Ethanol-Induced Alcohol Dehydrogenase E (AdhE) Potentiates Pneumolysin in <i>Streptococcus pneumoniae</i>

Luong, Truc Thanh,Kim, Eun-Hye,Bak, Jong Phil,Nguyen, Cuong Thach,Choi, Sangdun,Briles, David E.,Pyo, Suhkneung,Rhee, Dong-Kwon American Society for Microbiology 2015 Infection and immunity Vol.83 No.1

<P>Alcohol impairs the host immune system, rendering the host more vulnerable to infection. Therefore, alcoholics are at increased risk of acquiring serious bacterial infections caused by <I>Streptococcus pneumoniae</I>, including pneumonia. Nevertheless, how alcohol affects pneumococcal virulence remains unclear. Here, we showed that the <I>S. pneumoniae</I> type 2 D39 strain is ethanol tolerant and that alcohol upregulates alcohol dehydrogenase E (AdhE) and potentiates pneumolysin (Ply). Hemolytic activity, colonization, and virulence of <I>S. pneumoniae</I>, as well as host cell myeloperoxidase activity, proinflammatory cytokine secretion, and inflammation, were significantly attenuated in <I>adhE</I> mutant bacteria (Δ<I>adhE</I> strain) compared to D39 wild-type bacteria. Therefore, AdhE might act as a pneumococcal virulence factor. Moreover, in the presence of ethanol, <I>S. pneumoniae</I> AdhE produced acetaldehyde and NADH, which subsequently led Rex (redox-sensing transcriptional repressor) to dissociate from the <I>adhE</I> promoter. An increase in AdhE level under the ethanol condition conferred an increase in Ply and H<SUB>2</SUB>O<SUB>2</SUB> levels. Consistently, <I>S. pneumoniae</I> D39 caused higher cytotoxicity to RAW 264.7 cells than the Δ<I>adhE</I> strain under the ethanol stress condition, and ethanol-fed mice (alcoholic mice) were more susceptible to infection with the D39 wild-type bacteria than with the Δ<I>adhE</I> strain. Taken together, these data indicate that AdhE increases Ply under the ethanol stress condition, thus potentiating pneumococcal virulence.</P>

Inhibition of Autolysis by Lipase LipA in Streptococcus pneumoniae Sepsis

김규리,Truc Thanh Luong,박상상,이승엽,하정아,Cuong Thach Nguyen,안지혜,박기태,백만정,표석능,David E. Briles,이동권 한국분자세포생물학회 2017 Molecules and cells Vol.40 No.12

More than 50% of sepsis cases are associated with pneumonia. Sepsis is caused by infiltration of bacteria into the blood via inflammation, which is triggered by the release of cell wall components following lysis. However, the regulatory mechanism of lysis during infection is not well defined. Mice were infected with Streptococcus pneumoniae D39 wild-type (WT) and lipase mutant (∆lipA) intranasally (pneumonia model) or intraperitoneally (sepsis model), and survival rate and pneumococcal colonization were determined. LipA and autolysin (LytA) levels were determined by qPCR and western blotting. S. pneumoniae Spd_1447 in the D39 (type 2) strain was identified as a lipase (LipA). In the sepsis model, but not in the pneumonia model, mice infected with the ∆lipA displayed higher mortality rates than did the D39 WT-infected mice. Treatment of pneumococci with serum induced LipA expres-sion at both the mRNA and protein levels. In the presence of serum, the ∆lipA displayed faster lysis rates and higher LytA expression than the WT, both in vitro and in vivo. These results indicate that a pneumococcal lipase (LipA) represses autolysis via inhibition of LytA in a sepsis model.

Effect of decreased BCAA synthesis through disruption of ilvC gene on the virulence of Streptococcus pneumoniae

김규리,이승엽,Truc Thanh Luong,Cuong Thach Nguyen,박상상,Suhk neung Pyo,이동권 대한약학회 2017 Archives of Pharmacal Research Vol.40 No.8

Streptococcus pneumoniae (pneumococcus) isresponsible for significant morbidity and mortality worldwide. It causes a variety of life-threatening infections suchas pneumonia, bacteremia, and meningitis. In bacterialphysiology, the metabolic pathway of branched-chainamino acids (BCAAs) plays an important role in virulence. Nonetheless, the function of IlvC, one of the enzymesinvolved in the biosynthesis of BCAAs, in S. pneumoniaeremains unclear. Here, we demonstrated that downregulationof BCAA biosynthesis by ilvC ablation can diminishBCAA concentration and expression of pneumolysin (Ply)and LytA, and subsequently attenuate virulence. Infectionwith an ilvC mutant showed significantly reduced mortalityand colonization in comparison with strain D39 (serotype2, wild type), suggesting that ilvC can potentiate S. pneumoniaevirulence due to adequate BCAA synthesis. Takentogether, these results suggest that the function of ilvC inBCAA synthesis is essential for virulence factor and couldplay an important role in the pathogenesis of respiratoryinfections.

