• BMB Reports
• /
• v.39 no.6
• /
• pp.749-758
• /
• 2006

The cytosolic members of the HSP70 family of proteins play key roles in the molecular chaperone machinery of the cell. In the study we cloned and sequenced the full-length cDNA of Delia antiqua HSP70 gene, which is 2461 bp long and encodes 643 a.a. with a calculated molecular mass of 70,787 Da. We investigated gene copies of cytosolic HSP70 members of 4 insect species with complete genome available, and found that they are quite variable with species. In order to characterize this protein we carried out an alignment and a phylogenetic analysis with 41 complete protein sequences from insects. The analysis divided the cytosolic members of the family into two classes, HSP70 and HSC70, distinguishable on the basis of 15 residues. HSP70 class members were slightly shorter in length and smaller in molecular mass relative to the HSC70 class members, and the conservative and functional regions in these sequences were documented. Mainly, we investigated the expression of Delia antiqua HSP70 gene, in response to diapauses and thermal stresses. Both summer and winter diapauses elevated HSP70 transcript levels. Cold-stress led to increased HSP70 expression levels in summer- and winter-diapausing pupae, but heat-stress elevated the levels only in the winter-diapausing pupae. In all cases, the expression levels, after being elevated, gradually decreased with time. HSP70 expression was low in non-diapausing pupae but was up-regulated following cold- and heat-stresses. Heat-stress gradually increased the mRNA level with time whereas cold-stress gradually decreased levels after an initial increase.

• Journal of Microbiology and Biotechnology
• /
• v.26 no.12
• /
• pp.2106-2115
• /
• 2016

To identify the global effects of (p)ppGpp in the gram-positive bacterium Deinococcus radiodurans, which exhibits remarkable resistance to radiation and other stresses, RelA/SpoT homolog (RSHs) mutants were constructed by direct deletion mutagenesis. The results showed that RelA has both synthesis and hydrolysis domains of (p)ppGpp, whereas RelQ only synthesizes (p)ppGpp in D. radiodurans. The growth assay for mutants and complementation analysis revealed that deletion of relA and relQ sensitized the cells to $H_2O_2$, heat shock, and amino acid limitation. Comparative proteomic analysis revealed that the bifunctional RelA is involved in DNA repair, molecular chaperone functions, transcription, the tricarboxylic acid cycle, and metabolism, suggesting that relA maintains the cellular (p)ppGpp levels and plays a crucial role in oxidative resistance in D. radiodurans. The D. radiodurans relA and relQ genes are responsible for (p)ppGpp synthesis/hydrolysis and (p)ppGpp hydrolysis, respectively. (p)ppGpp integrates a general stress response with a targeted re-programming of gene regulation to allow bacteria to respond appropriately towards heat shock, oxidative stress, and starvation. This is the first identification of RelA and RelQ involvement in response to oxidative, heat shock, and starvation stresses in D. radiodurans, which further elucidates the remarkable resistance of this bacterium to stresses.

• Journal of Microbiology and Biotechnology
• /
• v.9 no.3
• /
• pp.346-351
• /
• 1999

Tumor cells may alter the expression of proteins involved in antigen processing and presentation, allowing them to avoid recognition and elimination by cytotoxic T cells. In order to investigate whether the major histocompatibility complex (MHC) class I-mediated antigen processing machinery is preserved in human lung cancer cell lines, we examined the expression of multiple components of the MHC class I antigen processing pathway, including transporter associated with antigen processing (TAP), $\beta_2$-microglobulin, MHC class I molecules, and chaperones which have not been previously examined in this context. Row cytometry analysis showed that the cell surface expression of MHC class I molecules was downregulated in all of the cell lines. While some cell lines showed no detectable expression of MHC class I molecules, pulse-chase experiments showed that MHC class I molecules were synthesized in the other cell lines but not transported from the endoplasmic reticulum to the cell surface. Low or nondetectable levels of TAP1 and/or TAP2 expression were demonstrated by Western blot analysis in all of the cell lines, representing a variety of lung tissue types. In some cases, this was accompanied by loss of tapasin expression. Our findings suggest that downregulation of antigen processing may be one of the strategies used by tumors to escape immune surveillance. This study provides further information for designing the potential therapeutic applications such as immunotherapy and gene therapy against cancers.

