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Metal homeostasis has evolved to tightly modulate the availability of metals within the cell, avoiding cytotoxic interactions due to excess and protein inactivity due to deficiency. Even in the presence of homeostatic processes, however, low bioavailability of these essential metal nutrients in soils can negatively impact crop health and yield. While research has largely focused on how plants assimilate metals, acclimation to metal-limited environments requires a suite of strategies that are not necessarily involved in metal transport across membranes. The identification of these mechanisms provides a new opportunity to improve metal-use efficiency and develop plant foodstuffs with increased concentrations of bioavailable metal nutrients. Here, we investigate the function of two distinct subfamilies of the nucleotide-dependent metallochaperones (NMCs), named ZNG1 and ZNG2, that are found in plants, using Arabidopsis thaliana as a reference organism. AtZNG1 (AT1G26520) is an ortholog of human and fungal ZNG1, and like its previously characterized eukaryotic relatives, localizes to the cytosol and physically interacts with methionine aminopeptidase type I (AtMAP1A). Analysis of AtZNG1, AtMAP1A, AtMAP2A, and AtMAP2B transgenic mutants are consistent with the role of Arabidopsis ZNG1 as a Zn transferase for AtMAP1A, as previously described in yeast and zebrafish. Structural modeling reveals a flexible cysteine-rich loop that we hypothesize enables direct transfer of Zn from AtZNG1 to AtMAP1A during GTP hydrolysis. Based on proteomics and transcriptomics, loss of this ancient and conserved mechanism has pleiotropic consequences impacting the expression of hundreds of genes, including those involved in photosynthesis and vesicle transport. Members of the plant-specific family of NMCs, ZNG2A1 (AT1G80480) and ZNG2A2 (AT1G15730), are also required during Zn deficiency, but their target protein(s) remain to be discovered. RNA-seq analyses reveal wide-ranging impacts across the cell when the genes encoding these plastid-localized NMCs are disrupted.

Common weedy rice plants are important genetic resources for modern breeding programs because they are the closest relatives to rice cultivars and their genomes contain elite genes. Determining the utility and copy numbers of WRKY and nucleotide-binding site (NBS) resistance-related genes may help to clarify their variation patterns and lead to crop improvements. In this study, the weedy rice line LM8 was examined at the whole-genome level. To identify the Oryza sativa japonica subpopulation that LM8 belongs to, the single nucleotide polymorphisms (SNPs) of 180 cultivated and 23 weedy rice varieties were used to construct a phylogenetic tree and a principal component analysis and STRUCTURE analysis were performed. The results indicated that LM8 with admixture components from japonica (GJ) and indica (XI) belonged to GJ-admixture (GJ-adm), with more than 60% of its genetic background derived from XI-2 (22.98%), GJ-tropical (22.86%), and GJ-subtropical (17.76%). Less than 9% of its genetic background was introgressed from weedy rice. Our results also suggested LM8 may have originated in a subtropical or tropical geographic region. Moreover, the comparisons with Nipponbare (NIP) and Shuhui498 (R498) revealed many specific structure variations (SVs) in the LM8 genome and fewer SVs between LM8 and NIP than between LM8 and R498. Next, 96 WRKY and 464 NBS genes were identified and mapped on LM8 chromosomes to eliminate redundancies. Three WRKY genes (ORUFILM02g002693, ORUFILM05g002725, and ORUFILM05g001757) in group III and one RNL [including the resistance to powdery mildew 8 (RPW8) domain, NBS, and leucine rich repeats (LRRs)] type NBS gene (ORUFILM12g000772) were detected in LM8. Among the NBS genes, the RPW8 domain was detected only in ORUFILM12g000772. This gene may improve plant resistance to pathogens as previously reported. Its classification and potential utility imply LM8 should be considered as a germplasm resource relevant for rice breeding programs.

