PGRP 2016
Discovery and Evaluation of Inbred-Specific and Hybrid-Specific Regulatory Modules
“Discovery and Evaluation of Inbred-Specific and Hybrid-Specific Regulatory Modules”, is supported by National Science Foundation grant IOS 1546899 awarded to Briggs and Springer.
Steven P. Briggs (PI), UC San Diego, sbriggs@ucsd.edu
Nathan M. Springer (co-PI), University of Minnesota, springer@umn.edu
Background
We produced genome-wide, paired transcriptome::proteome quantitative datasets of hybrids and their parents, including three or more replicates per sample type. One part of our study covered 23 tissues and stages of development. Another part compared leaf tissue for multiple inbreds and their hybrids. A major finding was that maize hybrids selectively over-express the protein complexes required for the light reactions of photosynthesis (PS). A second major finding was that inbreds express the unfolded protein response (UPR). We produced protein cysteine redox status data for selected comparisons between inbreds and their hybrid because both PS and the UPR are regulated by protein redox changes. We modified the sample preparation method for redox analysis to incorporate click chemistry which enabled 25-fold enrichment of cysteine-containing peptides and a much deeper profile of the redoxome.
At the outset of our prior NSF support, the field of heterosis lacked a clear description of the key differences in physiology that distinguish hybrids from their less vigorous inbred parents. The field also lacked an evidence-based hypothesis for the mechanism of heterosis, which gives rise to the robust physiology of hybrids. Our plan was to compare transcriptional and proteomic networks between inbreds and hybrids with the hope that we would observe allele-specific differences that are important in heterosis. The role of such differences was to be determined using near-isogenic lines that enable allele exchanges between inbreds
Our most recent article provides a compelling explanation for some of the differences in physiology between hybrids and inbreds (Birdseye, D. et al. PNAS 2021). There have been many interesting observations of molecular differences between inbreds and hybrids. The magnitude of gene expression differences has been assumed to reflect the importance of a gene to heterosis. The relevance of such molecular differences has been unclear because only one or two hybrids have been examined at a time. We presented a new and robust method to ascertain the relevance of gene expression to heterosis. We determined correlations between expression and trait values across a panel of 15 hybrids and their parents instead of using the magnitude of differences. We found high Pearson correlation values between gene expression levels in seedlings and trait heterosis levels in adult plants, for coherent sets of genes. We showed that the current method, of ranking differences based on their magnitudes, can be misleading. Proteomics data were more revealing than transcriptomics data because many of the correlated genes are encoded by the chloroplast genome and thus their expression levels were not included in the mRNA data, and because protein expression levels were generally better correlated with trait heterosis levels.
Based on our results, we can for the first time use seedling expression levels to predict adult plant heterosis levels. We made the unexpected finding that chloroplast ribosome levels in seedling leaves are robust predictors of trait heterosis in adult plants. This finding has both scientific and practical importance. These are unexpected and exciting results that bring organelle protein synthesis forward as a central player in heterosis physiology. We observed that expression levels of ethylene (ET) biosynthesis enzymes, including ACS, were repressed in hybrids. The hybrid-specific differences in photosynthesis and ribosome protein expression levels were recreated in an inbred that contained mutated ACS genes. Therefore, repression of ET biosynthetic enzyme levels is upstream of the heterosis molecular phenotypes. Repression of ET levels in Arabidopsis hybrids has been previously reported. Thus, our findings indicate that heterosis physiology may be at least partially conserved between monocots and dicots.
Expression heterosis and correlation values for genes of interest may be visualized on our web application: https://devonbirdseye.shinyapps.io/ExpressionViewer/
Technology advances
We modified our sample preparation method to enable 25-fold enrichment of cysteine-containing peptides and a much deeper profile of the redoxome. This will facilitate identification of proteins that are less oxidized in inbreds (i.e., that have fewer disulfide bridges) and, thus, are more likely to be unfolded triggers of the UPR. We were motivated to make these improvements because, using conventional methods, we observed that the overall population of protein cysteines was more oxidized in hybrid than inbred leaf tissue, as would be expected if the extent of protein folding/disulfide bond formation is greater in hybrids.
Data resources
Several large scale RNA-seq datasets were created by this project. These include an atlas of transcript levels for inbreds and hybrids at 23 developmental stages (PRJNA482146) and transcriptome data for collections of inbreds/hybrids or recombinant inbreds and hybrids (PRJNA747924).
Companion peptide mass spectrometry datasets were created by this project.
Leaf tissue mass spec raw data:
http://massive.ucsd.edu/ProteoSAFe/status.jsp?task=ab8ae1ab9fce4ef59f4b81b708c6a79f
To view the dataset’s files, log in to the MassIVE FTP server with this URL:
ftp://MSV000085916@massive.ucsd.edu
Non-green tissue mass spec raw data:
http://massive.ucsd.edu/ProteoSAFe/status.jsp?task=25d970675f314a60ac3200e73609b780
To view the dataset’s files, log in to the MassIVE FTP server with this URL:
ftp://MSV000088081@massive.ucsd.edu
Citations and links for articles
Zhou P, Hirsch CN, Briggs SP, Springer NM. Dynamic Patterns of Gene Expression Additivity and Regulatory Variation throughout Maize Development. Mol Plant. 2019 Mar 4;12(3):410-425. doi: 10.1016/j.molp.2018.12.015. Epub 2018 Dec 27. PMID: 30593858
Sartor RC, Noshay J, Springer NM, Briggs SP. Identification of the expressome by machine learning on omics data. Proc Natl Acad Sci U S A. 2019 Sep 3;116(36):18119-18125. doi: 10.1073/pnas.1813645116. Epub 2019 Aug 16. PMID: 31420517
Zhou P, Li Z, Magnusson E, Gomez Cano F, Crisp PA, Noshay JM, Grotewold E, Hirsch CN, Briggs SP, Springer NM. Meta Gene Regulatory Networks in Maize Highlight Functionally Relevant Regulatory Interactions. Plant Cell. 2020 May;32(5):1377-1396. doi: 10.1105/tpc.20.00080. Epub 2020 Mar 17. PMID: 32184350
Birdseye D, de Boer LA, Bai H, Zhou P, Shen Z, Schmelz EA, Springer NM, Briggs SP. Plant height heterosis is quantitatively associated with expression heterosis of the plastid ribosomal proteins. doi: