Cell-type Specific Strategies for Genome Maintenance

断裂的染色体对细胞来说是非常危险的，因为一个断裂可以导致数百个基因同时丢失。因此，所有生物都能检测到哪怕是一条双链断裂（DSB），从而启动包括诱导修复、阻止细胞分裂、改变发育命运，以及在某些情况下的程序性细胞死亡（PCD）在内的反应。这些反应因生物体和细胞类型而异。尽管损伤反应的细胞类型特异性很重要，但在任何系统中都没有很好地表征。最近的进展使得同时测量每个基因转录水平的变化成为可能。植物对dsb表现出强烈的转录反应，涉及数千个基因的诱导或抑制。新技术还允许从特定细胞类型中纯化细胞核。结合“转录组学”和表型分析将增加对转录调控在dsb修复和耐受中的作用的理解。在植物内的一个小而特殊的细胞群体中，经过PCD对断裂的反应将与周围细胞的转录反应进行比较，尽管经历了相同的dna损伤处理，但这些细胞并没有死亡。这种方法将允许鉴定断裂诱导的程序性细胞死亡所需的基因，为植物诱导PCD的早期和后期步骤的过程机制和标记提供见解。一些转录因子也被染色体断裂诱导。利用这些调控基因的突变体缺陷，研究反应中的子程序及其生物学意义。最后，使这种反应在植物中具有选择性优势的染色体断裂的自然来源尚不清楚。最近的研究表明，种子储存过程中的干燥和在富含金属的酸性土壤中生长引起的损伤都会引起上述转录反应，这一假设将得到进一步的研究。更广泛的影响：该项目将有助于理解细胞周期调控和植物细胞程序性死亡的机制。植物基因组稳定性的新测定和植物中dsb潜在天然来源的探索将导致与农业相关的发现。本科生、一名研究生和一名博士后学者将通过这项资助被雇用和培训，并且将从非常多样化的人群中招募。

Broken chromosomes are exceptionally dangerous to the cell, as a single break can result in the simultaneous loss of hundreds of genes. For this reason, all organisms can detect even a single double-strand break (DSB), initiating a response that includes the induction of repair, the arrest of cell division, changes in developmental fate, and in some cases programmed cell death (PCD). These responses vary from organism to organism, and with cell type. The cell-type specificity of damage response has not been well characterized in any system, in spite of its importance. Recent advances make it feasible to simultaneously measure changes in the transcription levels of every gene. Plants exhibit a robust transcriptional response to DSBs, involving the induction or suppression of thousands of genes. New technologies also allow purification of nuclei from specific cell types. Combining "transcriptomic" and phenotypic analysis will increase understanding of the role of transcriptional regulation in the repair and tolerance of DSBs. The transcriptional response in a small and specialized population of cells within the plant which undergo PCD in response to breaks will be compared to that of surrounding cells that do not die in spite of experiencing the same DNA-damaging treatment. This approach will allow identification of genes required for break-induced programmed cell death, providing insights into the mechanism of the process and markers for early and late steps in the induction of PCD in plants. Several transcription factors are also induced by chromosomal breaks. Using mutants defective in these regulatory genes, the subroutines within the response and their biological significance will be investigated.Finally, the natural source of chromosomal breaks that makes this response selectively advantageous in plants is unknown. Recent work indicates that both the desiccation experienced during seed storage and damage induced by growth in acidic, metal-rich soils provoke the transcriptional response described above, and this hypothesis will be investigated further.Broader impacts: This project will contribute to the understanding of cell cycle regulation and the mechanisms of programmed cell death in plants. Novel assays for genomic stability in plants and exploration of potential natural sources of DSBs in plants should result in findings of relevance to agriculture. Undergraduates, a graduate student, and a postdoctoral scholar will be employed and trained through this grant, and will be recruited from a very diverse population.