Abstract
BackgroundAnastral spindles assemble by a mechanism that involves microtubule nucleation and growth from chromatin. It is still uncertain whether γ-tubulin, a microtubule nucleator essential for mitotic spindle assembly and maintenance, plays a role. Not only is the requirement for γ-tubulin to form anastral Drosophila oocyte meiosis I spindles controversial, but its presence in oocyte meiosis I spindles has not been demonstrated and is uncertain.ResultsWe show, for the first time, using a bright GFP fusion protein and live imaging, that the Drosophila maternally-expressed γTub37C is present at low levels in oocyte meiosis I spindles. Despite this, we find that formation of bipolar meiosis I spindles does not require functional γTub37C, extending previous findings by others. Fluorescence photobleaching assays show rapid recovery of γTub37C in the meiosis I spindle, similar to the cytoplasm, indicating weak binding by γTub37C to spindles, and fits of a new, potentially more accurate model for fluorescence recovery yield kinetic parameters consistent with transient, diffusional binding.ConclusionsThe FRAP results, together with its mutant effects late in meiosis I, indicate that γTub37C may perform a role subsequent to metaphase I, rather than nucleating microtubules for meiosis I spindle formation. Weak binding to the meiosis I spindle could stabilize pre-existing microtubules or position γ-tubulin for function during meiosis II spindle assembly, which follows rapidly upon oocyte activation and completion of the meiosis I division.
Highlights
Anastral spindles assemble by a mechanism that involves microtubule nucleation and growth from chromatin
Chromatin has been shown to play an essential role [1], the involvement of the microtubule nucleator, g-tubulin, is still an open question. g-Tubulin localizes to centrosomes and other microtubule organizing centers in mitosis and is essential for nucleating microtubules in organisms as diverse as yeast, Drosophila, Xenopus, humans, and higher plants [2,3,4,5]. g-Tubulin is found on spindle microtubules, where it has been proposed to nucleate microtubules for spindle maintenance by functioning in a chromatinmediated nucleation pathway that augments the dominant pathway of nucleation by centrosomes [6,7]
Results gTub37C localizes to oocyte meiosis I (MI) spindles Females expressing gTub37C fused to a bright green fluorescent protein (GFP), regulated by the kinesin-14 ncd oocyte- and early embryo-specific promoter [21] were analyzed for this study
Summary
Anastral spindles assemble by a mechanism that involves microtubule nucleation and growth from chromatin. It is still uncertain whether g-tubulin, a microtubule nucleator essential for mitotic spindle assembly and maintenance, plays a role. Chromatin has been shown to play an essential role [1], the involvement of the microtubule nucleator, g-tubulin, is still an open question. G-tubulin is believed to play a central role in mitotic spindle assembly and maintenance in many organisms, its role in anastral spindles that lack centrosomes is less certain. Analysis of mutants that affect the Drosophila oocyte- and early embryo-specific gTub37C [16] has led to the conclusion that g-tubulin plays an essential role in nucleating microtubules for anastral oocyte meiosis I (MI) spindle assembly [17]. Attempts by several groups to stain oocyte MI spindles using anti-gTub37C antibodies have produced negative results [17,19,20], raising doubts as to whether g-tubulin is even present in the spindle
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
