Establishment of the meiotic cell cycle program through post-transcriptional regulation by MEIOC and YTHDC2

项目摘要/摘要 二倍体生殖细胞通过减数分裂转化为单倍体配子。减数分裂后 DNA合成，生殖细胞进入减数分裂前期I，这与有丝分裂前期有两个区别 主要特点。首先，减数分裂前期I显著长于有丝分裂前期。第二，减数分裂 第一阶段涉及一系列复杂的染色体事件，这些事件有助于建立 单倍体状态。前期I的延长长度对这些染色体事件的完成至关重要，因为 过早退出前期I会导致重组缺陷和染色体错误分离。在哺乳动物身上， Meioc和Ythdc2延长了减数分裂前期I。在男性和女性都缺乏这两种基因的情况下， 减数分裂生殖细胞未能完成减数分裂前期I，提前进入异常中期 最终的结果是。一些初步观察表明，MEIOC和YTHDC2作为一种 在减数分裂生殖细胞中转录后调节细胞周期因子的复合体。MEIOC和YTHDC2是 在减数分裂开始时首次在胚系中表达，它们彼此相互作用，以及与 重叠的转录本集合，包括关键的细胞周期调节因子。这些靶标被浓缩为RNA 修饰N6-甲基腺苷(M6A)，其优先被YTHDC2‘S YTH结构域识别和 它被用来靶向和转录后调节RNA。在Meioc缺席的情况下，MEIOC和 YTHDC2的S目标稳定性降低。转录稳定性的变化往往伴随着 在HeLa细胞中，YTHDC2在降低mRNA的同时提高翻译效率 富足。然而，目前尚不清楚MEIOC和YTHDC2是否影响减数分裂胚胎的翻译 细胞。这一提议将检验这样的假设，即MEIOC和YTHDC2作为一个复合体来识别和 翻译调节m6A修饰的mRNA，以建立减数分裂特异的细胞周期程序。这 假说将通过三个具体目标进行检验。第一个目的是评估经奥委员会影响YTHDC2 S的能力 通过鉴定YTHDC2与RNA的相互作用，特别是与细胞周期相关的转录本 在Meioc在场和不在场的情况下绑定到。此外，M6A是否对MEIOC和YTHDC2的S有贡献 还将评估与RNA相互作用的能力。第二个目标将确定以下翻译更改 当生殖细胞从有丝分裂过渡到减数分裂时，以及在野生型和Meioc空样本之间发生。在……里面 具体地说，这个目标将决定细胞周期因子是否在减数分裂开始时被翻译抑制。 以及Meioc的上调。第三个目标将决定MEIOC和YTHDC2是否连续 在减数分裂前期I所需的，以防止早熟细胞周期进展，以及识别 这一假定要求背后的机械性监管。总的来说，这些关于MEIOC的基础研究 而YTHDC2的S对减数分裂前期I长度的调控将为建立 以及对减数分裂细胞周期程序的调控，对生育、发育和癌症具有更广泛的影响。

PROJECT SUMMARY/ABSTRACT Diploid germ cells are transformed into haploid gametes via the reductive division of meiosis. After meiotic DNA synthesis, germ cells enter meiotic prophase I, which distinguishes itself from mitotic prophase by two major characteristics. First, meiotic prophase I is dramatically longer than mitotic prophase. Second, meiotic prophase I involves an intricate series of chromosomal events that contribute to the establishment of the haploid state. The extended length of prophase I is critical for the completion of these chromosomal events, as premature exit from prophase I causes recombination defects and chromosome mis-segregation. In mammals, meiotic prophase I is prolonged by Meioc and Ythdc2. In the absence of either gene in both males and females, meiotic germ cells fail to complete meiotic prophase I and prematurely enter an abnormal metaphase before ultimately apoptosing. Several preliminary observations suggest that MEIOC and YTHDC2 function as a complex to post-transcriptionally regulate cell cycle factors within meiotic germ cells. MEIOC and YTHDC2 are first expressed in the germ line at meiotic initiation, and they interact with one another as well as with an overlapping set of transcripts, including key cell cycle regulators. These targets are enriched for the RNA modification N6-methyladenosine (m6A), which is preferentially recognized by YTHDC2's YTH domain and which is used to target and post-transcriptionally regulate RNA. In the absence of Meioc, MEIOC and YTHDC2's targets exhibit reduced stability. Changes in transcript stability are oftentimes accompanied by changes in translation, and in HeLa cells, YTHDC2 enhances translational efficiency while decreasing mRNA abundance. However, it remains unknown whether MEIOC and YTHDC2 affect translation in meiotic germ cells. This proposal will test the hypothesis that MEIOC and YTHDC2 function as a complex to recognize and translationally regulate m6A-modified mRNA in order to establish a meiosis-specific cell cycle program. This hypothesis will be tested via three specific aims. The first aim will assess MEIOC's ability to affect YTHDC2's interactions with RNA, particularly cell cycle-related transcripts, by identifying the transcripts that YTHDC2 binds to in the presence and absence of Meioc. In addition, whether m6A contributes to MEIOC and YTHDC2's ability to interact with RNA will also be assessed. The second aim will identify the translational changes that occur as germ cells transition from mitosis to meiosis, as well as between wild-type and Meioc-null samples. In particular, this aim will determine whether cell cycle factors are translationally repressed at the onset of meiosis and by the upregulation of Meioc. The third aim will determine whether MEIOC and YTHDC2 are continuously required during meiotic prophase I to prevent precocious cell cycle progression, as well as identify the mechanistic regulation behind this putative requirement. Collectively, these fundamental studies on MEIOC and YTHDC2's regulation of the length of meiotic prophase I will provide key insights into the establishment and regulation of the meiotic cell cycle program, with broader implications for fertility, development, and cancer.