Gametes are the vessels of gene immortality. To escape the dying soma, these cells rely on a specialized form of the cell cycle in which a single round of DNA replication is followed by two successive rounds of chromosome segregation known as meiosis I and II. In the process of halving the somatic DNA content, however, meiotic cycles run the risk of additional rounds of chromosome mis-segregation following meiosis II. How haploid cells avoid this potential threat of “meiosis III” by exiting the cycle after meiosis II still remains poorly understood, possibly because the responsible machinery has additional essential functions.
The irreversibility of many cell cycle steps is mediated by the ubiquitin/proteasome system (UPS), most famously by inactivation of the master cell cycle kinase activity through targeted proteolysis of its cyclin subunits. We explored in collaboration with Soni Lacefield’s lab (University of Indiana) if autophagy – the other major protein degradation system used to target various damaged organelles and protein aggregates to the lysosome for destruction – plays a role during meiotic divisions using the budding yeast S. cerevisiae as our model. Yeast autophagy mutants are known to abort initiation of sporulation – the process of gametogenesis in this organism – in response to starvation due to defects in nutrient signaling pathways. To get around this technical obstacle, we allowed cells to undergo pre-meiotic DNA replication before using a chemical inhibitor to inactivate the kinase activity of Atg1, a master regulator of autophagy, or by engineering the UPS to degrade Atg14, another essential autophagy factor. These two approaches acutely inhibited autophagy at the onset of the first meiotic division and revealed a mutant phenotype that had not been previously reported: meiosis I and II, which we monitored by video microscopy of cells with marked chromosomes and spindles, occurred with largely normal timing but were then followed by additional rounds of chromosome (mis)segregation. This aberration from normal meiotic exit yielded gametes with sub-haploid DNA content that were inviable.
In search of a molecular mechanism that could account for this striking phenotype, we found evidence that autophagy facilitates degradation of Rim4, a meiosis-specific regulator of mRNA translation. Pioneering work from Angelika Amon’s lab (MIT) had shown that Rim4 assumes an amyloid-like but non-toxic state during meiosis in which it binds to its mRNA targets, including a specific cyclin, to repress their translation. The expression of these mRNAs is normally induced by Rim4 degradation during meiosis I, a process that was blocked in the absence of autophagy. When we inhibited autophagy and at the same time artificially expressed two key Rim4 mRNA targets, we could restore normal termination of meiosis following meiosis II. In summary, our working model is that autophagic destruction of Rim4 ensures proper timing of a cell developmental gene expression program necessary for meiotic exit. This conclusion has potential relevance to human reproductive health because DAZL is a mouse amyloid-like factor that plays a key role in male gametogenesis and is known to regulate translation. Thus, it will be important to test if DAZL and human DAZL-like proteins (absent in mice), which reside in a Y chromosome area deleted in patients with certain forms of male infertility, are also degraded by autophagy.
by Fei Wang and Vlad Denic
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