Adenylate kinase potentiates the capsular polysaccharide by modulating Cps2D in Streptococcus pneumoniae D39

Prachetash Ghosh,Truc Thanh Luong,Masaud Shah,Trung Thanh Thach,최상돈,이상호,이동권 생화학분자생물학회 2018 Experimental and molecular medicine Vol.50 No.-

Streptococcus pneumoniae is a polysaccharide-encapsulated bacterium. The capsule thickens during blood invasion compared with the thinner capsules observed in asymptomatic nasopharyngeal colonization. However, the underlying mechanism regulating differential CPS expression remains unclear. CPS synthesis requires energy that is supplied by ATP. Previously, we demonstrated a correlation between ATP levels and adenylate kinase in S. pneumoniae (SpAdK). A dose-dependent induction of SpAdK in serum was also reported. To meet medical needs, this study aimed to elucidate the role of SpAdK in the regulation of CPS production. CPS levels in S. pneumoniae type 2 (D39) increased proportionally with SpAdK levels, but they were not related to pneumococcal autolysis. Moreover, increased SpAdK levels resulted in increased total tyrosine kinase Cps2D levels and phosphorylated Cps2D, which is a regulator of CPS synthesis in the D39 strain. Our results also indicated that the SpAdK and Cps2D proteins interact in the presence of Mg-ATP. In addition, in silico analysis uncovered the mechanism behind this protein–protein interaction, suggesting that SpAdK binds with the Cps2D dimer. This established the importance of the ATP-binding domain of Cps2D. Taken together, the biophysical interaction between SpAdK and Cps2D plays an important role in enhancing Cps2D phosphorylation, which results in increased CPS synthesis.

Korean Red Ginseng inhibits apoptosis in neuroblastoma cells via estrogen receptor ${\beta}$-mediated phosphatidylinositol-3 kinase/Akt signaling

Nguyen, Cuong Thach,Luong, Truc Thanh,Kim, Gyu-Lee,Pyo, Suhkneung,Rhee, Dong-Kwon The Korean Society of Ginseng 2015 Journal of Ginseng Research Vol.39 No.1

Background: Ginseng has been shown to exert antistress effects both in vitro and in vivo. However, the effects of ginseng on stress in brain cells are not well understood. This study investigated how Korean Red Ginseng (KRG) controls hydrogen peroxide-induced apoptosis via regulation of phosphatidylinositol-3 kinase (PI3K)/Akt and estrogen receptor (ER)-${\beta}$ signaling. Methods: Human neuroblastoma SK-N-SH cells were pretreated with KRG and subsequently exposed to $H_2O_2$. The ability of KRG to inhibit oxidative stress-induced apoptosis was assessed in MTT cytotoxicity assays. Apoptotic protein expression was examined byWestern blot analysis. The roles of ER-${\beta}$, PI3K, and p-Akt signaling in KRG regulation of apoptosis were studied using small interfering RNAs and/or target antagonists. Results: Pretreating SK-N-SH cells with KRG decreased expression of the proapoptotic proteins p-p53 and caspase-3, but increased expression of the antiapoptotic protein BCL2. KRG pretreatment was also associated with increased ER-${\beta}$, PI3K, and p-Akt expression. Conversely, ER-${\beta}$ inhibition with small interfering RNA or inhibitor treatment increased p-p53 and caspase-3 levels, but decreased BCL2, PI3K, and p-Akt expression. Moreover, inhibition of PI3K/Akt signaling diminished p-p53 and caspase-3 levels, but increased BCL2 expression. Conclusion: Collectively, the data indicate that KRG represses oxidative stress-induced apoptosis by enhancing PI3K/Akt signaling via upregulation of ER-${\beta}$ expression.

