1. Volume 11, Issue 3, Pages 443-459 (March 2018)
Genome-wide Analysis of Transcriptional Variability in a Large Maize-Teosinte Population 
Xufeng Wang, Qiuyue Chen, Yaoyao Wu, Zachary H. Lemmon, Guanghui Xu, Cheng Huang, Yameng Liang, Dingyi Xu, Dan Li, John F. Doebley, Feng Tian 
Molecular Plant 
Volume 11, Issue 3, Pages (March 2018)
DOI: /j.molp
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2. Figure 1 Characterization of eQTL and eQTL-Regulated Genes.
(A) Number of eQTL mapped for each gene. The x axis and y axis represent the number of eQTL mapped for each gene and the number of genes in each group, respectively.
(B) The expression variation explained by local and distant eQTL. Local eQTL explains more expression variation than distant eQTL.
(C) The distribution of genes regulated by local and/or distant eQTL.
(D) Functional enrichment analysis for local eQTL-regulated genes. Each bubble stands for one functional class and the size of bubble indicates the number of enriched genes in each class. The y axis shows the P value of hypergeometric test with Benjamini and Hochberg multiple test correction.
Molecular Plant  , DOI: ( /j.molp )
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3. Figure 2 Expression Piggybacking and Co-regulated Gene Clusters.
(A) Genes with local eQTL more frequently occurred in adjacent positions than would be expected by chance. The x axis indicates the number of intervening genes between two annotated protein-coding genes from AGPv3. Red triangles represent the observed number of gene pairs with local eQTL mapped to them. The blue boxplots represent the distribution of gene pairs from 1000 permutations. The blue histogram in inset shows the distribution of adjacent gene pairs with no intervening genes based on 1000 permutations and the red arrow indicates the observed number of adjacent gene pairs having local eQTL mapped at both of them.
(B) Adjacent gene pairs having local eQTL mapped at both of them exhibited the highest expression correlations in the RILs. The red line indicates the distribution of correlation coefficients of adjacent genes with local eQTL mapped at both of them (class I; the average expression correlation coefficient is 0.25). The orange line indicates the expression correlations of adjacent genes with local eQTL detected at either gene (class II; the average expression correlation coefficient is 0.10). The purple line represents the expression correlations of adjacent genes without local eQTL mapped to both genes (class III; the average expression correlation coefficient is 0.08). The black dotted line indicates the expression correlations of non-adjacent genes with local eQTL mapped at both of them (class IV; the average expression correlation coefficient is 0.06). The red boxes with peaks on the top in the legend indicate genes with local eQTL detected.
(C) The number of co-regulated gene clusters. The x axis is the cluster size, and the y axis is the number of clusters in each size group. One-thousand permutations were performed to examine how often the co-regulated gene clusters occur by chance alone at P < 0.05.
Molecular Plant  , DOI: ( /j.molp )
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4. Figure 3
The Benzoxazinoid Metabolic Pathway Is under Coordinated Local Regulation.
(A) Benzoxazinoid metabolic pathway in maize. A total of 12 Bx genes are known to be involved in the maize benzoxazinoid metabolic pathway. The eight genes (Bx1–Bx8) shown in red reside in a cluster on chromosome four that primarily controls DIMBOA-Glc biosynthesis. The three genes (Bx10–Bx12) shown in blue reside in the cluster on chromosome 1 and are involved in the conversion of DIMBOA-Glc to HDMBOA-Glc. Bx9, which encodes the same enzyme as Bx8, is located on the long arm of chromosome 1. The benzoxazinoid synthesizing enzymes encoded by these Bx genes belong to four different gene families (labeled by color): Cyt P450, cytochrome-P450-dependent mono-oxygenase; UDP-GT, UDP-glucosyltransferase; 2-ODD, 2-oxoglutarate-dependent dioxygenase; and O-MT, O-methyltransferase.
(B and C) The local eQTL effects of the Bx genes. The boxplot shows the expression differences between maize and teosinte alleles at each Bx gene. The y axis represents the expression residual of each e-trait after accounting for confounders.
(D) Physical order of the Bx genes in the maize genome, and expression correlations among Bx genes.
(E) qRT–PCR assay for Bx genes with local effects. Student's t-test was used to detect significant expression differences between the maize and teosinte subgroups (P values are shown above the bars in red). The heights of the bars represent the means of the relative expression of 16 samples with SEs. M, maize allele (red bar); T, teosinte allele (blue bar).
Molecular Plant  , DOI: ( /j.molp )
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5. Figure 4 Genomic Distribution of Distant eQTL Hotspots.
The emanative bars along the outside of the circle comprising the 10 chromosomes indicate the distribution of the identified distant eQTL hotspots. Each bar represents one distant eQTL hotspot, and red color indicates hotspots whose targets are significantly enriched in specific metabolic pathways. The width and height of each bar indicate the physical interval of each hotspot and the number of targets regulated by the corresponding hotspot, respectively. Centromeres are indicated by black dots. Significant functional enrichments for genes regulated by distant eQTL hotspots are listed in detail in Supplemental Dataset 5.
