eQTLs play critical roles in regulating gene expression and identifying key regulators in rice.

The regulation of gene expression plays an essential role in both the phenotype and adaptation of plants. Transcriptome sequencing enables simultaneous identification of exonic variants and quantification of gene expression. Here, we sequenced the leaf transcriptomes of 287 rice accessions from around the world and obtained a total of 177 853 high-quality single nucleotide polymorphisms after filtering. Genome-wide association study identified 44 354 expression quantitative trait loci (eQTLs), which regulate the expression of 13 201 genes, as well as 17 local eQTL hotspots and 96 distant eQTL hotspots. Furthermore, a transcriptome-wide association study screened 21 candidate genes for starch content in the flag leaves at the heading stage. HS002 was identified as a significant distant eQTL hotspot with five downstream genes enriched for diterpene antitoxin synthesis. Co-expression analysis, eQTL analysis, and linkage mapping together demonstrated that bHLH026 acts as a key regulator to activate the expression of downstream genes. The transgenic assay revealed that bHLH026 is an important regulator of diterpenoid antitoxin synthesis and enhances the disease resistance of rice. These findings improve our knowledge of the regulatory mechanisms of gene expression variation and complex regulatory networks of the rice genome and will facilitate genetic improvement of cultivated rice varieties.

The coordination of OsbZIP72 and OsMYBS2 with reverse roles regulates the transcription of OsPsbS1 in rice.

Nonphotochemical quenching (NPQ), an intricate photoprotective process, plays fundamental roles in maintaining plant fitness. The PsbS protein is essential for the rapid induction of NPQ, and acts in a dose-dependent manner in leaves. However, little information is known on the transcriptional control of PsbS in land plants. Here we demonstrated that the expression of OsPsbS1 is directly upregulated by OsbZIP72 while repressed by OsMYBS2 in rice. We identified a new cis-element GACAGGTG in japonica OsPsbS1 promoter, to which OsbZIP72 could strongly bind and activate the expression of OsPsbS1. The new cis-element CTAATC confers specific binding for OsMYBS2 in japonica OsPsbS1 promoter. OsbZIP72 can be activated by SAPK1, and acts depending on the abscisic acid (ABA) signalling pathway. GF14A protein affects the repression activity of OsMYBS2 by regulating its nucleocytoplasmic shuttling, and Ser53 is necessary for OsMYBS2 to be retained in the cytoplasm. The inducibility of OsPsbS1 transcription under high light conditions in OsbZIP72 knockout lines was greatly impaired, while the repression of OsPsbS1 transcription under a low light environment in OsMYBS2 knockout lines was significantly alleviated. These results reveal cross-talk among NPQ processes, the ABA signalling pathway and abiotic stress signalling. The elaborate mechanisms may help enhance photoprotection and improve photosynthesis in rice.

Genome-wide association study of flowering time reveals complex genetic heterogeneity and epistatic interactions in rice.

Since domestication, rice has cultivated in a wide range of latitudes with different day lengths. Selection of diverse natural variations in heading date and photoperiod sensitivity is critical for adaptation of rice to different geographical environments. To unravel the genetic architecture underlying natural variation of rice flowering time, we conducted a genome wide association study (GWAS) using several association analysis strategies with a diverse worldwide collection of 529 O. sativa accessions. Heading date was investigated in three environments under long-day or short-day conditions, and photosensitivity was evaluated. By dividing the whole association panel into subpopulations and performing GWAS with both linear mixed models and multi-locus mixed-models, we revealed hundreds of significant loci harboring novel candidate genes as well as most of the known flowering time genes. In total, 127 hotspots were detected in at least two GWAS. Universal genetic heterogeneity was found across subpopulations. We further detected abundant interactions between GWAS loci, especially in indica. Functional gene families were revealed from enrichment analysis of the 127 hotspots. The results demonstrated a rich of genetic interactions in rice flowering time genes and such epistatic interactions contributed to the large portions of missing heritability in GWAS. It suggests the increased complexity of genetic heterogeneity might discount the power of increasing the sample sizes in GWAS.

Gut microbiota correlates with fiber and apparent nutrients digestion in goose.

