Jun-Cheol Moon

A gene family encoding RING finger proteins in rice: their expansion, expression diversity, and co-expressed genes

The proteins harboring RING finger motif(s) have been shown to mediate protein–protein interactions that are relevant to a variety of cellular processes. In an effort to elucidate the evolutionary dynamics of the rice RING finger protein family, we have attempted to determine their genomic locations, expression diversity, and co-expressed genes via in silico analysis and semi-quantitative RT–PCR. A total of 425 retrieved genes appear to be distributed over all 12 of the chromosomes of rice with different distributions, and are reflective of the evolutionary dynamics of the rice genome. A genome-wide dataset harboring 155 gene expression omnibus sample plates evidenced some degree of differential evolutionary fates between members of RING-H2 and RING-HC types. Additionally, responses to abiotic stresses, such as salinity and drought, demonstrated that some degree of expression diversity existed between members of the RING finger protein genes. Interestingly, we determined that one RING-H2 finger protein gene (Os04g51400) manifested striking differences in expression patterns in response to abiotic stresses between leaf and culm-node tissues, further revealing responses highly similar to the majority of randomly selected co-expressed genes. The gene network of genes co-expressed with Os04g51400 may suggest some role in the salt response of the gene. These findings may shed further light on the evolutionary dynamics and molecular functional diversity of these proteins in complex cellular regulations.

Molecular dissection of the response of a rice leucine-rich repeat receptor-like kinase (LRR-RLK) gene to abiotic stresses

Leucine-rich repeat (LRR) receptor-like kinase (RLK) proteins play key roles in a variety of biological pathways. In a previous study, we analyzed the members of the rice LRR-RLK gene family using in silico analysis. A total of 23 LRR-RLK genes were selected based on the expression patterns of a genome-wide dataset of microarrays. The Oryza sativa gamma-ray induced LRR-RLK1 (OsGIRL1) gene was highly induced by gamma irradiation. Therefore, we studied its expression pattern in response to various different abiotic and phytohormone treatments. OsGIRL1 was induced on exposure to abiotic stresses such as salt, osmotic, and heat, salicylic acid (SA), and abscisic acid (ABA), but exhibited downregulation in response to jasmonic acid (JA) treatment. The OsGIRL1 protein was clearly localized at the plasma membrane. The truncated proteins harboring juxtamembrane and kinase domains (or only harboring a kinase domain) exhibited strong autophosphorylation. The biological function of OsGIRL1 was investigated via heterologous overexpression of this gene in Arabidopsis plants subjected to gamma-ray irradiation, salt stress, osmotic stress, and heat stress. A hypersensitive response was observed in response to salt stress and heat stress, whereas a hyposensitive response was observed in response to gamma-ray treatment and osmotic stress. These results provide critical insights into the molecular functions of the rice LRR-RLK genes as receptors of external signals.

Comprehensive Analysis of the Rice RING E3 Ligase Family Reveals Their Functional Diversity in Response to Abiotic stress

A large number of really interesting new gene (RING) E3 ligases contribute to the post-translational modification of target proteins during plant responses to environmental stresses. However, the physical interactome of RING E3 ligases in rice remains largely unknown. Here, we evaluated the expression patterns of 47 Oryza sativa RING finger protein (OsRFP) genes in response to abiotic stresses via semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) and in silico analysis. Subsequently, molecular dissection of nine OsRFPs was performed by the examination of their E3 ubiquitin ligase activity, subcellular localization, and physical interaction with target proteins. Most of the OsRFPs examined possessed E3 ligase activity and showed diverse subcellular localization. Yeast two-hybrid analysis was then employed to construct a physical interaction map of seven OsRFPs with their 120 interacting proteins. The results indicated that these OsRFPs required dynamic translocation and partitioning for their cellular activation. Heterogeneous overexpression of each of the OsRFP genes in Arabidopsis suggested that they have functionally diverse responses to abiotic stresses, which may have been acquired during evolution. This comprehensive study provides insights into the biological functions of OsRFPs, which may be useful in understanding how rice plants adapt to unfavourable environmental conditions.

Differentially expressed transcripts in adipose tissue between Korean native pig and Yorkshire breeds.

