The genetic bases of secondary metabolite diversity in a hybrid species complex

描述（由申请人提供）：本研究旨在提高对遗传距离较远的生物之间的杂交如何促进新表型的进化以及环境压力如何塑造最终表型多样性的理解。具体目标是：1)确定植物杂交种复合体中特定类别萜类代谢物的结构和含量多样性，比较自然存在的和实验创造的杂交种，以及测量发育过程中和对胁迫刺激的反应的变化；2)确定特定生物合成酶的转录反应变化和功能结构变化如何影响这些天然和人工杂交物种复合物的代谢输出；3)确定萜类代谢产物结构变异的遗传基础。利用现有的HPLC-MS和NMR分析方法对亲本和杂交物种进行萜烯结构的生化分析。编码负责萜类生物合成早期阶段的酶的遗传位点将从现有文献或通过与已知酶的同源性来确定。这些特定酶的DNA序列和mRNA转录物积累的变化将通过标准测序方案和定量PCR来测量。控制结构变异的其他候选基因座将通过结构定位群体和与模式植物物种中萜类修饰位点的同源性来确定，并通过酵母和细菌系统中的异源表达来评估这些候选酶的活性。来自不同亲本的基因组贡献如何相互作用产生独特的表型是一个生物学问题，对于理解包括人类在内的有性生殖物种的遗传基础和进化生物学特征具有重要意义。植物系统对这类研究特别有用，因为种间杂交在遗传上是可行的，在伦理上是允许的。由于萜类代谢物的致敏性和潜在的治疗用途，以及这些研究在代谢工程中的潜在效用，有待探索的代谢表型还与公共卫生相关。

DESCRIPTION (provided by applicant): This research is proposed to improve understanding of how hybridization between genetically distant organisms contributes to the evolution of novel phenotypes and how environmental pressures shape the resultant phenotypic diversity. The specific aims are: 1) to determine structural and content diversity of specific classes of terpenoid metabolites in plant hybrid species complexes, with comparison of naturally- existing and experimentally-created hybrids as well as measurement of variation across development and in response to stress stimuli; 2) to determine how variation in transcriptional response and functional variation in structure of specific biosynthetic enzymes impacts the metabolic output of these natural and artificial hybrid species complexes; and 3) to identify the genetic bases of structural variation in terpenoid metabolites. Biochemical analyses of terpene structures will be conducted on parental and hybrid species using modifications of existing analytical protocols for HPLC-MS and NMR. Genetic loci encoding enzymes responsible for early stages of terpenoid biosynthesis will be identified from existing literature or via homology to known enzymes. Variation in DNA sequence and mRNA transcript accumulation for these specific enzymes will be measured via standard sequencing protocols and quantitative PCR. Additional candidate loci controlling structural variation will be identified using structured mapping populations and homology to terpenoid modification loci identified in model plant species, with activity of these candidate enzymes evaluated via heterologous expression in yeast and bacterial systems. How genomic contributions from diverse parents interact to create unique phenotypes is a biological question of basic importance to understanding the genetic bases and evolution of biological traits in sexually-reproducing species, including humans. Plant systems are particularly useful for such studies as interspecific crosses are both genetically feasible and ethically permissible. The metabolic phenotypes to be explored are additionally relevant to public health due to the allergenicity and potential therapeutic uses of terpenoid metabolites, and the potential utility of these studies in metabolic engineering.