The weedy rice (Oryza sativa f. spontanea) pericarp has diverse colors (e.g., purple, red, light-red, and white). However, research on pericarp colors has focused on red and purple, but not green. Unlike many other common weedy rice resources, LM8 has a green pericarp at maturity. In this study, the coloration of the LM8 pericarp was evaluated at the cellular and genetic levels. First, an examination of their ultrastructure indicated that LM8 chloroplasts were normal regarding plastid development and they contained many plastoglobules from the early immature stage to maturity. Analyses of transcriptome profiles and differentially expressed genes revealed that most chlorophyll (Chl) degradation-related genes in LM8 were expressed at lower levels than Chl a/b cycle-related genes in mature pericarps, suggesting that the green LM8 pericarp was associated with inhibited Chl degradation in intact chloroplasts. Second, the F2 generation derived from a cross between LM8 (green pericarp) and SLG (white pericarp) had a pericarp color segregation ratio of 9:3:4 (green:brown:white). The bulked segregant analysis of the F2 populations resulted in the identification of 12 known genes in the chromosome 3 and 4 hotspot regions as candidate genes related to Chl metabolism in the rice pericarp. The RNA-seq and sqRT-PCR assays indicated that the expression of the Chl a/b cycle-related structural gene DVR (encoding divinyl reductase) was sharply up-regulated. Moreover, genes encoding magnesium-chelatase subunit D and the light-harvesting Chl a/b-binding protein were transcriptionally active in the fully ripened dry pericarp. Regarding the ethylene signal transduction pathway, the CTR (encoding an ethylene-responsive protein kinase) and ERF (encoding an ethylene-responsive factor) genes expression profiles were determined. The findings of this study highlight the regulatory roles of Chl biosynthesis- and degradation-related genes influencing Chl accumulation during the maturation of the LM8 pericarp.

Genes have been lost or weakened from cultivated rice during rice domestication and breeding. Weedy rice (Oryza sativa f. spontanea) is usually recognized as the progeny between cultivated rice and wild rice and is also known to harbor an gene pool for rice breeding. Therefore, identifying genes from weedy rice germplasms is an important way to break the bottleneck of rice breeding. To discover genes from weedy rice germplasms, we constructed a genetic map based on w-hole-genome sequencing of a F2 population derived from the cross between LM8 and a cultivated rice variety. We further identified 31 QTLs associated with 12 important agronomic traits and revealed that ORUFILM03g000095 gene may play an important role in grain length regulation and participate in grain formation. To clarify the genomic characteristics from weedy rice germplasms of LM8, we generated a high-quality genome assembly using single-molecule sequencing, Bionano optical mapping, and Hi-C technologies. The genome harbored a total size of 375.8 Mb, a scaffold N50 of 24.1 Mb, and originated approximately 0.32 million years ago (Mya) and was more closely related to Oryza sativa ssp. japonica. and contained 672 unique genes. It is related to the formation of grain shape, heading date and tillering. This study generated a high-quality reference genome of weedy rice and high-density genetic map that would benefit the analysis of genome evolution for related species and suggested an effective way to identify genes related to important agronomic traits for further rice breeding.

Growing cultivated rice with a moderate heading date is the key to expanding its cultivation area and maintaining stable yields. The genes that regulate heading date are largely cloned; however, it remains unclear how genetic mutations and their combinations affect the heading date and adaptability of cultivated rice. Here, we report the analysis of genetic variation in eight long-day flowering suppressor genes (Hd1, DTH8, Ghd7, OsCOL4, DTH7, Hd6, Se5, and PhyB) and the phylogenetic relationship of eight genes. Genetic variations in DTH8, Ghd7, Hd1, DTH7, PhyB, and OsCOL4 are correlated with differences in heading date and the correlation between the genetic diversity of Hd6 and Se5 and rice heading data are weak. One group of haplotypes of DTH8, Ghd7, Hd1, DTH7, PhyB, and OsCOL4 are associated with earlier heading dates and appear to have accumulated during the northward expansion of rice cultivation. A minimum of four group A alleles of DTH8, Ghd7, Hd1, DTH7, PhyB, and OsCOL4 are required for the growth of cultivated rice at latitudes above 30°N. This study presents a preliminary investigation of the genetic patterns and adaptation mechanisms of long-day flowering suppressor genes and provides a useful reference for the molecular breeding of rice cultivars for various environments and farming systems.