Intellectual Merit:

At the outset of our 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 manuscript provides a compelling explanation for some of the differences in physiology associated with heterosis (Birdseye, D. et al. 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.

To achieve our goals, 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. We produced protein cysteine redox status data for selected comparisons between inbreds and their hybrid because photosynthesis can be regulated by protein redox changes.

Technology development:

We modified our sample preparation method to enable 25-fold enrichment of cysteine-containing peptides and a much deeper profile of the redoxome. 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.

Citations and links to project publications:

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 plastid ribosomal proteins. Proc Natl Acad Sci USA 2021 (in press); bioRxiv doi: https://doi.org/10.1101/2021.02.16.431485

Broader Impacts:

Two postdoctoral scholars were trained. One accepted a position in academia and the other in industry.

Three PhD students were trained or are in training.

One MS student was trained and accepted a position in industry.

Several undergraduates and PhD rotation students participated in this research.

Last Modified: 09/20/2021
Modified by: Steven P Briggs