Molecular Genetics of Arabidopsis Titan Mutants

拟南芥泰坦突变体的分子遗传学

• 批准号: 0091754
• 负责人: David Meinke
• 金额: $ 33万
• 依托单位: Oklahoma State University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-03-01 至 2005-02-28
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0091754&HistoricalAwards=false
• 关键词: Molecular; Genetics; Arabidopsis; Titan; Mutants

0091754MeinkeThe titan (ttn) mutants of Arabidopsis exhibit dramatic alterations inmitosis and cell cycle control during seed development. The commonfeature among these mutants is that endosperm nuclei become enlarged andhighly polyploid. Mutant embryos differ in cell size, morphology, andviability, depending on the locus involved. Three titan mutants withdistinct phenotypes were first identified in the Meinke laboratory 4 yearsago. During the previous grant period, 10 additional titan mutants wererecovered and 5 TITAN genes cloned in collaboration with Novartis(Syngenta). Several of these genes encode condensin (SMC) proteinsrequired for chromosome condensation and dosage compensation. Anothergene product is related to the ARF class of GTP binding proteinsassociated with vesicle trafficking. The remaining genes are involvedwith other cellular processes. What remains to be determined is how thesediverse proteins interact to influence nuclear division and cell viabilityduring seed development. This project will make it possible to continue the analysis of titanmutants and determine the relationship between gene function, nucleardynamics, and cell growth during seed development. Emphasis will beplaced on the SMC class of titans because these proteins appear to be mostdirectly linked to the mutant phenotype. Project objectives are toidentify the remaining SMC gene knockouts through reverse genetics,characterize nuclear and cytoskeletal defects in more detail throughfluorescence and electron microscopy, analyze phenotypes of multiple SMCknockouts, and determine how SMC functions in plants compare with thoseestablished in other organisms. The ARF class of titans will be examinedfirst by attempting to complement ttn1 with an ARF-GAP that maps to theTTN1 region. This should complete a map-based cloning effort initiatedduring the last grant period. The other class of titans will be examinedin part by analyzing more tagged endosperm mutants from the Novartiscollection, including those that have intermediate titan phenotypes.These additional mutants will be compared with existing titans toestablish the complex network of genes that influence endospermdevelopment. The titan phenotype demonstrates that Arabidopsis endosperm tissueprovides an ideal environment for proliferation of aberrant nucleidefective in mitosis and cell cycle control. Detailed analysis of thesemutants should provide valuable insights into novel features of nucleardynamics during endosperm development and evolution. The long-term goalis to understand the complex mechanisms that regulate endosperm formationin angiosperms.

0091754MeinkeTITAN(Ttn)突变体在种子发育过程中表现出极大的有丝分裂和细胞周期控制变化。这些突变体的共同特征是胚乳细胞核变大，高度多倍性。突变胚胎在细胞大小、形态和存活率上有所不同，这取决于所涉及的基因座。4年前，在Meinke实验室首次发现了三个具有不同表型的泰坦突变体。在之前的赠款期间，与诺华公司(先正达)合作，又回收了10个泰坦突变体，克隆了5个泰坦基因。其中几个基因编码凝集素(SMC)蛋白，是染色体凝聚和剂量补偿所必需的。另一个基因产物与ARF类GTP结合蛋白有关，与囊泡运输有关。剩下的基因与其他细胞过程有关。有待确定的是，在种子发育过程中，这些不同的蛋白质如何相互作用，影响核分裂和细胞存活率。该项目将使继续分析钛突变体并确定种子发育过程中基因功能、核动力学和细胞生长之间的关系成为可能。重点将放在SMC类巨人身上，因为这些蛋白质似乎与突变表型最直接相关。项目目标是通过反向遗传学识别剩余的SMC基因敲除，通过荧光和电子显微镜更详细地表征核和细胞骨架缺陷，分析多个SMC敲除的表型，并确定SMC在植物中的功能与在其他生物体中建立的SMC功能的比较。首先将通过尝试用映射到TTN1区域的ARF-GAP来补充ttn1来检查ARF类巨人。这应该会完成上一次授权期间启动的基于地图的克隆工作。另一类泰坦将通过分析诺华收集的更多有标签的胚乳突变体，包括那些具有中间泰坦表型的突变，来部分地进行检查。这些额外的突变体将与现有的泰坦进行比较，以建立影响胚乳发育的复杂基因网络。TITAN表型表明，拟南芥胚乳组织为异常核的增殖提供了理想的环境，这对有丝分裂和细胞周期控制有影响。对这些突变体的详细分析将为了解胚乳发育和进化过程中核动力学的新特征提供有价值的见解。长期的目标是了解调节被子植物胚乳形成的复杂机制。