibly dense with tubulin, containing a concentration much higher than found in the cytoplasm (~60 uM vs <1uM). Different theories have been proposed as to what leads to this high tubulin concentration gradient. One such theory focuses on the spindle somehow being a stabilizing environment that prevents nonkinetochore microtubules that had been nucleated elsewhere from depolymerizing. The results of this paper cast a serious doubt on this theory.
 +
 +
 +
The experiments were carried out in ''Xenopus laevis'' egg extracts where a small amount of fluorescently labeled tubulin was added. This tubulin incorporates into the microtubules as shown in Fig 1(C). The location of each labeled tubulin could be found accurately from these images by fitting a 2D gaussian function to the intensity from each point, as shown in Fig 1(A). Individual tubulin are added and removed from the microtubules in the spindle. By tracking each of these "dots" in the image from the time it appears to the time that it disappears, one can define the lifetime of each tubulin in the spindle. A plot of the relative frequency of each lifetime vs. the lifetime is found in Fig (2). The red fit line is the equation...
 +
 +
 +
<math>~~~P(v) ~ \alpha  ~ t^{-3/2}e^{-t/ \tau}</math>
 +
 +
 +
where tau is 4 times the expected lifetime of a microtubule of average length. This is the equation for a biased random walk. Here, tau is an adjustable parameter and the fit in Fig 2 has tau = 72 +-3 s. Even though this is a spectacular fit to the data, the authors stress that the use of a biased random walk in this context is simply a phenomenological model that serves as an approximation to  the "true" microscopic model of tubulin dynamics.
 +
 +
To check and see if there was a variation in local stabilization in different parts of the spindle each image was divided into 5 regions (2 pole regions, 2 midbody regions, and 1 center region) as shown in Fig 3 (A). The ration of deaths (tubulin leaving the spindle) to births (tubulin entering the spindle) was calculated and is shown for each region in figure 3 (B). It is evident that the ratio is similar in all parts of the spindle. Fig 3 (C) shows a similar plot to the one in Fig 2, but using the data collected from each region of the spindle. The overlap of points leads further support to the notion that no region of the spindle seems to be more stable than another.
 +
  
 
[[Image:Dn90fig2.png|thumb|300px| Figure 2, taken from [1].]]
 
[[Image:Dn90fig2.png|thumb|300px| Figure 2, taken from [1].]]

Revision as of 03:28, 20 October 2011

In progress, Fall 2011....

Original Entry: Peter Foster, AP 225, Fall 2011

Figure 1, taken from [1].

General Information

Authors: Daniel J. Needleman, Aaron Groen, Ryoma Ohi, Tom Maresca,Leonid Mirny, and Tim Mitchison

Publication: Needleman et al. Fast microtubule dynamics in meiotic spindles measured by single molecule imaging: Evidence that the spindle environment does not stabilize microtubules. Molecular Biology of the Cell (2010) vol. 21 (2) pp. 323

Summary

The meiotic spindle is a piece of cellular machinery composed of microtubules (which are in turn composed of tubulin subunits) that serves to separate chromosomes during cell division. The spindle is incredibly dense with tubulin, containing a concentration much higher than found in the cytoplasm (~60 uM vs <1uM). Different theories have been proposed as to what leads to this high tubulin concentration gradient. One such theory focuses on the spindle somehow being a stabilizing environment that prevents nonkinetochore microtubules that had been nucleated elsewhere from depolymerizing. The results of this paper cast a serious doubt on this theory.


The experiments were carried out in Xenopus laevis egg extracts where a small amount of fluorescently labeled tubulin was added. This tubulin incorporates into the microtubules as shown in Fig 1(C). The location of each labeled tubulin could be found accurately from these images by fitting a 2D gaussian function to the intensity from each point, as shown in Fig 1(A). Individual tubulin are added and removed from the microtubules in the spindle. By tracking each of these "dots" in the image from the time it appears to the time that it disappears, one can define the lifetime of each tubulin in the spindle. A plot of the relative frequency of each lifetime vs. the lifetime is found in Fig (2). The red fit line is the equation...


<math>~~~P(v) ~ \alpha ~ t^{-3/2}e^{-t/ \tau}</math>


where tau is 4 times the expected lifetime of a microtubule of average length. This is the equation for a biased random walk. Here, tau is an adjustable parameter and the fit in Fig 2 has tau = 72 +-3 s. Even though this is a spectacular fit to the data, the authors stress that the use of a biased random walk in this context is simply a phenomenological model that serves as an approximation to the "true" microscopic model of tubulin dynamics.

To check and see if there was a variation in local stabilization in different parts of the spindle each image was divided into 5 regions (2 pole regions, 2 midbody regions, and 1 center region) as shown in Fig 3 (A). The ration of deaths (tubulin leaving the spindle) to births (tubulin entering the spindle) was calculated and is shown for each region in figure 3 (B). It is evident that the ratio is similar in all parts of the spindle. Fig 3 (C) shows a similar plot to the one in Fig 2, but using the data collected from each region of the spindle. The overlap of points leads further support to the notion that no region of the spindle seems to be more stable than another.


Figure 2, taken from [1].
Figure 3, taken from [1].

Discussion

References

[1] Needleman et al. Fast microtubule dynamics in meiotic spindles measured by single molecule imaging: Evidence that the spindle environment does not stabilize microtubules. Molecular Biology of the Cell (2010) vol. 21 (2) pp. 323