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Updated: Jul 10, 2025
Meiotic Spindle Assessment in Mouse Oocytes by siRNA-mediated Silencing
Published on: October 11, 2015
Nuclear-cytoplasmic asynchrony in oocyte maturation caused by TUBB8 variants via impairing microtubule function: a
Tianli Chang1, Jing Zhao1, Qi Li1
1Reproductive Medicine Center, Xiangya Hospital, Central South University, Changsha, 410078, Hunan, China.
Background:
TUBB8, a crucial gene encoding microtubule protein, plays a pivotal role in cellular processes. Deleterious TUBB8 variants have been shown to significantly hinder oocyte maturation. In this study, we conducted an in vitro investigation using TUBB8 mutant mouse oocytes to elucidate the pathogenic mechanisms of TUBB8 variants in oocyte nuclear and cytoplasmic maturation.
Methods:
A mutant model was successfully established in mouse oocytes via microinjection to further investigate the effects of four novel discovered TUBB8 mutations on the nuclear and cytoplasmic maturation of mouse oocytes. Immunofluorescence and confocal microscopy were performed to observe the cortical polarity and spindle and of mutant oocytes. Active mitochondrial staining was performed to analyze mitochondrial distribution patterns. Endoplasmic reticulum and Ca2+ staining were conducted to assess ER distribution and cytoplasmic calcium ion concentration in oocytes.
Results:
In mouse oocytes, TUBB8 variants (p.A313V, p.C239W, p.R251Q, and p.G96R) resulted in a reduction of the first polar body extrusion rate, disruption of spindle assembly, and abnormal chromosome distribution. Additionally, these variants induced oocyte organelle abnormalities, including anomalies in mitochondrial redistribution and endoplasmic reticulum stress compared to the wild-type.
Conclusion:
Deleterious TUBB8 variants could disrupt microtubule function, affecting critical processes such as spindle assembly, chromosome distribution, and organelle rearrangement during oocyte meiosis. These disruptions culminate in compromised nuclear-cytoplasmic maturation, consequently giving rise to oocyte maturation defects.
Insights
Deleterious TUBB8 variants disrupt microtubule function, leading to oocyte maturation defects by compromising spindle assembly and organelle function. This study clarifies the pathogenic mechanisms of TUBB8 mutations in female meiosis.
Area of Science:
- Reproductive Biology
- Cell Biology
- Genetics
Background:
- The TUBB8 gene encodes a crucial microtubule protein essential for cellular processes.
- Deleterious TUBB8 variants are linked to impaired oocyte maturation.
- Understanding TUBB8 variant pathogenicity is vital for reproductive health.
Purpose of the Study:
- To investigate the pathogenic mechanisms of TUBB8 variants in mouse oocyte nuclear and cytoplasmic maturation.
- To elucidate how TUBB8 mutations affect key meiotic events in vitro.
- To analyze the impact of novel TUBB8 mutations on oocyte development.
Main Methods:
- Established a TUBB8 mutant mouse oocyte model via microinjection.
- Utilized immunofluorescence and confocal microscopy to assess spindle and cortical polarity.
- Performed active mitochondrial and endoplasmic reticulum (ER) staining to evaluate organelle function and distribution.
- Assessed Ca2+ ion concentration in oocytes.
Main Results:
- TUBB8 variants (p.A313V, p.C239W, p.R251Q, p.G96R) reduced first polar body extrusion rates.
- Observed disrupted spindle assembly and abnormal chromosome distribution in mutant oocytes.
- Identified oocyte organelle abnormalities, including mitochondrial redistribution anomalies and ER stress.
Conclusions:
- Deleterious TUBB8 variants impair microtubule function during oocyte meiosis.
- Disruptions in spindle assembly, chromosome distribution, and organelle rearrangement lead to maturation defects.
- TUBB8 mutations compromise oocyte nuclear-cytoplasmic maturation, impacting fertility.
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