Nature Biotechnology: Key genetic regulatory mechanisms for maize resistance and yield balance

Nanhu News Network News (correspondent Sun Xiaopeng) On October 13, the National Key Laboratory of Crop Genetic Improvement, Hubei Hongshan Laboratory, Professor Dai Mingqiu, Professor Li Lin's research group, Hubei Hongshan Laboratory, and the Key Laboratory of Horticultural Plant Biology of the Ministry of Education Professor Li Feng's research group in Nature — The journal Nature Biotechnology published a paper titled " The role of transposon invertedrepeats in balancing drought tolerance and yield-related traits in maize” Using multi-omics and gene editing strategies, the genetic and molecular mechanisms of maize yield and resistance balance were elaborated in detail, which laid a theoretical foundation for the precise molecular design and breeding of maize with high resistance and high yield, and provided excellent genetic resources
.

Maize is the largest and most productive food crop in China, which is crucial
to ensuring national food security.
Maize is a tall crop that needs water throughout its life and is sensitive to drought; The main corn planting areas in China are mostly located in arid and semi-arid areas, and the corn yield is greatly reduced and huge economic losses
are caused by drought every year.

Therefore, the cultivation of drought-resistant maize is of great practical significance
.
Crop breeding is often aimed at yield, and the genome that controls excellent production traits is fixed by artificial selection; Natural selection, on the other hand, aims at the reproduction of species and focuses on the selection and utilization
of environmentally adaptive genomes.
In the past hundred years, breeders have greatly increased maize yields, but this has been accompanied by a significant decline in maize drought resistance, indicating that yield traits and stress tolerance traits are often antagonistic to each other, and the genetic loci that control them are closely linked
genomicly.
How to break this chain and achieve the optimal balance between yield and resistance is a research topic with important application value in crop breeding, and finding the negative regulatory site of drought stress response to control yield traits and revealing its genetic and molecular mechanism is a major bottleneck scientific problem
facing the breeding of high-resistance and high-yield crops.

Small RNA (sRNA) is a class of short, non-coding RNA that is widely found
in eukaryotes.
sRNAs, especially miRNAs, have been reported to play an important regulatory role
in plant growth and development and responses to stress stress.
A class of inverted repeats (IRs) is prevalent in plant genomes that produce large amounts of sRNA
.
Although the sRNA produced by these IRs is known to regulate gene expression at the transcriptional and post-transcriptional levels, the specific regulatory function of IR on crop development has not been reported
in soybeans until recently.
Crop yields and environmental stress resistance are complex agronomic traits controlled by micro-potent polygenes, involving the expression and regulation
of many genes on a genome-wide scale.
Whether and how IR can control maize yield and environmental stress tolerance by generating sRNA to regulate gene expression is unknown
.

Through high-throughput sequencing, the authors obtained the sRNA expression of 338 maize-associated populations under normal watering (WW) and drought stress (DS) growth conditions and transcriptomes
of 197 of them.
More than 30,000 sRNAs in response to drought stress were detected and expressed at the population level
.
sRNA has a regulatory effect
on gene expression.
A total of 6158 pairs of sRNA-gene regulatory relationships were found through population-wide regression analysis, of which many known miRNA-gene regulatory relationships (such as miR168 regulation of AGO1c, miR169 regulation of NF-YA8, miR167 regulation of ARF6, etc.
), and many new sRNA regulatory relationships
on genes were also discovered 。 Genome-wide association analysis (eGWAS) of the above sRNA expression traits showed that more than 6,000 eQTLs regulated sRNA expression were obtained, of which 29 were eQTL hotspots, which could regulate the expression
of more than 20 sRNAs at the same time.
A comparative analysis of eQTL regulating sRNA expression and eQTL regulating gene expression showed that 4722 eQTLs could control the expression of sRNA and genes at the same time, indicating that there were a large number of genome-wide synergistic regulatory relationships
between sRNA and genes.

Figure 1.
Population-wide identification of sRNA and gene expression and resolution of their genetic regulatory mechanisms

Through predictive analysis, it was found that the sRNA generated by 29 eQTL hotspots regulates 2764 potential target genes
.
Among them, 530 genes (19.
2%) were associated with plant reproduction and 891 genes (32.
2%) were associated with stress response, indicating that these seQTL hotspots play an important role
in regulating maize yield and environmental adaptation.
To further elucidate the molecular regulatory mechanisms of crop yield and environmental adaptation, the authors cloned a drought-specific superhotspot eQTL DRESH8 on chromosome 8 that correlates with thousands of sRNA expressions and is detected
only under DS conditions.
Further analysis revealed that DRESH8 is a 21.
4 kb reverse repeating structure (TE-IR) variant
consisting of a gypsy-like transposable element (TE).
This TE-IR inserts the third intron of the ZmPP2C16 gene and is regulated by the promoter of the gene to induce initiation expression
in arid environments.
In addition, DRESH8 has no obvious regulatory effect on the inserted local gene, which may regulate the gene at the post-transcriptional level by producing sRNA
.