We measured and compared the transcripts of adipose tissue from Korean native pig (KNP) and Yorkshire (YS) breeds to investigate breed-specific transcription changes. We employed both the Affymetrix porcine genome array and quantitative reverse transcription-polymerase chain reaction (qRT-PCR). We found eight genes showing significant changes between the two breeds. Based on a literature review, these genes were indicative of differences in extracellular structure density and differences in the potential to metabolize xenobiotic chemicals and lipids. The differentially expressed genes indicated that KNP has a lower extracellular structure density and a lower potential to metabolize xenobiotic chemicals than YS.

Molecular dissection of Oryza sativa salt‐induced RING Finger Protein 1 (OsSIRP1): possible involvement in the sensitivity response to salinity stress

Ubiquitination-mediated protein degradation via Really Interesting New Gene (RING) E3 ligase plays an important role in plant responses to abiotic stress conditions. Many plant studies have found that RING proteins regulate the perception of various abiotic stresses and signal transduction. In this study, Oryza sativa salt-induced RING Finger Protein 1 (OsSIRP1) gene was selected randomly from 44 Oryza sativa RING Finger Proteins (OsRFPs) genes highly expressed in rice roots exposed to salinity stress. Transcript levels of OsSIRP1 in rice leaves after various stress treatments, including salt, heat, drought and hormone abscisic acid (ABA), were observed. Poly-ubiquitinated products of OsSIRP1 were investigated via an in vitro ubiquitination assay.35S:OsSIRP1-EYFP was distributed in the cytosol of untreated and salt-treated rice protoplasts. Heterogeneous overexpression of OsSIRP1 in Arabidopsis reduced tolerance for salinity stress during seed germination and root growth. Our findings indicate that OsSIRP1 acts as a negative regulator of salinity stress tolerance mediated by the ubiquitin 26S proteasome system.

Chloroplast markers for detecting rice grain-derived food ingredients in commercial mixed-flour products

The development of DNA techniques such as quantitative real-time PCR (qRT-PCR) has led to advancements in the field of illegal food product detection. Here, we report a PCR-based method to detect rice grain flour in commercial mixed-flour products. To select the chloroplast genes available for a rice-specific marker, we analyzed chloroplast DNA (cpDNA) polymorphisms in several gene families from five plant species, including rice, adlay, barley, maize, and wheat by using comparative sequence analysis. We found two potential rice-specific marker genes, rpoB and rpoC2, which exhibited relatively high numbers of segregating sites compared to other genes. We designed gene-specific primers for rpoB and rpoC2 on the basis of sequence differences, and identified the appropriate PCR amplification in grain flour samples derived from six Korean rice varieties using the linearity test of the qRT-PCR assay. To test the applicability of these cpDNA markers, we performed a qRT-PCR assay on total DNA obtained from different commercial food products, and successfully detected the rice-specific cpDNA region (rpoB and rpoC2) in several commercial food products that were declared to contain rice. Thus, the reported qRT-PCR assay may prove to be a useful tool for the detection of various rice flours in commercial mixed-flour products such as Sunsik.

Transcriptome Analysis of Flowering Time Genes under Drought Stress in Maize Leaves

Flowering time is an important factor determining yield and seed quality in maize. A change in flowering time is a strategy used to survive abiotic stresses. Among abiotic stresses, drought can increase anthesis-silking intervals (ASI), resulting in negative effects on maize yield. We have analyzed the correlation between flowering time and drought stress using RNA-seq and bioinformatics tools. Our results identified a total of 619 genes and 126 transcripts whose expression was altered by drought stress in the maize B73 leaves under short-day condition. Among drought responsive genes, we also identified 20 genes involved in flowering times. Gene Ontology (GO) enrichment analysis was used to predict the functions of the drought-responsive genes and transcripts. GO categories related to flowering time included reproduction, flower development, pollen–pistil interaction, and post-embryonic development. Transcript levels of several genes that have previously been shown to affect flowering time, such as PRR37, transcription factor HY5, and CONSTANS, were significantly altered by drought conditions. Furthermore, we also identified several drought-responsive transcripts containing C2H2 zinc finger, CCCH, and NAC domains, which are frequently involved in transcriptional regulation and may thus have potential to alter gene expression programs to change maize flowering time. Overall, our results provide a genome-wide analysis of differentially expressed genes (DEGs), novel transcripts, and isoform variants expressed during the reproductive stage of maize plants subjected to drought stress and short-day condition. Further characterization of the drought-responsive transcripts identified in this study has the potential to advance our understanding of the mechanisms that regulate flowering time under drought stress.