• The Plant Pathology Journal
• /
• v.36 no.1
• /
• pp.43-53
• /
• 2020

Ralstonia solanacearum (Rso) is a causal agent of bacterial wilt in Solanaceae crops worldwide including Republic of Korea. Rso virulence predominantly relies on type III secreted effectors (T3Es). However, only a handful of Rso T3Es have been characterized. In this study, we investigated subcellular localization of and manipulation of plant immunity by 8 Rso T3Es predicted to harbor a nuclear localization signal (NLS). While 2 of these T3Es elicited cell death in both Nicotiana benthamiana and N. tabacum, only one was dependent on suppressor of G2 allele of skp1 (SGT1), a molecular chaperone of nucleotide-binding and leucine-rich repeat immune receptors. We also identified T3Es that differentially regulate flg22-induced reactive oxygen species production and gene expression. Interestingly, several of the NLS-containing T3Es translationally fused with yellow fluorescent protein accumulated in subcellular compartments other than the cell nucleus. Our findings bring new clues to decipher Rso T3E function in planta.

• Journal of Environmental Impact Assessment
• /
• v.26 no.3
• /
• pp.207-216
• /
• 2017

Several bioassays have been performed for assessment of the impact of polluted sediments. The direct exposure method using sediments is limited by difficulty controlling feeding and its effects on organisms. Furthermore, only macro-organisms and benthic organisms are used. To evaluate the potential application of sediment elutriate as a complementary strategy for impact assessment, copepods, small organisms with a short life cycle, were exposed to sediment elutriates, and several end-points were measured. As a result, sediment elutriates prepared from polluted sites caused growth retardation in marine copepods. In terms of molecular biomarkers, antioxidant-related and chaperone protein gene expression levels were increased in a dose- and time-dependent manner. Thus, we suggest that sediment elutriate tests can provide an effective alternative for toxicity assessment using whole sediment samples. Further studies are required to obtain sufficient data for future applications.

• YAKHAK HOEJI
• /
• v.54 no.3
• /
• pp.180-186
• /
• 2010

Aldosterone, mineralocorticoid hormone, has important functions related to the regulation of blood pressure and balance of fluids and electrolytes in the distal region of the nephron. By genomic and non-genomic action of aldosterone, the physiological kidney functions are modulated. However, many of them except several kind of sodium channel have not been identified and analyzed yet. In this study, proteomic technologies with two-dimensional gel electrophoresis (2-DE) gel using aldosterone rat model were applied to analyze and identify the aldosterone dependently expressed proteins in rat kidney cortex. As a result, the established aldosterone rat model exhibited the normal physiological responses to aldosterone and modulated proteins were identified, which included 15 increased and 3 decreased proteins on 2-DE analysis. Among them, 11 proteins were identified as changed proteins by LC-MS/MS analysis. These proteins identified as aldosterone induced proteins were involved in several cellular pathways such as cytoskeleton remodeling, energy metabolism, amino acid metabolism, and chaperone process. In conclusion, our data could provide the insights into the new mechanism underlying regulation of kidney functions by aldosterone.

• Journal of Microbiology and Biotechnology
• /
• v.27 no.7
• /
• pp.1345-1358
• /
• 2017

The impact of overproduction of a heterologous protein on the metabolic system of host Lactococcus lactis was investigated. The protein expression profiles of L. lactis IL1403 containing two near-identical plasmids that expressed high- and low-level of the green fluorescent protein (GFP) were examined via shotgun proteomics. Analysis of the two strains via high-throughput LC-MS/MS proteomics identified the expression of 294 proteins. The relative amount of each protein in the proteome of both strains was determined by label-free quantification using the spectral counting method. Although expression level of most proteins were similar, several significant alterations in metabolic network were identified in the high GFP-producing strain. These changes include alterations in the pyruvate fermentation pathway, oxidative pentose phosphate pathway, and de novo synthesis pathway for pyrimidine RNA. Expression of enzymes for the synthesis of dTDP-rhamnose and N-acetylglucosamine from glucose was suppressed in the high GFP strain. In addition, enzymes involved in the amino acid synthesis or interconversion pathway were downregulated. The most noticeable changes in the high GFP-producing strain were a 3.4-fold increase in the expression of stress response and chaperone proteins and increase of caseinolytic peptidase family proteins. Characterization of these host expression changes witnessed during overexpression of GFP was might suggested the metabolic requirements and networks that may limit protein expression, and will aid in the future development of lactococcal hosts to produce more heterologous protein.

