Source: IOWA STATE UNIVERSITY submitted to
ADVANCING UNDERSTANDING OF HEAT STRESS RESPONSE MECHANISMS BY INTEGRATED MOLECULAR, BIOCHEMICAL, AND WHOLE-PLANT ANALYSIS
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
EXTENDED
Funding Source
Reporting Frequency
Annual
Accession No.
1008557
Grant No.
2016-67013-24602
Project No.
IOW05460
Proposal No.
2015-06894
Multistate No.
(N/A)
Program Code
A1101
Project Start Date
Jan 1, 2016
Project End Date
Dec 31, 2020
Grant Year
2016
Project Director
Yu, J.
Recipient Organization
IOWA STATE UNIVERSITY
2229 Lincoln Way
AMES,IA 50011
Performing Department
Agronomy
Non Technical Summary
High temperature stress severely limits plant productivity and causes extensive economic loss to US agriculture. The negative impact of heat stress on yield stability and agricultural production has been exacerbated in recent years, and what is predicted for the future from climate change models is even more alarming. Hence, understanding the mechanisms of crop response to heat stress is critical to sustain agricultural production. Little progress has been made in this area, however, primarily due to the challenge of having access to field-based heat stress conditions, the complexity of the crop genomes, and the inadequate integration of physiology and genetics research. Without dedicated effort in advancing research in heat stress response in crops, we are certainly ill-prepared for global warming and climate change, which have shown the impact of climatological extremes, including high temperature stress, on agriculture production.The overarching rationale of the proposed research is to fill a critical research gap in understanding the mechanisms of crop response to abiotic stress, specifically heat stress response in maize. The first objective is to quantify the natural variation in heat stress response across a diverse set of maize inbred lines and identify underlying mechanisms through integrating analysis at the gene-expression, biochemical, and whole-plant level. To achieve this goal, we plan to conduct genome-wide gene expression profiling through of a set of diverse maize accessions that have been identified from extensive field screening. The second objective is to identify the genes that contribute to the differential heat stress response through joint analysis of two populations. Two mapping populations will be developed through the doubled haploid process, and field-based phenotyping and subsequent genetic mapping will be conducted.This project is designed to obtain a much improved understanding of heat stress by integrating biochemical and whole-plant characterization with molecular mechanisms. Knowledge generated from this project is expected to facilitate further breeding research in not only maize, but also many other crops. The societal benefit of this project is its scientific contribution to the sustainable agriculture production under changing climates.
Animal Health Component
0%
Research Effort Categories
Basic
80%
Applied
20%
Developmental
(N/A)
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
2031510102070%
2011510108030%
Goals / Objectives
High temperature stress severely limits plant productivity and causes extensive economic loss to US agriculture. Little progress has been made in this area, however, primarily due to the challenge of having access to field-based heat stress conditions, the complexity of the crop genomes, and the inadequate integration of physiology and genetics research.Our project goal is to elucidate the genetic and physiological mechanisms of heat stress response in maize.Objective 1: Quantify the natural variation in heat stress response across a diverse set of maize inbred lines and identify underlying mechanisms through integrating analysis at the gene-expression, biochemical, and whole-plant level. A set of 24 diverse accessions have been identified from extensive field screening. We plan to conduct genome-wide transcriptional profiling through RNA-sequencing.Objective 2: Identify the genetic loci that contribute to the differential heat stress response through meta-analysis of twopopulations. Doubled haploid mapping populations from between a heat tolerant line (B76) and two sensitive lines (B106 and NC350) are being developed for field-based phenotyping and subsequent genetic mapping.These two complementary approaches facilitate the integration of biochemical and whole-plant characterization with molecular mechanisms. This project is expected to generate gene expression network that facilitate research in not only maize, but also many other crops. Additional maize lines that have either comparable or superior heat stress tolerance than the mapping parent (B76) will be available for futher mechanistic study and breeding research.
Project Methods
Objective 1. Quantify the natural variation in heat stress response across a diverse set of maize inbred lines and identify underlying mechanisms through integrating analysis at the gene-expression, biochemical, and whole-plant level. A set of 24 diverse inbreds have been identified from extensive field screening of 600 maize inbred lines and genetic diversity analysis with high density single nucleotide polymorphism (SNP) markers. These inbreds showed significant difference in heat stress response at early, mid, and late season: leaf firing, tassel blasting, plant death, leaf bleaching, tassel barren, and delay of silking. We plan to conduct genome-wide transcriptional profiling through RNA-sequencing (RNA-seq) by subjecting this set of 24 maize inbreds to controlled heat stress conditions that produce rankings similar to available field screening results in leaf firing, the most prevalent trait with high repeatability. Gene expression networks will be constructed and compared with established networks in model plant species. Together with transcriptional profiling under non-stress conditions, the most cutting-edge Differential Network (DN) analysis method will be used to identify genes critical to heat response mechanisms. Membrane lipid profiling will allow us to examine the roles of phosphatidic acid (PA) on heat stress response and to validate the potential correlation of PA with heat tolerance traits across diverse genetic backgrounds.Objective 2. Identify the genetic loci that contribute to the differential heat stress response of three representative maize inbred lines through quantitative trait locus (QTL) analysis of two populations. Two doubled haploid (DH) mapping populations from a heat-tolerant line (B76) and two heat-sensitive lines (B106 and NC350) will be developed for field-based phenotyping of heat stress response and subsequent genetic mapping. As an approach adopted by industry, DH significantly reduces time and cost of population development compared with repeated selfing to obtain recombinant inbred lines. Genotyping-by-sequencing (GBS), a widely adopted genotyping method in maize and many other crop species for its high throughput capacity, will be applied to obtain high density SNP markers for QTL analysis of these DH populations. Specific DHs with different combinations of QTL alleles will be selected for membrane lipid profiling and physiological characterization.