Molecular Plant  , DOI: ( /j.molp )
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6. Figure 5
Genes Regulated by the Distant eQTL Hotspot 108 Are Significantly Enriched in the Glycolysis Pathway.
(A) Glycolysis pathway in maize. Hotspot 108 regulates seven target genes (blue) in the glycolysis pathway. The gene in red is hex9, encoding a key hexokinase in glycolysis; hex9 is located in the hotspot region and associated with significant local regulatory difference.
(B) hex9 negatively regulated the seven downstream genes in the glycolysis pathway. The red lines indicate the corresponding eQTL support intervals for hex9 and the seven target genes, respectively. The red triangle indicates the physical position of hex9 on chromosome 8. A significant local eQTL was detected at hex9, with the maize allele (red box) associated with higher expression than the teosinte allele (blue box). The distant eQTL at hotspot 108 showed consistent allelic effects on the expression of the seven downstream genes, with teosinte alleles (blue boxes) associated with higher expression than maize alleles (red boxes).
Molecular Plant  , DOI: ( /j.molp )
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7. Figure 6 trans Regulator of the Flavonoid Biosynthesis Pathway.
(A) The eight genes involved in the flavonoid biosynthetic pathway are regulated by the R1 locus underlying hotspot 125 on chromosome 10.
(B) R1 positively regulated the eight downstream genes in the flavonoid biosynthesis pathway. The y axis represents the expression residual of each e-trait after accounting for confounders. A significant local eQTL was detected at R1, and the teosinte allele (blue box) was associated with higher expression than the maize allele (red box). The distant eQTL at hotspot 125 showed consistent allelic effects on the expression of the eight downstream genes, and teosinte alleles were associated with higher expression than maize alleles.
Molecular Plant  , DOI: ( /j.molp )
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8. Figure 7
Impact of Domestication and Improvement in Shaping Global Gene Expression Pattern.
(A) The distribution of eQTL additive effects. Positive and negative additive effects indicate maize and teosinte allele up-regulated expression, respectively. The maize allele showed a more highly expression than the teosinte allele at 53% of mapped local eQTL, significantly different from a 50:50 unbiased expectation (P = 1.31E-11; Supplemental Table 5). “Dom” and “Imp” stand for the selection targets affected by maize domestication and improvement identified by Hufford et al. (2012), respectively. Dom and Imp genes exhibited stronger skew toward higher expression of maize alleles over teosinte alleles (60.7% and 61.4%, respectively), indicating the role of domestication selection in shaping the global gene expression pattern.
(B) Stronger bias toward higher expression of maize alleles was observed for large-effect local eQTL. The x axis indicates the bin interval of expression variation explained by local eQTL. The y axis represents the relative percentages of local eQTL with higher expression of maize alleles (red) and local eQTL with higher expression of teosinte alleles (gray). The numbers at the top of bars indicate numbers of local eQTL in the corresponding class.
Molecular Plant  , DOI: ( /j.molp )
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9. Figure 8
Bx Genes Showed Coordinated Regulatory Divergence between Maize and Teosinte.
(A) Genes exhibiting significant expression divergences or cis-regulatory differences between maize and teosinte are more likely detected with local eQTL.
(B) Cis effects of Bx genes across multiple maize-teosinte F1s in the ASE study of Lemmon et al. (2014). For the clustered Bx genes on chromosome 4, maize alleles showed higher expression than teosinte alleles. For Bx12, the teosinte allele was more highly expressed than the maize allele. Binomial test P values are shown to the right of bars indicating significant deviation from a 1:1 expression ratio in maize-teosinte F1 hybrids (Lemmon et al., 2014). N.A., not available.
(C) Comparison of the DIMBOA-Glc concentrations between maize and teosinte accessions. The left panel indicates that the DIMBOA-Glc concentration in maize inbreds (black bar) is significantly higher than that in teosinte accessions (red bar). The right panel shows significant differences detected between temperate maize (blue bar) and teosinte lines (red bar).
(D) Maize lines carrying the CACTA transposon insertion in Bx12 exhibited greatly decreased genetic diversity, as compared with teosinte lines. The upper schematic diagram displays the location of the transposon insertion. The red and green lines indicate the upstream and downstream portions of the transposon insertion, respectively, that were sequenced for nucleotide diversity analysis. The lower panels of the bar plots show nucleotide diversity of the upstream (red) and downstream (green) regions of the transposon insertion in teosinte accessions and in maize inbreds with (Maize+) or without (Maize−) the transposon insertion. The y axis represents the nucleotide diversity (π) for each subclass.
Molecular Plant  , DOI: ( /j.molp )
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