• Biomedical Science Letters
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• v.18 no.3
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• pp.310-312
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• 2012

Cytochrome $b_5$ is involved in the reduction of methemoglobin back to hemoglobin, thereby maintaining normal function of the blood to carry oxygen around. Congenital abnormal condition of this enzyme causes a rare disease called methemoglobinemia. At least 4 different retropseudogenes are reported so far for cytochrome $b_5$. However, type III pseudogene has attracted most attention because it contains open reading frame in its structure. Although there is no evidence yet if this pseudogene is actually expressed in the cell or the blood the possibility of its expression needs to be elucidated. We have isolated type III pseudogene by polymerase chain reaction and cloned into pGEX-4T-1 expression vector followed by SDS-PAGE. Protein was expressed and the size of the expressed protein was 28 kDa as expected in its genetic code. This result also shows that the protein is not harmful for the viability of the microorganism. This study may contribute to the genetic diagnosis of cardiac diseases, possibly caused by cytochrome $b_5$.

• YAKHAK HOEJI
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• v.53 no.3
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• pp.156-160
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• 2009

The present study was undertaken to determine whether transgelin participates in the regulation of vascular smooth muscle contraction and, if so, to investigate the mechanism. By PCR homology cloning, the cDNA sequence of ferret transgelin was determined and phosphorothioate antisense and random oligonucleotides were synthesized and introduced into strips of ferret aorta by a chemical loading procedure. Treatment of ferret aorta with transgelin antisense oligonucleotides resulted in a significant decrease in protein levels of transgelin to sham- or random sequence-loaded muscles, but no change in the protein levels of actin. Contraction in response to a phorbol ester was significantly decreased in antisense-treated muscles compared to sham- or random sequence-loaded controls. Neither basal intrinsic tone nor the contraction in response to phenylephrine was significantly affected by the antisense treatment. The data indicate that transgelin plays a significant role in the regulation of contraction and suggest that in a tonically active smooth muscle transgelin may function as a signalling protein to facilitate PKC or ERK-dependent signalling rather than thick filament regulation including $Ca^{2+}$ or calmodulin dependent regulation of myosin light chain kinase.

• BMB Reports
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• v.50 no.5
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• pp.235-236
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• 2017

The circadian clock is an internal system that is synchronized by external stimuli, such as light and temperature, and influences various physiological and developmental processes in living organisms. In the model plant Arabidopsis, transcriptional, translational and post-translational processes are interlocked by feedback loops among morning- and evening-phased genes. In a post-translational loop, plant-specific single-gene encoded GIGANTEA (GI) stabilize the F-box protein ZEITLUPE (ZTL), driving the targeted-proteasomal degradation of TIMING OF CAB EXPRESSION 1 (TOC1) and PSEUDO-RESPONSE REGULATOR 5 (PRR5). Inherent to this, we demonstrate the novel biochemical function of GI as a chaperone and/or co-chaperone of Heat-Shock Protein 90 (HSP90). GI prevents ZTL degradation as a chaperone and facilitates ZTL maturation together with HSP90/HSP70, enhancing ZTL activity in vitro and in planta. GI is known to be involved in a wide range of physiology and development as well as abiotic stress responses in plants, but it could also interact with diverse client proteins to increase protein maturation. Our results provide evidence that GI helps proteostasis of ZTL by acting as a chaperone and a co-chaperone of HSP90 for proper functioning of the Arabidopsis circadian clock.

• IMMUNE NETWORK
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• v.2 no.2
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• pp.72-78
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• 2002

Background: The precise mechanism by which CTLA-4 regulates T cell immune responses is still not fully understood. Previously we proposed that CTLA-4 could downregulate T cell function by modulating a signaling cascade initiated from the T cell receptor complex. The evidence for this notion comes from our findings that CTLA-4 associated with the T cell receptor zeta (TCR zeta) chain, and hence regulated TCR zeta phosphorylation by co-associated SHP-2 tyrosine phosphatase (1). In this report, we investigated whether any other signaling molecules could be involved in the CTLA-4 signaling pathway. Methods: We have taken biochemical approaches, such as immunoprecipitation followed by autoradiography or immunoblotting, to identify the molecules associated with CTLA-4. To perform these assays, we used activated primary T cells and ectopically transfected 293 cells. Various truncation mutants of CTLA-4 were used to map the interaction site on CTLA-4. Results: We found that in addition to TCR zeta and SHP-2, a recently cloned small adaptor molecule, SAP (SLAM-associated protein), was also able to associate with CTLA-4. We identified the domain of SAP association in CTLA-4 being a motif involving GVYVKM. This motif has been previously found to bind SHP-2 through its phosphorylated tyrosine interaction with SH-2 domain of SHP-2. Indeed, co-expression of SAP and SHP-2 reduced their binding to CTLA-4 significantly, suggesting that SAP and SHP-2 compete for the common binding site, GVYVKM. Thus, by blocking SHP-2 recruitment SAP could function as a negative regulator of CTLA-4. Conclusion: Taken together, our data suggest the existence of complicate signaling cascade in regulating CTLA-4 function, and further provide evidence that SAP can act either as a positive or negative regulator depending on the nature of the associating receptors.