To explore the relationship among the level of fiber, gut microbiota, and nutritional substances, we applied the next generation sequencing technology for the identification of the composition and structure of microbiota in the gastrointestinal tract. In this study, a total of 25 phyla and 298 genera were identified from the gastrointestinal tract; Firmicutes, Bacteroidetes, and Proteobacteria were the predominant phyla. The ability of cecum in carbohydrate metabolism was significantly higher than that of the gizzard and ileum (P < 0.05). The bacterial community structure in various stages of the development of the cecum was different. In the different growth stages of cecum, the increase in the microbiota structure of the fiber level elevates the ability of carbon hydration. Second, the apparent metabolic rates of the other nutrients were affected by the fiber and period except for acid detergent fiber (P < 0.05); the apparent utilization rate of the nutrients increased with time. However, with the increase in the fiber level, the apparent utilization of nutrients was initially increased, followed by a decrease. Therefore, a correlation was established between the fiber level and gastrointestinal microbiota and apparent nutrient utilization rate of the 3 phyla. Our results suggest that the fiber level and growth stages could impact the composition of gut microbiota.

Genetic Architecture of Natural Variation in Rice Nonphotochemical Quenching Capacity Revealed by Genome-Wide Association Study.

The photoprotective processes conferred by nonphotochemical quenching (NPQ) serve fundamental roles in maintaining plant fitness and sustainable yield. So far, few loci have been reported to be involved in natural variation of NPQ capacity in rice (Oryza sativa), and the extents of variation explored are very limited. Here we conducted a genome-wide association study (GWAS) for NPQ capacity using a diverse worldwide collection of 529 O. sativa accessions. A total of 33 significant association loci were identified. To check the validity of the GWAS signals, three F2 mapping populations with parents selected from the association panel were constructed and assayed. All QTLs detected in mapping populations could correspond to at least one GWAS signal, indicating the GWAS results were quite reliable. OsPsbS1 was repeatedly detected and explained more than 40% of the variation in the whole association population in two years, and demonstrated to be a common major QTL in all three mapping populations derived from inter-group crosses. We revealed 43 single nucleotide polymorphisms (SNPs) and 7 insertions and deletions (InDels) within a 6,997-bp DNA fragment of OsPsbS1, but found no non-synonymous SNPs or InDels in the coding region, indicating the PsbS1 protein sequence is highly conserved. Haplotypes with the 2,674-bp insertion in the promoter region exhibited significantly higher NPQ values and higher expression levels of OsPsbS1. The OsPsbS1 RNAi plants and CRISPR/Cas9 mutants exhibited drastically decreased NPQ values. OsPsbS1 had specific and high-level expression in green tissues of rice. However, we didn't find significant function for OsPsbS2, the other rice PsbS homologue. Manipulation of the significant loci or candidate genes identified may enhance photoprotection and improve photosynthesis and yield in rice.

Genetic Architecture of Natural Variation in Rice Chlorophyll Content Revealed by a Genome-Wide Association Study.

Chlorophyll content is one of the most important physiological traits as it is closely related to leaf photosynthesis and crop yield potential. So far, few genes have been reported to be involved in natural variation of chlorophyll content in rice (Oryza sativa) and the extent of variations explored is very limited. We conducted a genome-wide association study (GWAS) using a diverse worldwide collection of 529 O. sativa accessions. A total of 46 significant association loci were identified. Three F2 mapping populations with parents selected from the association panel were tested for validation of GWAS signals. We clearly demonstrated that Grain number, plant height, and heading date7 (Ghd7) was a major locus for natural variation of chlorophyll content at the heading stage by combining evidence from near-isogenic lines and transgenic plants. The enhanced expression of Ghd7 decreased the chlorophyll content, mainly through down-regulating the expression of genes involved in the biosynthesis of chlorophyll and chloroplast. In addition, Narrow leaf1 (NAL1) corresponded to one significant association region repeatedly detected over two years. We revealed a high degree of polymorphism in the 5' UTR and four non-synonymous SNPs in the coding region of NAL1, and observed diverse effects of the major haplotypes. The loci or candidate genes identified would help to fine-tune and optimize the antenna size of canopies in rice breeding.