Progress 01/01/18 to 12/31/18

Outputs
Target Audience:Target audiences include research scientists in plant physiology, plant breeding, genetics and genomics, and agronomy, and growers. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?One gradaute student worked on this project traveled to Lubbock Texas and gained experinece in field evaluation of heat stress response of a mapping population. A second graduate student worked on this project gained experience in RNA extraction and reviewed the literature in RNA sequencing in crops. Several undergradaute students participated the field experiment to increase the seeds for two other mapping population and the greenhouse experirment for tisuss sampling of the mapping population for genotyping. How have the results been disseminated to communities of interest?We published the paper in Crop Science in 2018, representing a starting point to further elucidate the genetic control of heat stress responsein North American maize germplasm. To our knowledge, this is the first peer-reviewed paper of a genetic mapping study in field-based heat-tress responseusing North American maize germplasm. Given the anticipated interest, we published the paper in Open Access. This paper was selected to be highlighted in aCSA News article by Crop Science Soceity of America. What do you plan to do during the next reporting period to accomplish the goals?We will conduct field experiments with the threemapping populations, as well as the 28-line set. We will conduct RNA extraction and sequencing andcomplete data analysis for membrane thermostability measurements andlipid profiling.

Impacts
What was accomplished under these goals? IMPACT: We worked on elucidating the genetic and physiological mechanisms of heat stress response in maize. The project addresses the knowledge gap between model species and crops on heat stress response. With the initial experiments, we have collected extensive physiological data in-greenhouse to differentiate maize inbreds and developed genetic populations for further field characterization of heat stress response and genetic mapping. Objective 1: Quantify the natural variation in heat stress response across a diverse set of maize inbred lines and identify underlying mechanisms through integrating analysis at the gene-expression, biochemical, and whole-plant level. We completed two sets ofexperiments: lipid profiling and membrane thermostability analysis, and made proress in the third expeirments.We worked on comparining diffiernet RNA extraction protocols and examined the sequencing reads and costs betweent the estbalished RNA-seq and the new 3' RNA-seq. Objective 2: Identify the genetic loci that contribute to the differential heat stress response through meta-analysis of two populations. We conducted the field experiments at Lubbock Texas withthe first mapping population, B76/B106, a doubled haploid (DH) population with 224 individuals. We completed the development (seed increase) of two other mapping population at Ames Iowa, B76/NC350with 123 recombinant inbred lines (RILs), andNC350/B106with 144 RILs.

Publications

  • Type: Journal Articles Status: Published Year Published: 2018 Citation: McNellie, J.P., J. Chen, X. Li, and J. Yu. 2018. Genetic mapping of foliar and tassel heat stress tolerance in maize. Crop Science 58:2484-2493.


Progress 01/01/17 to 12/31/17

Outputs
Target Audience:Other scientists working on maize heat stress Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?On the physiology side, we have trained 3 undergraduate students for sample collection, lipid extraction, and membrane stability measurement. On the breeding sides, summer nursery to develop three mapping populations provided an opportunity to expose 3 undergraduate students to plant breeding and genetics research. How have the results been disseminated to communities of interest?One graduate student presented a poster at the 59th Annual Maize Genetics Conference. What do you plan to do during the next reporting period to accomplish the goals?We will conduct RNA extraction and sequencing, complete data analysis for membrane thermostability measurements, complete lipid profiling and data analysis, and conduct field experiments with the developed mapping populations.