Korean Red Ginseng inhibits apoptosis in neuroblastoma cells via estrogen receptor β-mediated phosphatidylinositol-3 kinase/Akt signaling

Cuong Thach Nguyen,Truc Thanh Luong,Gyu-Lee Kim,Suhkneung Pyo,Dong-Kwon Rhee 고려인삼학회 2015 Journal of Ginseng Research Vol.39 No.3

Background: Ginseng has been shown to exert antistress effects both in vitro and in vivo. However, the effects of ginseng on stress in brain cells are not well understood. This study investigated how Korean Red Ginseng (KRG) controls hydrogen peroxide-induced apoptosis via regulation of phosphatidylinositol- 3 kinase (PI3K)/Akt and estrogen receptor (ER)-β signaling. Methods: Human neuroblastoma SK-N-SH cells were pretreated with KRG and subsequently exposed to H₂O₂. The ability of KRG to inhibit oxidative stress-induced apoptosis was assessed in MTT cytotoxicity assays. Apoptotic protein expression was examined byWestern blot analysis. The roles of ER-β, PI3K, and p-Akt signaling in KRG regulation of apoptosis were studied using small interfering RNAs and/or target antagonists. Results: Pretreating SK-N-SH cells with KRG decreased expression of the proapoptotic proteins p-p53 and caspase-3, but increased expression of the antiapoptotic protein BCL2. KRG pretreatment was also associated with increased ER-β, PI3K, and p-Akt expression. Conversely, ER-β inhibition with small interfering RNA or inhibitor treatment increased p-p53 and caspase-3 levels, but decreased BCL2, PI3K, and p-Akt expression. Moreover, inhibition of PI3K/Akt signaling diminished p-p53 and caspase-3 levels, but increased BCL2 expression. Conclusion: Collectively, the data indicate that KRG represses oxidative stress-induced apoptosis by enhancing PI3K/Akt signaling via upregulation of ER-β expression.

Korean Red Ginseng inhibits apoptosis in neuroblastoma cells via estrogen receptor b-mediated phosphatidylinositol-3 kinase/Akt signaling

Cuong Thach Nguyen,Truc Thanh Luong,김규리,표석능,이동권 고려인삼학회 2015 Journal of Ginseng Research Vol.39 No.1

Background: Ginseng has been shown to exert antistress effects both in vitro and in vivo. However, theeffects of ginseng on stress in brain cells are not well understood. This study investigated how KoreanRed Ginseng (KRG) controls hydrogen peroxide-induced apoptosis via regulation of phosphatidylinositol-3 kinase (PI3K)/Akt and estrogen receptor (ER)-b signaling. Methods: Human neuroblastoma SK-N-SH cells were pretreated with KRG and subsequently exposed toH2O2. The ability of KRG to inhibit oxidative stress-induced apoptosis was assessed in MTT cytotoxicityassays. Apoptotic protein expression was examined byWestern blot analysis. The roles of ER-b, PI3K, andp-Akt signaling in KRG regulation of apoptosis were studied using small interfering RNAs and/or targetantagonists. Results: Pretreating SK-N-SH cells with KRG decreased expression of the proapoptotic proteins p-p53and caspase-3, but increased expression of the antiapoptotic protein BCL2. KRG pretreatment was alsoassociated with increased ER-b, PI3K, and p-Akt expression. Conversely, ER-b inhibition with smallinterfering RNA or inhibitor treatment increased p-p53 and caspase-3 levels, but decreased BCL2, PI3K,and p-Akt expression. Moreover, inhibition of PI3K/Akt signaling diminished p-p53 and caspase-3 levels,but increased BCL2 expression. Conclusion: Collectively, the data indicate that KRG represses oxidative stress-induced apoptosis byenhancing PI3K/Akt signaling via upregulation of ER-b expression.

