Soon Il Kwon

Molecular characterization of Oryza sativa WRKY6, which binds to W-box-like element 1 of the Oryza sativa pathogenesis-related (PR) 10a promoter and confers reduced susceptibility to pathogens

WRKY proteins are transcription factors (TFs) that regulate the expression of defense-related genes. The salicylic acid (SA)-inducible Oryza sativa WRKY6 (OsWRKY6) was identified as a positive regulator of Oryza sativa pathogenesis-related 10a (OsPR10a) by transient expression assays. A physical interaction between OsWRKY6 and W-box-like element 1 (WLE1), which positively regulates OsPR10a/probenazole induced protein 1 expression, was verified in vitro. Several pathogenesis-related (PR) genes were constitutively activated, including OsPR10a, and transgenic rice (Oryza sativa) plants overexpressing (ox) OsWRKY6 exhibited enhanced disease resistance to pathogens. By contrast, PR gene induction was compromised in transgenic OsWRKY6-RNAi lines, suggesting that OsWRKY6 is a positive regulator of defense responses. OsWRKY6-ox lines displayed leaf lesions, and increased OsWRKY6 levels caused cell death. Salicylic acid (SA) concentrations were higher in OsWRKY6-ox lines than in wild-type (WT) plants, and transcript levels of Oryza sativa isochorismate synthase 1 (OsICS1), which encodes a major enzyme involved in SA biosynthesis, were higher in OsWRKY6-ox lines than in WT. OsWRKY6 directly bound to the OsICS1 promoter in vivo. This indicates that OsWRKY6 can directly regulate OsICS1 expression and thereby increase SA concentrations. OsWRKY6 autoregulates its own expression. OsWRKY6 protein degradation is possibly regulated by ubiquitination. Our results suggest that OsWRKY6 positively regulates defense responses through activation of OsICS1 expression and OsWRKY6 stabilization.

Expression analysis of rice VQ genes in response to biotic and abiotic stresses

WRKY transcription factors are encoded by a large gene superfamily with a broad range of roles in plants. Proteins containing a short VQ (FxxxVQxLTG) motif have been recently shown to interact with WRKY transcription factors, implying that AtVQ proteins are important in the plant defense responses in Arabidopsis, either as positive or negative cofactors of WRKY transcription factors. Thirty-nine Oryza sativa genes containing the VQ motif (OsVQs) were identified and the genome structures of OsVQ proteins were characterized through genome-wide analysis in rice. Also, phylogenetic tree analysis was performed with the VQ domain of Arabidopsis and rice. The expression patterns of these OsVQ genes in plants under several stress treatments were assessed, specifically, following infection with the bacterial pathogen Xanthomonas oryzae pv. oryzae (Xoo), treatment with abscisic acid (ABA), or exposure to drought. The cellular localization of a few OsVQ proteins was examined using rice protoplast system. Based on our results, we suggest that OsVQ proteins function as important co-regulators during the plant defense response to biotic and abiotic stresses.

Identification of S-allele genotypes of Korean apple cultivars by using allele-specific polymerase chain reaction

By using allele-specific polymerase chain reaction, self-incompatibility (S)-genotypes of the Korean apple cultivars were analyzed to identify their self-incompatibility relationship. S-genotypes of the cultivars were determined as ‘Hongro’, ‘Saenara’, ‘Hongso’, and ‘Hongan’, S1S3; ‘Picnic’, S1S5; ‘Gamhong’, ‘Manbok’, ‘Hwarang’, ‘Yeohong’, ‘Danhong’, and ‘Hwayoung’, S1S9; ‘Seokwang’, S3S5; ‘Honggeum’ and ‘Greenball’, S3S7; ‘Sunhong’, ‘Seohong’, ‘Chukwang’, and ‘Hwahong’, S3S9; and ‘Summer Dream’, S7S9. These results will help to select compatible pollinizers and design apple orchards.

Identification of AtWRKY75 as a transcriptional regulator in the defense response to Pcc through the screening of Arabidopsis activation-tagged lines

The necrotrophic pathogen Pectobacterium carotovorum ssp. carotovorum (Pcc) causes soft rot in a broad range of plant hosts. Approximately 60,000 independent seeds from Arabidopsis activation tagging lines were inoculated with Pcc and screened for resistant mutants. An Rpe1 (resistance protein to Pectobacterium 1) mutant, which had more resistance to Pcc than wild-type (WT) plants, was selected for further study. The T-DNA inserting locus in Rpe1 was located on the middle of chromosome V by flanking sequence analysis. Through expression analysis with several genes adjacent to the T-DNA tagging region, AtWRKY75 gene was highly up-regulated in the Rpe1 mutant compared to the WT plant. The up-regulation of AtWRKY75 gene was shown to be correlated on the induction of the PDF1.2, VSP1 and PR1 genes compared to the WT plant. AtWRKY75 over-expression lines exhibited reduced Pcc bacterial growth compared to WT. Taken together, our data suggest that AtWRKY75 should be a positive regulator in the JA- or SA-mediated defense signaling responses to Pcc.