Molecular characterization of a heat inducible rice gene, OsHSP1, and implications for rice thermotolerance

Higher plants have acquired complex molecular mechanisms to withstand heat stress through years of natural evolutionary processes. Although physiological responses to elevated temperatures have been well studied, thermotolerance mechanisms at the molecular level are poorly understood in rice plants. In order to identify the genes involved in the thermotolerance of rice, we used a publicly available microarray dataset and identified a number of heat stress-responsive genes. Herein, we report details of the rice gene OsHSP1, which is upregulated by heat stress. In addition, OsHSP1 is highly expressed when exposed to salt and osmotic treatments but not cold treatment. Sequence analysis indicated that OsHSP1 belongs to the heat shock protein 90 family of genes. The biological function of OsHSP1 was investigated by heterologous overexpression in Arabidopsis. Transgenic Arabidopsis overexpressing the OsHSP1 gene exhibited enhanced thermotolerance but was hypersensitive under salt and osmotic stresses. Subcellular localization analysis indicated that the OsHSP1 protein is predominantly targeted to the cytosol and nucleus under heat stress. The coexpression network showed 39 interactions for the functionally interacting genes of OsHSP1. Taken together, these findings suggest that OsHSP1 is a heat-inducible gene that may play an important role in the thermotolerance of rice.

Characterization of Expressed Genes Under Ozone Stress in Soybean

To identify the genes specifically or predominantly expressed in ozone-fumigated leaves of two soybean cultivars: Jinpumkong and Cheongjakong, expression levels of mRNA were investigated using differential banding patterns on agarose gel. A total of 408 bands differently expressed after ozone fumigation was identified; 153 of which were up-regulated while 225 were down-regulated. Using BLASTx, the putative functions of the expressed sequence tags were determined. The 178 ozone-regulated differentially expressed genes (DEGs) matched with the previously known genes with high significance. The putative functional classes of these DEGs were categorized by two databases: Gene Ontology and MIPS. Based on the Gene Ontology database, majority of the DEGS have molecular function related to transferase activity. Most of them are involved in the cellular and metabolic processes. Cytoplasmic part and cell part were the primary types of cellular component in the ozone-responding DEGs. Whereas findings using the MIPS database revealed the function distribution of up-regulated DEGs across all classes. Most of the ozone-regulated genes identified in this study are related to biotic and abiotic stresses. The characterized ESTs will serve as useful data to provide a better understanding of the molecular basis and transcript profiles.

Transcriptome analysis for identifying possible gene regulations during maize root emergence and formation at the initial growth stage.

The root plays an important role during plant development and growth, i.e., the plant body maintenance, nutrient storage, absorption of water, oxygen and nutrient from the soil, and storage of water and carbohydrates, etc. The objective of this study was attempted to determine root-specific genes at the initial developmental stages of maize by using network-based transcriptome analysis. The raw data obtained using RNA-seq were filtered for quality control of the reads with the FASTQC tool, and the filtered reads were pre-proceed using the TRIMMOMATIC tool. The enriched BINs of the DEGs were detected using PageMan analysis with the ORA_FISHER statistical test, and genes were assigned to metabolic pathways by using the MapMan tool, which was also used for detecting transcription factors (TFs). For reconstruction of the co-expression network, we used the algorithm for the reconstruction of accurate cellular networks (ARACNE) in the R package, and then the reconstructed co-expression network was visualized using the Cytoscape tool. RNA-seq. was performed using maize shoots and roots at different developmental stages of root emergence (6–10 days after planting, VE) and 1 week after plant emergence (V2). A total of 1286 differentially expressed genes (DEGs) were detected in both tissues. Many DEGs involved in metabolic pathways exhibited altered mRNA levels between VE and V2. In addition, we observed gene expression changes for 113 transcription factors and found five enriched cis-regulatory elements in the 1-kb upstream regions of both DEGs. The network-based transcriptome analysis showed two modules as co-expressed gene clusters differentially expressed between the shoots and roots during plant development. The DEGs of one module exhibited gene expressional coherence in the maize root tips, suggesting that their functional relationships are associated with the initial developmental stage of the maize root. Finally, we confirmed reliable mRNA levels of the hub genes in the potential sub-network related to initial root development at the different developmental stages of VE, V2, and 2 weeks after plant emergence.