Calmodulin (CaM), a ubiquitous calcium-binding protein, regulates diverse cellular functions by modulating the activity of a variety of enzymes and proteins. Plants express numerous CaM isoforms that exhibit differential activation and/or inhibition of CaM-dependent enzymes in vitro. However, the specific biological functions of plant CaM are not well known. In this study, we isolated a cDNA encoding a CaM binding transcription factor, MYB2, that regulates the expression of salt- and dehydration-responsive genes in Arabidopsis. This was achieved using a salt-inducible CaM isoform (GmCaM4) as a probe from a salt-treated Arabidopsis expression library. Using domain mapping, we identified a Ca2+-dependent CaM binding domain in MYB2. The specific binding of CaM to CaM binding domain was confirmed by site-directed mutagenesis, a gel mobility shift assay, split ubiquitin assay, and a competition assay using a Ca2+/CaM-dependent enzyme. Interestingly, the specific CaM isoform GmCaM4 enhances the DNA binding activity of AtMYB2, whereas this was inhibited by a closely related CaM isoform (GmCaM1). Overexpression of Gm-CaM4 in Arabidopsis up-regulates the transcription rate of AtMYB2-regulated genes, including the proline-synthesizing enzyme P5CS1 (Delta1-pyrroline-5-carboxylate synthetase-1), which confers salt tolerance by facilitating proline accumulation. Therefore, we suggest that a specific CaM isoform mediates salt-induced Ca2+ signaling through the activation of an MYB transcriptional activator, thereby resulting in salt tolerance in plants.

Calmodulin (CaM) regulates diverse cellular functions by modulating the activities of a variety of enzymes and proteins. However, direct modulation of transcription factors by CaM has been poorly understood. In this study, we isolated a putative transcription factor by screening a rice cDNA expression library by using CaM:horseradish peroxidase as a probe. This factor, which we have designated OsCBT (Oryza sativa CaM-binding transcription factor), has structural features similar to Arabidopsis AtSRs/AtCAMTAs and encodes a 103-kDa protein because it contains a CG-1 homology DNA-binding domain, three ankyrin repeats, a putative transcriptional activation domain, and five putative CaM-binding motifs. By using a gel overlay assay, gel mobility shift assays, and site-directed mutagenesis, we showed that OsCBT has two different types of functional CaM-binding domains, an IQ motif, and a Ca 2؉ -dependent motif. To determine the DNA binding specificity of OsCBT, we employed a random binding site selection method. This analysis showed that OsCBT preferentially binds to the sequence 5-TWCG(C/T)GTKKKKTKCG-3 (W and K represent A or C and T or G, respectively). OsCBT was able to bind this sequence and activate ␤-glucuronidase reporter gene expression driven by a minimal promoter containing tandem repeats of these sequences in Arabidopsis leaf protoplasts. Green fluorescent protein fusions of two putative nuclear localization signals of OsCBT, a bipartite and a SV40 type, were predominantly localized in the nucleus. Most interestingly, the transcriptional activation mediated by OsCBT was inhibited by co-transfection with a CaM gene. Taken together, our results suggest that OsCBT is a transcription activator modulated by CaM.

Calmodulin (CaM) is a ubiquitous Ca 2+ -binding protein known to regulate diverse cellular functions by modulating the activity of various target proteins. We isolated a cDNA encoding AtWRKY7, a novel CaM-binding transcription factor, from an Arabidopsis expression library with horseradish peroxidase-conjugated CaM. CaM binds specifically to the Ca 2+ -dependent CaM-binding domain (CaMBD) of AtWRKY7, as shown by site-directed mutagenesis, a gel mobility shift assay, a split-ubiquitin assay, and a competition assay using a Ca 2+ / CaM-dependent enzyme. Furthermore, we show that the CaMBD of AtWRKY7 is a conserved structural motif (C-motif) found in group IId of the WRKY protein family.

Transient influx of Ca2؉ constitutes an early event in the signaling cascades that trigger plant defense responses. However, the downstream components of defense-associated Ca 2؉ signaling are largely unknown. Because Ca 2؉ signals are mediated by Ca 2؉ -binding proteins, including calmodulin (CaM), identification and characterization of CaM-binding proteins elicited by pathogens should provide insights into the mechanism by which Ca 2؉ regulates defense responses. In this study, we isolated a gene encoding rice Mlo (Oryza sativa Mlo; OsMlo) using a protein-protein interactionbased screening of a cDNA expression library constructed from pathogen-elicited rice suspension cells. OsMlo has a molecular mass of 62 kDa and shares 65% sequence identity and scaffold topology with barley Mlo, a heptahelical transmembrane protein known to function as a negative regulator of broad spectrum disease resistance and leaf cell death. By using gel overlay assays, we showed that OsMlo produced in Escherichia coli binds to soybean CaM isoform-1 (SCaM-1) in a Ca 2؉ -dependent manner. We located a 20-amino acid CaMbinding domain (CaMBD) in the OsMlo C-terminal cytoplasmic tail that is necessary and sufficient for Ca 2؉ -dependent CaM complex formation. Specific binding of the conserved CaMBD to CaM was corroborated by sitedirected mutagenesis, a gel mobility shift assay, and a competition assay with a Ca 2؉ /CaM-dependent enzyme. Expression of OsMlo was strongly induced by a fungal pathogen and by plant defense signaling molecules. We propose that binding of Ca 2؉ -loaded CaM to the C-terminal tail may be a common feature of Mlo proteins.

