NAD-Glycohydrolase Depletes Intracellular NAD+ and Inhibits Acidification of Autophagosomes to Enhance Multiplication of Group A Streptococcus in Endothelial Cells.

Cheng-Lu Hsieh,Chuan Chiang-Ni,Shu-Ying Hsieh,Yee-Shin Lin,Ching-Chuan Liu,Jiunn-Jong Wu,Po-Xing Zheng,Pei-Jane Tsai,Shu-Ying Wang,Hsuan-Min Huang

doi:10.3389/fmicb.2018.01733

Abstract

Group A Streptococcus (GAS) is a human pathogen causing a wide spectrum of diseases, from mild pharyngitis to life-threatening necrotizing fasciitis. GAS has been shown to evade host immune killing by invading host cells. However, how GAS resists intracellular killing by endothelial cells is still unclear. In this study, we found that strains NZ131 and A20 have higher activities of NADase and intracellular multiplication than strain SF370 in human endothelial cells (HMEC-1). Moreover, nga mutants of NZ131 (SW957 and SW976) were generated to demonstrate that NADase activity is required for the intracellular growth of GAS in endothelial cells. We also found that intracellular levels of NAD+ and the NAD+/NADH ratio of NZ131-infected HMEC-1 cells were both lower than in cells infected by the nga mutant. Although both NZ131 and its nga mutant were trapped by LC3-positive vacuoles, only nga mutant vacuoles were highly co-localized with acidified lysosomes. On the other hand, intracellular multiplication of the nga mutant was increased by bafilomycin A1 treatment. These results indicate that NADase causes intracellular NAD+ imbalance and impairs acidification of autophagosomes to escape autophagocytic killing and enhance multiplication of GAS in endothelial cells.

Highlights

Group A Streptococcus (GAS) is an important human pathogen responsible for causing wide spectrum of diseases, ranging from superficial infections to life-threatening manifestations including necrotizing fasciitis and streptococcal toxic-shock syndrome (Cunningham, 2008)
To demonstrate whether NADase activity is required for GAS multiplication in HMEC-1 cells, the ribosomal rpsL promoter was utilized to drive an endogenous inhibitor of NADase, IFS, in the wild-type NZ131
The results demonstrated that overexpression of ifs in NZ131 (SW960) resulted in the similar phenotypes, including NADase and intracellular survival, compared to the nga mutant (Figures 1E,F)

Summary

Introduction

Group A Streptococcus (GAS) is an important human pathogen responsible for causing wide spectrum of diseases, ranging from superficial infections to life-threatening manifestations including necrotizing fasciitis and streptococcal toxic-shock syndrome (Cunningham, 2008). NADase Enhance Intracellular Multiplication of GAS that GAS can invade epithelial cells to escape killing by host immune responses and antibiotics (LaPenta et al, 1994; Osterlund et al, 1997; Kaplan et al, 2006). SLO is a pore-forming toxin that forms oligomeric pores to disrupt cell membranes and facilitate autophagy formation, which contributes to enhance GAS survival in the intracellular niche of host cells (Sierig et al, 2003; Nakagawa et al, 2004). Recent studies showed that the epidemic M1 and M89 GAS strains, which are rapidly spreading globally, produce higher levels of NADase and SLO to cause severe tissue destruction (Turner et al, 2015; Zhu et al, 2015a,b, 2016), indicating the importance of NADase and SLO in GAS pathogenesis

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Results

Conclusion