• Journal of Microbiology and Biotechnology
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• v.17 no.9
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• pp.1452-1459
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• 2007

The Dps protein, which is overexpressed in harsh environments, is known to playa critical role in the protection of DNA against oxidative stresses. In this study, the roles of Fur in the expression of the dps gene in Salmonella and the protection mechanisms against oxidative stress in Salmonella cells preexposed to iron-stress were investigated. Two putative Fur boxes were predicted within the promoter region of the S. typhimurium dps gene. The profile of dps expression performed by the LacZ reporter assay revealed growth-phase dependency regardless of iron-status under the culture conditions. The fur mutant, $_X4659$, evidenced a reduced level of ${\beta}$-galactosidase as compared to the wild-type strain. The results observed after the measurement of the Dps protein in various Salmonella regulatory mutants were consistent with the results acquired in the reporter assay. This evidence suggested that Fur performs a function as a subsidiary regulator in the expression of dps. The survival ability of Salmonella strains after exposure to oxidative stress demonstrated that the Dps protein performs a pivotal function in the survival of stationary-phase S. typhimurium against oxidative stress. Salmonella cells grown in iron-restricted condition required Dps for full protection against oxidative stress. The CK24 (${\Delta}dps$) cells grown in iron-replete condition survived at a rate similar to that observed in the wild-type strain, thereby suggesting the induction of an unknown protection mechanism(s) other than Dps in this condition.

• Biomolecules & Therapeutics
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• v.29 no.5
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• pp.506-518
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• 2021

The imprinted tumour suppressor NOEY2 is downregulated in various cancer types, including ovarian cancers. Recent data suggest that NOEY2 plays an essential role in regulating the cell cycle, angiogenesis and autophagy in tumorigenesis. However, its detailed molecular function and mechanisms in ovarian tumours remain unclear. In this report, we initially demonstrated the inhibitory effect of NOEY2 on tumour growth by utilising a xenograft tumour model. NOEY2 attenuated the cell growth approximately fourfold and significantly reduced tumour vascularity. NOEY2 inhibited the phosphorylation of the signalling components downstream of phosphatidylinositol-3'-kinase (PI3K), including phosphoinositide-dependent protein kinase 1 (PDK-1), tuberous sclerosis complex 2 (TSC-2) and p70 ribosomal protein S6 kinase (p70S6K), during ovarian tumour progression via direct binding to vascular endothelial growth factor receptor-2 (VEGFR-2). Particularly, the N-terminal domain of NOEY2 (NOEY2-N) had a potent anti-angiogenic activity and dramatically downregulated VEGF and hypoxia-inducible factor-1α (HIF-1α), key regulators of angiogenesis. Since no X-ray or nuclear magnetic resonance structures is available for NOEY2, we constructed the three-dimensional structure of this protein via molecular modelling methods, such as homology modelling and molecular dynamic simulations. Thereby, Lys15 and Arg16 appeared as key residues in the N-terminal domain. We also found that NOEY2-N acts as a potent inhibitor of tumorigenesis and angiogenesis. These findings provide convincing evidence that NOEY2-N regulates endothelial cell function and angiogenesis by interrupting the VEGFR-2/PDK-1/GSK-3β signal transduction and thus strongly suggest that NOEY2-N might serve as a novel anti-tumour and anti-angiogenic agent against many diseases, including ovarian cancer.

• Korean Journal of Plant Tissue Culture
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• v.21 no.5
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• pp.253-275
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• 1994

Many flowering plants possess genetically controlled self -incompatibility (SI) system that prevents inbreeding and promotes outcrosses. SI is usually controlled by a single, multiallelic S-locus. In gametophytically controlled system, SI results when the S-allele of the pollen is matched by one of the two S-alleles in the style, while in the sporophytic system self-incompatible reaction occurs by the interaction between the pistil genotype and genotype of, not the pollen, but the pollen parent In the former system the self-incompatible phenotype of pollen is determined by the haploid genome of the pollen itself but in the latter the pollen phenotype is governed by the genotype of the pollen parent along with the occurrence of either to-dominant or dominant/recessive allelic interactions. In the sporophytic type the inhibition reaction occurs within minutes following pollen-stigma contact, the incompatible pollen grains usually failing to germinate, whereas in gametophytic system pollen tube inhibition takes place during growth in the transmitting tissue of the style. Recognition and rejection of self pollen are the result of interaction between the S-locus protein in the pistil and the pollen protein. In the gametophytic SI the S-associated glycoprotein which is similar to the fungal ribonuclease in structure and function are localized at the intercellular matrix in the transmitting tissue of the style, with the highest concentration in the collar of the stigma, while in the sporophytic SI deposit of abundant S-locus specific glycoprotein (SLSG).is detected in the cell wall of stigmatic papillae of the open flowers. In the gametophytic system S-gene is expressed mostly at the stigmatic collar the upper third of the style length and in the pollen after meiosis. On the other hand, in the sporophytic SI S-glycoprotein gene is expressed in the papillar cells of the stigma as well as in e sporophytic tape is cells of anther wall. Recognition and rejection of self pollen in the gametophytic type is the reaction between the ribonuclease in the transmitting tissue of the style and the protein in the cytoplasm of pollen tube, whereas in the sporophytic system the inhibition of selfed pollen is caused by the interaction between the Sycoprotein in the wall of stigmatic papillar cell and the tapetum-origin protein deposited on the outer wall of the pollen grain. The claim that the S-allele-associated proteins are involved in recognition and rejection of self pollen has been made merely based on indirect evidence. Recently it has been verified that inhibition of synthesis of S$_3$ protein in Petunia inflata plants of S$_2$S$_3$ genotype by the antisense S$_3$ gene resulted in failure of the transgenic plant to reject S$_3$ pollen and that expression of the transgenic encoding S$_3$ protein in the S$_1$S$_2$ genotype confers on the transgenic plant the ability to reject S$_3$ pollen. These finding Provide direct evidence that S-proteins control the s elf-incompatibility behavior of the pistil.