Globally, soybean is a major protein and oil crop. Enhancing our understanding of the soybean domestication and improvement process helps boost genomics-assisted breeding efforts. Here we present a genome-wide variation map of 10.6 million single-nucleotide polymorphisms and 1.4 million indels for 781 soybean individuals which includes 418 domesticated (Glycine max), 345 wild (Glycine soja), and 18 natural hybrid (G. max/G. soja) accessions. We describe the enhanced detection of 183 domestication-selective sweeps and the patterns of putative deleterious mutations during domestication and improvement. This predominantly selfing species shows 7.1% reduction of overall deleterious mutations in domesticated soybean relative to wild soybean and a further 1.4% reduction from landrace to improved accessions. The detected domestication-selective sweeps also show reduced levels of deleterious alleles. Importantly, genotype imputation with this resource increases the mapping resolution of genome-wide association studies for seed protein and oil traits in a soybean diversity panel.

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This is an Open-Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.Plant Breed. Biotech. 2015 (September) is one of the most important crops in the world and in Korea as well. Since the official start of soybean breeding program in Korea at which a landrace 'Jangdanbaekmok' was first released to promote cultivation in 1913, approximately one century has elapsed. Currently, a total of 178 soybean varieties are registered at two representative Korean national institutes, the RDA-Genebank Information Center (http://www.genebank.go.kr) and the Korea Seed & Variety Service (http://www.seed.go.kr). Of these, 155 varieties (87.1%) have been developed through hybridization-based breeding technologies, of which most cultivars (133 varieties, 85.8%) have been released in the last twenty five years. In this review, we attempted to integrate all the information for individual cultivars and to rebuild a breeding pedigree including the entirety of registered Korean soybean varieties. The analysis has resulted in a total of four pedigrees involving 168 cultivars (94.4% out of 178 cultivars), which form the broadest network of pedigrees. Each of pedigrees highlights different key varieties within the context of progenitor networks derived from crossing of various elite parental lines as follows; pedigree I-'Kwangkyo', 'Hwangkeumkong', 'Paldalkong' and 'Sinpaldalkong2', pedigree II-'Baegunkong', 'Jangyeobkong' and 'Keunolkong', pedigree III-'Danyeob', 'Pangsa' and 'Eunhakong'. These pedigrees also reveal purpose (i.e., desirable traits)-driven development of characteristic soybean varieties during the past century of breeding history in Korea. We expect that the pedigree reconstructed in this study will provide breeders with information useful to design breeding schema and guidance towards the genomics-assisted soybean improvement in the future.

Key message Genotyping data of a comprehensive Korean soybean collection obtained using a large SNP array were used to clarify global distribution patterns of soybean and address the evolutionary history of soybean. Abstract Understanding diversity and evolution of a crop is an essential step to implement a strategy to expand its germplasm base for crop improvement research. Accessions intensively collected from Korea, which is a small but central region in the distribution geography of soybean, were genotyped to provide sufficient data to underpin population genetic questions. After removing natural hybrids and duplicated or redundant accessions, we obtained a non-redundant set comprising 1957 domesticated and 1079 wild accessions to perform population structure analyses. Our analysis demonstrates that while wild soybean germplasm will require additional sampling from diverse indigenous areas to expand the germplasm base, the current domesticated soybean germplasm is saturated in terms of genetic diversity. We then showed that our genome-wide polymorphism map enabled us to detect genetic loci underlying flower color, seed-coat color, and domestication syndrome. A representative soybean set consisting of 194 accessions was divided into one domesticated subpopulation and four wild subpopulations that could be traced back to their geographic collection areas. Population genomics analyses suggested that the monophyletic group of domesticated soybeans was likely originated at a Japanese region. The results were further substantiated by a phylogenetic tree constructed from domestication-associated single nucleotide polymorphisms identified in this study. Electronic supplementary material The online version of this article (10.1007/s00122-018-3271-7) contains supplementary material, which is available to authorized users.