The role of VRK-1 during the meiotic divisions of C. elegans embryos

VRK-1是线虫体内的一种痘苗相关蛋白，在整个发育过程中具有多种功能。它是胚胎发生过程中核膜形成所必需的，也是幼虫发育过程中生殖系增殖和外阴形态发生所必需的。果蝇NHK-1是一种组蛋白激酶，其突变等位基因影响卵母细胞的染色体形态。我们一直对影响线虫减数分裂过程中染色体形态和行为的蛋白质感兴趣。我们已经开始使用RNAi检测VRK-1基因缺失的胚胎的表型。我们观察到了高度渗透性的胚胎致死性。这些胚胎在减数分裂过程中表现出缺陷，也不能使染色质解聚。因此，这些胚胎作为带有很少DNA的多细胞胚胎停止发育。这种缺陷不是发育中的卵母细胞核膜缺陷的结果，也不是生殖系发育的任何其他方面的结果。我们认为VRK-1蛋白正在影响卵母细胞减数分裂染色体的组蛋白修饰，从而扰乱了受精后减数分裂染色体正确组织的能力。我们正在使用表达GFP标记的组蛋白的活胚胎的实时成像来追踪VRK-1耗尽时发生的发育缺陷。我们还使用免疫细胞化学来确定此时是否有任何特定的组蛋白修饰被干扰，以解释观察到的减数分裂缺陷。 我们还在筛选VRK-1的突变等位基因，这将使我们能够研究它在整个发育过程中的许多功能。缺失等位基因确实存在，并导致不育。我们正在使用TILLING来筛选VRK-1基因点突变。有了VRK-1的有用等位基因，我们可能能够使用遗传抑制筛选来进一步剖析线虫中的VRK-1途径。

VRK-1, a vaccinia-related kinase in C. elegans, has multiple functions throughout development. It is required for nuclear envelope formation during embryogenesis and for germ line proliferation and vulval morphogenesis during larval development. The Drosophila ortholog, NHK-1, is a histone kinase, mutant alleles of which affect chromosome morphology in oocytes. We have an ongoing interest in proteins that influence chromosome morphology and behavior during the meiotic divisions in C. elegans. We have begun to examine the phenotypes of embryos depleted of the vrk-1 gene, using RNAi. We have observed highly penetrant embryonic lethality. These embryos display defects in the meiotic divisions as well as a failure to decondense chromatin. As a result, these embryos arrest as multicellular embryos with very little DNA. This defect is not a result of nuclear envelope defects in the developing oocytes nor in any other aspect of germ line development. We believe the VRK-1 protein is affecting the histone modifications of the meiotic chromosomes of the oocyte and thus is perturbing their ability to properly organize for the meiotic divisions that follow fertilization. We are using real time imaging of live embryos expressing GFP-tagged histones to follow the developmental defects that occur upon vrk-1 depletion. We are also using immunocytochemistry to determine whether any specific histone modifications are perturbed at this time to account for the observed meiotic defects. We are also in the process of screening for mutant alleles of vrk-1, which would allow us to study its many functions throughout development. A deletion allele does exist and results in sterility. We are using TILLING to screen for point mutations in the vrk-1 gene. With useful alleles of vrk-1, we may be able to further dissect the vrk-1 pathway in C. elegans using a genetic suppressor screen.