• Bulletin of the Korean Chemical Society
• /
• v.23 no.12
• /
• pp.1773-1777
• /
• 2002

Reactive oxygen species(ROS) have been investigated to have pivotal roles on amyloidogenecity of $\beta-amyloidpeptide(A\beta)$, the major component of senile plaques in Alzheimer's disease(AD) brain. Addition of radical scavengers is one of the on-going strategies for therapeutic treatment for AD patients. Hsp104 protein including two ATP binding sites from Saccharomyces cerevisiae, as a molecular chaperone, was known to function as a protector of ROS generation when exposed to oxidative stress in our previous study. This observation has led us to investigate Hsp104 protein as a molecular mediator of $A{\beta}$ aggregation in this study. We have developed a new way of expression for Hsp104 protein using GST-fusion tag. As we expected, formation of $A{\beta}$ aggregate was protected by wild type Hsp104 protein, but not by the two ATP-binding site mutant, based on Thioflavin-T fluorescence. Interestingly, Hsp104 protein was observed to keep $A{\beta}$ from forming aggregates independent of ATP binding. On the other hand, disaggregation of $A{\beta}$ aggregates by wild type Hsp104 was totally dependent on the presence of ATP. On the other hand, mutant Hsp104 with two ATP binding sites altered exhibited no inhibition. Another effective antioxidant, hydrazine analogs of curcumin were also effective in $A{\beta}$ fibrilization as protectors against oxidative stress. Based on these observations we conclude that Hsp104 and curcumin derivatives, as protectors of oxidative stress, inhibit $A{\beta}$ aggregation in virto and can be candidates for therapeutic approaches in cure of some neurodegenerative disease.

• BMB Reports
• /
• v.40 no.1
• /
• pp.72-81
• /
• 2007

In order to screen the aging related proteins in human normal colon epithelia, the comparative proteomics analysis was applied to get the two-dimensional electrophoresis (2-DE) profiles with high resolution and reproducibility from normal colon epithelial tissues of young and aged people. Differential proteins between the colon epithelia of two age groups were found with PDQuest software. The thirty five differential protein-spots were identified by peptide mass fingerprint (PMF) based on matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) and database searching. Among them there are sixteen proteins which are significantly up-regulated in the colonic mucosal epithelia of young people group, which include ATP synthase beta chain, electron transfer flavoprotein alpha-subunit, catalase, glutathione peroxidase 1, annexin A2 and heat shock cognate 71 kDa protein, etc.; There are nineteen proteins which are significantly up-regulated in the colonic mucosal epithelia of aged people group, which include far upstream element-binding protein 1, nucleoside diphosphate kinase B, protein disulfide-isomerase precursor and VDAC-2, etc.. The identified differential proteins appear to be involved in metabolism, energy generation, chaperone, antioxidation, signal transduction, protein folding and apoptosis. The data will help to understand the molecular mechanisms of human colon epithelial aging.

• Journal of Ginseng Research
• /
• v.44 no.1
• /
• pp.161-167
• /
• 2020

Background: The ascomycete fungi Cylindrocarpon destructans (Cd) and Fusarium solani (Fs) cause ginseng root rot and significantly reduce the quality and yield of ginseng. Cd produces the secondary metabolite radicicol, which targets the molecular chaperone Hsp90. Fs is resistant to radicicol, whereas other fungal genera associated with ginseng disease are sensitive to it. Radicicol resistance mechanisms have not yet been elucidated. Methods: Transcriptome analyses of Fs and Cd mycelia treated with or without radicicol were conducted using RNA-seq. All of the differentially expressed genes (DEGs) were functionally annotated using the Fusarium graminearum transcript database. In addition, deletions of two transporter genes identified by RNA-seq were created to confirm their contributions to radicicol resistance. Results: Treatment with radicicol resulted in upregulation of chitin synthase and cell wall integrity genes in Fs and upregulation of nicotinamide adenine dinucleotide dehydrogenase and sugar transporter genes in Cd. Genes encoding an ATP-binding cassette transporter, an aflatoxin efflux pump, ammonium permease 1 (mep1), and nitrilase were differentially expressed in both Fs and Cd. Among these four genes, only the ABC transporter was upregulated in both Fs and Cd. The aflatoxin efflux pump and mep1 were upregulated in Cd, but downregulated in Fs, whereas nitrilase was downregulated in both Fs and Cd. Conclusion: The transcriptome analyses suggested radicicol resistance pathways, and deletions of the transporter genes indicated that they contribute to radicicol resistance.