In the population, maize lacking DRESH8 had low sRNA expression, but its survival rate under drought stress was higher than that of maize
carrying DRESH8.
Furthermore, transgenic maize missing DRESH8 was obtained by gene editing, and it was found that the transgenic maize was more drought-resistant than wild-type control maize, and the sRNA produced was significantly reduced compared with the wild-type, indicating that DRESH8 was the causal variation site of drought resistance variation and sRNA expression variation in the maize population
.
Through mRNA cleavage experiments, it was found that the sRNA produced by DRESH8 had a cleavage effect
on about 30 genes downstream, including ZmMYBR38.
By overexpressing ZmMYBR38 gene in maize and Arabidopsis thaliana to detect the drought resistance of this gene, it was found that both groups of transgenic plants overexpressing ZmMYBR38 were more drought-tolerant than the corresponding wild-type non-transgenic plants, indicating that ZmMYBR38 played a conservative and positive role
in plant drought resistance.
These data suggest that siRNA produced by DRESH8 regulates drought tolerance in plants by mediating cleavage of downstream target mRNA
.

Figure 2.
DRESH8 inhibits drought resistance in maize by regulating the expression of target genes through the sRNA it produces

Next, the authors examine the evolution and selection
of DRESH8 in detail.
Using different analysis methods, it is found that there is a strong selection signal
at the DRESH8 site.
A total of thousands of maize germplasm resources, including maize ancestral grass, farm seeds and modern maize inbred lines, were used to further explore the origin and spread
of DRESH8.
The results show that the approximate origin of DRESH8 transposon is 51800 years ago, the deletion of DRESH8 may have been naturally selected during the adaptation of the more recent grass to environmental stress, and the presence of the DRESH8 allele may have been retained by artificial selection during the domestication and spread of maize
.
Considering that DRESH8 is not conducive to corn drought resistance, what drives DRESH8 to be chosen to be retained? To answer this important question, the authors obtained two sources of data
.
One is the relationship between
rainfall and DRESH8.
They found that rainfall was higher
in areas planted with DRESH8 corn than in areas without DRESH8.
The second is the relationship between
yield and DRESH8.
They found that in terms of yield traits (such as ear length, ear diameter, grain length, grain width, grain thickness, 100 grain weight, etc.
), wild-type corn containing DRESH8 was 5%-14% higher than gene-edited GM corn without DRESH8, that is, the presence of DRESH8 was conducive to increasing corn yield
。 Based on these results, the authors propose a model to explain how DRESH8 sites are selected: in an environment with abundant rainfall, farmers may choose to plant corn with DRESH8 to increase yields; In an arid environment, farmers choose to plant corn lacking DRESH8, which can remove the inhibitory effect of the sRNA produced by DRESH8 on drought-resistant genes, thereby improving the drought resistance of maize
.
These data show that DRESH8 can mediate the selective balance between drought tolerance and yield traits, which in turn affects the spread
of maize worldwide.

Figure 3.
DRESH8 EVOLUTION AND SELECTION

To more fully characterize the universal role of IR in maize in responding to drought, the authors studied the structure
of IR on a genome-wide scale.
Based on the B73 genome, a total of 8261 IRs were identified; Although these IRs account for only 1.
2% of the maize genome, they control the expression of about 42% of drought-responsive sRNAs, suggesting that IRs play a broad role
in maize's drought response.
85% of the corn genome is TE, and among the 8261 IRs, TE-IR accounts for 90.
7%, producing 86.
9% of IR sRNA
.
TE mainly produces 22-nt sRNA after forming IR structure, while TE without IR structure mainly produces 24-nt sRNA
.
In addition, 22-ntsRNA produced by IR accounts for about
70% of genome-wide 22-nt sRNA.
In TE-IR, most DNA transposons (except DHH and DTA) form a shorter IR (10 kb).

Most of the 21- and 22-nt sRNAs produced by IR are upregulated after drought, and these IR sRNAs also mediate responses including low temperature and salt stress, suggesting a universal role
in regulating plant responses to different environmental stresses.
It was further found that in the process of domestication and improvement, both IR length and number expanded, and the selection of grain length may be one of
the important driving factors for IR expansion.
In addition, there are 23 eQTL hotspots with TE-IR structural variations, of which 21 TE-IR-generated sRNA regulate potential target genes that are enriched to gene sets
related to yield and stress tolerance.
In summary, these data suggest that IR, especially TE-IR, may have a general role
in regulating the balance of environmental adaptability and yield traits in maize.

The authors identified a large number of environment-specific genetic regulators associated with drought adaptation and yield traits
.
The findings highlight the key role
that TE plays in the formation of IR structures.
These IR structures mainly control the expression of sRNA through DCL2 and are involved in post-transcriptional regulation, and this mode of IR is a key genetic and molecular regulatory mechanism
for the balance between maize environmental adaptability and crop yield traits.
This equilibrium mechanism, driven by the TE-IR structure, paves the way
for the cultivation of high-resistance and high-yield crops through precise genomic design.

Dr.
Xiang Yanli (now postdoctoral fellow at Ghent University), Dou Nannan, doctoral student Dou Nannan, and Zhang Hui, postdoctoral fellow of Huazhong Agricultural University, are the first authors
of the paper.
Professors Dai Mingqiu and Li Lin from the National Key Laboratory of Crop Genetic Improvement and Hongshan Laboratory, and Professor Li Feng from the Key Laboratory of Horticultural Plant Biology of the Ministry of Education are the corresponding authors
of the paper.
Professor Yan Jianbing from the National Key Laboratory of Crop Genetic Improvement and the Hongshan Laboratory also participated in the work
.
This research was supported by the International Cooperation and General Projects of the NSFC, the 13th Five-Year Key R&D Program, the Central University Fund Project, and the Hubei Hongshan Laboratory Key R&D Project
.

Reviewed by Dai Mingqiu

Online paper link: _msthash="101754" _msttexthash="4729556">Extended reading: [News Feature]" Drought? Yield? Fish and bear's paws can have both! ”