Inhibition of Autolysis by Lipase LipA in Streptococcus pneumoniae Sepsis

Kim, Gyu-Lee,Luong, Truc Thanh,Park, Sang-Sang,Lee, Seungyeop,Ha, Jung Ah,Nguyen, Cuong Thach,Ahn, Ji Hye,Park, Ki-Tae,Paik, Man-Jeong,Pyo, Suhkneung,Briles, David E.,Rhee, Dong-Kwon Korean Society for Molecular and Cellular Biology 2017 Molecules and cells Vol.40 No.12

More than 50% of sepsis cases are associated with pneumonia. Sepsis is caused by infiltration of bacteria into the blood via inflammation, which is triggered by the release of cell wall components following lysis. However, the regulatory mechanism of lysis during infection is not well defined. Mice were infected with Streptococcus pneumoniae D39 wild-type (WT) and lipase mutant (${\Delta}lipA$) intranasally (pneumonia model) or intraperitoneally (sepsis model), and survival rate and pneumococcal colonization were determined. LipA and autolysin (LytA) levels were determined by qPCR and western blotting. S. pneumoniae Spd_1447 in the D39 (type 2) strain was identified as a lipase (LipA). In the sepsis model, but not in the pneumonia model, mice infected with the ${\Delta}lipA$ displayed higher mortality rates than did the D39 WT-infected mice. Treatment of pneumococci with serum induced LipA expression at both the mRNA and protein levels. In the presence of serum, the ${\Delta}lipA$ displayed faster lysis rates and higher LytA expression than the WT, both in vitro and in vivo. These results indicate that a pneumococcal lipase (LipA) represses autolysis via inhibition of LytA in a sepsis model.

Korean Red Ginseng inhibits apoptosis in neuroblastoma cells via estrogen receptor β-mediated phosphatidylinositol-3 kinase/Akt signaling

Cuong Thach Nguyen,Truc Thanh Luong,Gyu-Lee Kim,Suhkneung Pyo,Dong-Kwon Rhee 고려인삼학회 2015 Journal of Ginseng Research Vol.39 No.1

Background: Ginseng has been shown to exert antistress effects both in vitro and in vivo. However, the effects of ginseng on stress in brain cells are not well understood. This study investigated how Korean Red Ginseng (KRG) controls hydrogen peroxide-induced apoptosis via regulation of phosphatidylinositol- 3 kinase (PI3K)/Akt and estrogen receptor (ER)-b signaling. Methods: Human neuroblastoma SK-N-SH cells were pretreated with KRG and subsequently exposed to H₂O₂. The ability of KRG to inhibit oxidative stress-induced apoptosis was assessed in MTT cytotoxicity assays. Apoptotic protein expression was examined byWestern blot analysis. The roles of ER-β, PI3K, and p-Akt signaling in KRG regulation of apoptosis were studied using small interfering RNAs and/or target antagonists. Results: Pretreating SK-N-SH cells with KRG decreased expression of the proapoptotic proteins p-p53 and caspase-3, but increased expression of the antiapoptotic protein BCL2. KRG pretreatment was also associated with increased ER-β, PI3K, and p-Akt expression. Conversely, ER-b inhibition with small interfering RNA or inhibitor treatment increased p-p53 and caspase-3 levels, but decreased BCL2, PI3K, and p-Akt expression. Moreover, inhibition of PI3K/Akt signaling diminished p-p53 and caspase-3 levels, but increased BCL2 expression. Conclusion: Collectively, the data indicate that KRG represses oxidative stress-induced apoptosis by enhancing PI3K/Akt signaling via upregulation of ER-b expression.

TLR4 Mediates Pneumolysin-Induced ATF3 Expression through the JNK/p38 Pathway in Streptococcus pneumoniae-Infected RAW 264.7 Cells

Nguyen, Cuong Thach,Kim, Eun-Hye,Luong, Truc Thanh,Pyo, Suhkneung,Rhee, Dong-Kwon Korean Society for Molecular and Cellular Biology 2015 Molecules and cells Vol.38 No.1

Activating transcription factor-3 (ATF3) acts as a negative regulator of cytokine production during Gram-negative bacterial infection. A recent study reported that ATF3 provides protection from Streptococcus pneumoniae infection by activating cytokines. However, the mechanism by which S. pneumoniae induces ATF3 after infection is still unknown. In this study, we show that ATF3 was upregulated via Toll-like receptor (TLR) pathways in response to S. pneumoniae infection in vitro. Induction was mediated by TLR4 and TLR2, which are in the TLR family. The expression of ATF3 was induced by pneumolysin (PLY), a potent pneumococcal virulence factor, via the TLR4 pathway. Furthermore, ATF3 induction is mediated by p38 mitogen-activated protein kinase (MAPK) and c-Jun N-terminal kinase (JNK). Thus, this study reveals a potential role of PLY in modulating ATF3 expression, which is required for the regulation of immune responses against pneumococcal infection in macrophages.