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Kinetochore microtubules in crane-fly spermatocytes: untreated, 2 degrees C-treated, and 6 degrees C-grown spindles
Abstract:
We investigated the involvement of kinetochore microtubules (kMTs) in mediating chromosome-to-pole connections in crane-fly (Nephrotoma suturalis and Nephrotoma ferruginea) spermatocytes. Two experimental treatments were used to yield spindles with reduced numbers of nonkinetochore microtubules (nkMTs). Short-term (10-15 min) exposure of spermatocytes to 2 degrees C caused depolymerization of the majority of nkMTs, resulting in a kMT:(kMT + nkMT) ratio of 0.76. Long-term (24h) exposure to 2 degrees C followed by recovery at 6 degrees C resulted in a kMT:(kMT + nkMT) ratio of 0.55, the spindle having more nkMTs than a 2 degrees C-treated spindle but fewer than an untreated spindle, in which the kMT:(kMT + nkMT) ratio was 0.27. The numbers and lengths of kMTs in 6 degrees C-grown spindles were similar to those in untreated cells, suggesting that the overall inhibition of MT assembly at 6 degrees C apparently did not affect the mechanism by which kMTs are formed. We observed most kMTs of early anaphase spindles to be long (greater than 3 microns), and many extended to the polar regions of the spindle. Thus, the crane-fly spindle appears not to be as atypical as it was previously suggested to be.
Insights
Crane-fly spermatocytes utilize kinetochore microtubules (kMTs) for chromosome-to-pole connections. Experiments show kMTs are essential for spindle function, even with reduced nonkinetochore microtubules (nkMTs).
Area of Science:
- Cell Biology
- Cytoskeleton Dynamics
- Meiosis
Background:
- Kinetochore microtubules (kMTs) are crucial for chromosome segregation during cell division.
- The structure and function of crane-fly spindles have been debated, with suggestions of atypical organization.
Purpose of the Study:
- To investigate the role of kMTs in establishing chromosome-to-pole connections in crane-fly spermatocytes.
- To determine if crane-fly spindles are atypical in their microtubule composition.
Main Methods:
- Crane-fly spermatocytes were subjected to cold treatments (2°C for short and long durations) to selectively depolymerize nonkinetochore microtubules (nkMTs).
- Microtubule composition (kMTs vs. nkMTs) and kMT lengths were analyzed in treated and untreated spindles.
- The ratio of kMTs to total microtubules (kMTs + nkMTs) was calculated under different experimental conditions.
Main Results:
- Cold treatments significantly reduced the proportion of nkMTs, altering the kMT:(kMT + nkMT) ratio.
- Spindles with reduced nkMTs still maintained functional chromosome-to-pole connections mediated by kMTs.
- kMTs in treated spindles were predominantly long and extended towards the spindle poles, similar to untreated cells.
Conclusions:
- The crane-fly spindle is not as atypical as previously suggested, with kMTs playing a conserved role in chromosome attachment.
- The mechanism of kMT formation appears robust and unaffected by overall microtubule assembly inhibition.