• Childhood Kidney Diseases
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• v.19 no.2
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• pp.65-70
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• 2015

It is well known that proteins present in the primary urine are reabsorbed in the renal proximal tubules, and that this reabsorption is mediated via the megalin-cubilin complex and the neonatal $Fc{\gamma}$ receptor. However, the reabsorption is also thought to be influenced by an electrostatic interaction between protein molecules and the microvilli of the renal proximal tubules. By analyzing the charge diversity of urinary IgG, we showed that this reabsorption process occurs in a cationic charge-preferential manner. The charge-selective molecular sieving function of the glomerular capillary walls has long been a target of research since Brenner et al. demonstrated the existence of this function by a differential clearance study by using the anionic dextran sulfate polymer. However, conclusive evidence was not obtained when the study was performed using differential clearance of serum proteins. We noted that immunoglobulin (Ig) A and IgG have similar molecular sizes but distinct molecular isoelectric points. Therefore, we studied the differential clearance of these serum proteins (clearance IgA/clearance IgG) in podocyte diseases and glomerulonephritis. In addition, we studied this differential clearance in patients with Dent disease rather than in normal subjects because the glomerular sieving function is considered to be normal in subjects with Dent disease. Our results clearly showed that the charge-selective barrier is operational in Dent disease, impaired in podocyte disease, and lacking in glomerulonephritis.

• BMB Reports
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• v.53 no.2
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• pp.100-105
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• 2020

While liver histopathology is heterogeneous in diabetes, the underlying mechanisms remain unclear. We investigated whether glycemic variation resulting from differential diets can induce heterogeneity in diabetic liver and the underlying molecular mechanisms. We generated end-stage non-obese diabetic model rats by subtotal-pancreatectomy in male Sprague-Dawley rats and ad libitum diet for 7 weeks (n = 33). The rats were then divided into three groups, and fed a standard- or a low-protein diet (18 or 6 kcal%, respectively), for another 7 weeks: to maintain hyperglycemia, 11 rats were fed ad libitum (18AL group); to achieve euglycemia, 11 were calorie-restricted (18R group), and 11 were both calorie- and protein-restricted with the low-protein diet (6R group). Overnight-fasted liver samples were collected after the differential diets together with sham-control (18S group), and histology and molecular changes were compared. Hyperglycemic-18AL showed glycogenic hepatopathy (GH) without steatosis, with the highest GSK-3β inactivation because of Akt activation during hyperglycemia; mitochondrial function was not impaired, compared to the 18S group. Euglycemic-18R showed neither GH nor steatosis, with intermediate GSK-3β activation and mitochondrial dysfunction. However, euglycemic-6R showed both GH and steatosis despite the highest GSK-3β activity and no molecular evidence of increased lipogenesis or decreased ApoB expression, where mitochondrial dysfunction was highest among the groups. In conclusion, heterogeneous liver histopathology developed in end-stage non-obese diabetic rats as the glycemic levels varied with differential diets, in which protein content in the diets as well as glycemic levels differentially influenced GSK-3β activity and mitochondrial function in insulin-deficient state.

• KSBB Journal
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• v.22 no.5
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• pp.341-344
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• 2007

Obesity is a major obstacle for human health, which induces various diseases such as cardiac injury and type 2 diabetes. Ginsenosides, active components of ginseng extract, exert various physiological effects. However, There are still no evidence for their anti obesity effects. In this study, we investigated the effects of ginsenoside Rd on adipocyte differentiation in 3T3-L1 cells. Our data show that ginsenoside Rd (80 uM) was effective in adipocyte differentiation inhibition. These inhibitory effects of ginsenosides on adipocyte differentiation were accompanied by PPAR gamma inhibition in rosiglitazone-treated cells. We also tested whether AMP-activated protein kinase (AMPK) activation was involved in the effects of these ginsenosides. AMPK is a master target for obesity, ginsenoside Rd significantly activated AMPK. Taken together, these results suggest that the anti obesity effects of ginsenoside Rd involve the AMPK signaling pathway and PPAR-gamma inhibition.