OsWRKY51, a rice transcription factor, functions as a positive regulator in defense response against Xanthomonas oryzae pv. oryzae

Key messageOsWRKY51 functions as a positive transcriptional regulator in defense signaling againstXanthomonas oryzae pv. oryzaeby direct DNA binding to the promoter of defense related gene, OsPR10a.AbstractOsWRKY51 in rice (Oryza sativa L.) is induced by exogenous salicylic acid (SA) and inoculation with Xanthomonas oryzae pv. oryzae (Xoo). To examine the role of OsWRKY51 in the defense response of rice, we generated OsWRKY51 overexpressing and underexpressing transgenic rice plants. OsWRKY51-overexpressing transgenic rice lines were more resistant to Xoo and showed greater expression of defense-related genes than wild-type (WT) plants, while OsWRKY51-underexpressing lines were more susceptible to Xoo and showed less expression of defense-associated genes than WT plants. Transgenic lines overexpressing OsWRKY51 showed growth retardation compared to WT plants. In contrast, transgenic lines underexpressing OsWRKY51 by RNA interference showed similar plant height with WT plants. Transient expression of OsWRKY51-green fluorescent protein fusion protein in rice protoplasts revealed that OsWRKY51 was localized in the nucleus. OsWRKY51 bound to the W-box and WLE1 elements of the OsPR10a promoter. Based on these results, we suggest that OsWRKY51 is a positive transcriptional regulator of defense signaling and has direct DNA binding ability to the promoter of OsPR10a, although it is reported to be a negative regulator in GA signaling.

Genetic identification of apple cultivars bred in Korea using simple sequence repeat markers

Among the many DNA markers currently available, simple sequence repeat (SSR) markers are well developed and widely used for gene identification and cultivar discrimination by apple breeders and researchers. We developed SSR fingerprints for identification and to reveal the genetic diversity of 22 apple cultivars released from Korea. We used a total of 22 SSR markers. Among them, 15 SSR markers were developed from the complete sequence of ‘Golden Delicious’ and 7 SSR markers were chosen from the HiDRAS database. Except for somatic mutants, cultivar identification of all Korean bred apple genotypes was accomplished by using only two SSR markers (CH03d07 and KFBG-7). Parent-offspring relationships analysis was performed to investigate the real parents of three suspicious cultivars (‘Gamhong’, ‘Hongan’, and ‘Green Ball’). Based on the SSR markers experiment and reported S-RNase allele, we confirmed that ‘Fuji’ was used as paternal parent of ‘Gamhong’ and either ‘Hongro’ or ‘Natumidori’ were used as paternal parent of ‘Hongan’. Moreover, ‘Jonathan’ rather than ‘Fuji’ seems to be a paternal parent of ‘Green Ball’.

Inbreeding, coancestry, and founding clones of apple cultivars released from Korea

Inbreeding and coancestry coefficients were calculated for 20 apple (Malus × domestica Borkh.) cultivars released from Korea using their recorded pedigree information. The overall mean inbreeding coefficient was 0.034 for all 20 cultivars. Frequently used founding clones were ‘Grimes Golden’, ‘Delicious’, ‘Ralls Janet’, ‘Earli Blaze’, and ‘Esopus Spitzenburg’. Coefficients of coancestry among all 20 cultivars and the above mentioned five founding clones averaged 0.075, 0.075, 0.0075, 0.069, and 0.024, respectively. The average coancestry within the apple cultivars released from breeding program was 0.136 and mean coefficient ranged from 0.115 to 0.166, which is comparable to 0.125 for half-sib relationship. The redundant use of the same parents and their progenies can narrow the genetic base, thereby limiting genetic gain. Strategies are suggested for maintaining and increasing the genetic diversity, as well as for careful examining of pedigrees for future apple breeding in Korea.

Assessment of apple core collections constructed using phenotypic and genotypic data

Several types of information can be used to select core collections, including passport data, agronomic data, and molecular data. However, little is known about the ability of core collections to retain the genetic diversity and structure of the whole collection for characters that were not considered during the selection, particularly when molecular markers are used. In this study, two core subsets were established for the apple (Malus spp) germplasm bank curated at the Apple Research Station, National Institute of Horticultural and Herbal Science, Korea, based upon genetic diversity estimated with 14 simple sequence repeat markers, and phenotypic diversity based on 23 traits. Comparisons between these two subsets and with the whole collection were used to determine the effect of the data used in the selection on phenotypic and genetic diversity, and population structure. The two subsets had a similar diversity and did not differ from the original collection, according to the Nei and Shannon diversity indices. Allele and class frequencies were also maintained in the two subsets. Overall, the type of data used to construct the core collection had little influence on the phenotypic and genetic diversity retained. Therefore, in the case of apple collections, the use of molecular markers is preferable, because they allow rapid and reliable characterization.