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Microtubule tracks can be detected in mouse oocytes with an antibody directed against a calcium transporter
H de Pennart1, C Cibert, C Petzelt
1Département de Biologie du Développement, Institut Jacques Monod, CNRS-Université Paris 7, France.
Abstract:
In metaphase II-arrested mouse oocytes, most microtubules are found in the meiotic spindle, a structure that remains stable for hours despite microtubule instability. Microtubule organizing centres (MTOCs) are present at the poles of the spindle and in the cytoplasm, but the latter nucleate very few microtubules. This particular organization of the microtubule network enabled us to observe the unexpected behaviour of a protein that can associate with microtubules. We compared the distribution of a mitosis-activated calcium transport system with that of the microtubule network, by immunofluorescence, using two monoclonal antibodies, one directed against a component of the calcium transport system (7/13), and the other against the common tyrosinated form of alpha-tubulin (YL1/2). The 7/13 staining was associated with the spindle microtubules and with the kinetochore area. In addition, we observed many asters in the cytoplasm, around the cytoplasmic MTOCs. The majority of these asters were not stained with the antitubulin antibody. Moreover, these 7/13 asters either disappeared after nocodazole treatment or were enlarged after taxol treatment. Using a confocal microscope, we observed single fibres that were stained with both antibodies: the extremity furthest from the MTOC (corresponding to the + end of the microtubule) being detected by the 7/13 antibody only. All these observations suggest that the 7/13 antigen is associated with microtubule tracks that persist a few minutes after microtubule depolymerization. The possible role of these tracks in microtubule regrowth is discussed.
Insights
Researchers identified a novel protein (7/13 antigen) associated with stable microtubule tracks in mouse oocytes. This protein persists after microtubule depolymerization, suggesting a role in microtubule regrowth.
Area of Science:
- Cell Biology
- Molecular Biology
- Reproductive Biology
Background:
- Mouse oocytes in metaphase II feature a stable meiotic spindle despite inherent microtubule instability.
- Cytoplasmic microtubule organizing centers (MTOCs) nucleate few microtubules, creating a unique microtubule organization.
Purpose of the Study:
- To investigate the distribution of a mitosis-activated calcium transport system in relation to the microtubule network in mouse oocytes.
- To characterize the behavior of the 7/13 antigen, a component of the calcium transport system, and its association with microtubules.
Main Methods:
- Immunofluorescence microscopy using monoclonal antibodies against the 7/13 antigen and tyrosinated alpha-tubulin (YL1/2).
- Confocal microscopy to visualize detailed microtubule-microtubule organizing center (MTOC) interactions.
- Treatment with nocodazole and taxol to assess microtubule dynamics and the 7/13 antigen's response.
Main Results:
- The 7/13 antigen localized to spindle microtubules and the kinetochore area.
- Cytoplasmic asters, stained by 7/13 but not antitubulin, appeared around MTOCs.
- These 7/13 asters were sensitive to nocodazole and taxol, indicating microtubule dependence.
- Confocal imaging revealed the 7/13 antibody specifically labeling the plus-end extremity of single microtubule fibers.
Conclusions:
- The 7/13 antigen is associated with persistent microtubule tracks that remain visible for minutes after depolymerization.
- These findings suggest a potential role for the 7/13 antigen in regulating microtubule regrowth processes.