Impacts
What was accomplished under these goals? We worked on elucidating the genetic and physiological mechanisms of heat stress response in maize. The project addresses the knowledge gap between model species and crops on heat stress response. With the initial experiments, we have collected extensive physiological data in greenhouse to differentiate maize inbreds and developed genetic populations for further field characterization of heat stress response and genetic mapping. Objective 1: Quantify the natural variation in heat stress response across a diverse set of maize inbred lines and identify underlying mechanisms through integrating analysis at the gene-expression, biochemical, and whole-plant level. We conducted three experiments: tissue sampling for RNA sequencing, lipid profiling, and membrane thermostability analysis. These experiments were conducted in a coordinated fashion so that data generated from three experiments can be analyzed together. A set of 28 inbreds were assembled to include the core set of 24 inbreds lines and 4 additional inbreds (Mo17, B73, B104, and CML277). These inbreds were grown under optimal condition in a greenhouse with an appropriate experimental design. The plants were divided into two groups: one for heat treatment and one as control. Leaf punches were collected for RNA extraction and sequencing. A separate set of leaf punches were collected for lipid extraction and lipid profiling is ongoing. A third set of leaf punches were collected for electrolyte leakage analysis, and preliminary results were obtained. Objective 2: Identify the genetic loci that contribute to the differential heat stress response through meta-analysis of two populations. We completed the development of a set of connected mapping populations. The first one, B76/B106, is a doubled haploid (DH) population with 224 individuals. The second one, B76/NC350, is a population with 123 recombinant inbred lines (RILs), The third population, NC350/B106, with 144 RILs.

Publications

  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: McNellie, J.P., X. Li, J. Chen, J. Yu. 2017. Advancing understanding of heat stress response mechanisms by integrated, molecular, biochemical, and whole-plant analysis. The 59th Annual Maize Genetics Conference, p201.


Progress 01/01/16 to 12/31/16

Outputs
Target Audience:We discussed the project and initial findings with other scientists working on maize heat stress. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?We have trained 3 undergraduate students for sample collection, lipid extraction, membrane stability measurement, and gas exchange and chlorophyll fluorescence measurement How have the results been disseminated to communities of interest?We discussed the project and initial findings with other scientists working on maize heat stress. What do you plan to do during the next reporting period to accomplish the goals?We will collect samples for lipidomics and RNA sequencing, perform membrane stability and chlorophyll content measurements, increase seeds of three mapping populations, and start analyzing the data collected.

Impacts
What was accomplished under these goals? Our project goal is to elucidate the genetic and physiological mechanisms of heat stress response in maize. The project addresses the knowledge gap between model species and crops on heat stress response. With the initial experiments, we have collected extensive physiological data in greenhouse to differentiate maize inbreds and developed genetic populations for further field characterization of heat stress response and genetic mapping. Objective 1: Quantify the natural variation in heat stress response across a diverse set of maize inbred lines and identify underlying mechanisms through integrating analysis at the gene-expression, biochemical, and whole-plant level. We conducted 3 greenhouse experiments and performed two types of heat stress measurements for the 27 inbreds under controlled environments. The original inbred list was expanded to include B73, Mo17 and B104. First, we phenotyped the heat stress responses of maize plants at the whole-plant level under controlled environment. Second, we conducted a subset of experiments to a) examine variation of heat stress treatment performed in greenhouse vs. in walk-in growth chambers, b) test how to effectively coordinate different physiological and biochemical measurements and sampling process during the heat treatment, and c) train students for sample collections, lipid extraction, membrane stability measurement, gas exchange and chlorophyll fluorescence measurements. Third, we conducted a time-course experiment to measure chlorophyll fluorescence and leaf-level net carbon assimilation of young and mature leaves. Objective 2: Identify the genetic loci that contribute to the differential heat stress response through meta-analysis of two populations. Two mapping population are being developed. The first one, B76/B106, is a doubled haploid (DH) population with 234 individuals. The second one, B76/NC350, is a recombinant inbred line population with 126 individuals. As a remedy to the relatively small size of the second population (due to the adaptation issue from NC350), we have also developed a third population, NC350/B106, with 150 individuals. Seed increase will be conducted in summer nursery at Ames Iowa in 2017.

Publications