Surface export of GAPDH/SDH, a glycolytic enzyme, is essential for Streptococcus pyogenes virulence.
Surface export of GAPDH/SDH, a glycolytic enzyme, is essential for Streptococcus pyogenes virulence.
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作者：
Jin H;Agarwal S;Agarwal S;Pancholi V
链球菌表面脱氢酶（SDH）（甘油醛-3-磷酸脱氢酶[GAPDH]）是A群链球菌（GAS）中的一种无锚定的主要多功能表面蛋白，能够结合包括纤溶蛋白（原）在内的重要哺乳动物蛋白。虽然SDH的一些生物学特性提示其可能在GAS毒力中起作用，但其在GAS发病机制中的直接作用尚未确定，因为它对GAS存活至关重要。因此，SDH/GAPDH“如何以及为什么”输出到细菌表面仍然是一个谜。目前的研究重点是“为什么”SDH被输出到GAS表面。通过在SDH的c末端插入疏水尾巴（M1-SDHHBtail），从而防止其输出到GAS表面，使用特定突变体进行了基于差分微阵列的全基因组转录物丰度分析。该分析显示，大多数与GAS毒力有关的基因和属于碳水化合物和氨基酸代谢的基因下调，与脂质代谢相关的基因上调。该突变体在小鼠模型中的毒力完全衰减，野生型和突变型菌株分别与SDHHBtail和SDH互补后毒力下降和增加，表明SDH表面输出确实调节了GAS的毒力。M1-SDHHBtail也表现出不变的生长模式，增加了细胞内ATP浓度和Hpr双磷酸化，显著降低了pH耐受性、链溶素S和SpeB活性。尽管已建立的转录调节因子的表达水平没有改变，但在突变体中观察到的这些表型和生理变化进一步强调了SDH与许多调节因子相结合的事实，其表面输出对GAS毒力至关重要。链球菌表面脱氢酶（SDH）是一种经典的无锚定胞质定位糖酵解酶，它通过一种迄今未知的机制被输出到a群链球菌（GAS）表面。目前尚不清楚为什么GAS或其他原核生物会将这种蛋白质输出到表面。通过遗传操作，我们创造了一种新的表达SDH的GAS突变株，其c端具有12个氨基酸的疏水尾部，因此能够在不改变其酶活性或生长模式的情况下阻止其表面输出。有趣的是，在小鼠腹膜炎模型中，该突变体的毒力完全减弱。该突变体的全球基因表达谱显示，表面输出SDH是维持GAS毒力的必要条件。GAS作为一种成功的病原体，能够将SDH定位在细胞质和表面，这在生理上和动态上是必要的，可以微调许多转录调节因子的功能，也可以利用其感染的毒力特性。
Streptococcal surface dehydrogenase (SDH) (glyceraldehyde-3-phosphate dehydrogenase [GAPDH]) is an anchorless major multifunctional surface protein in group A Streptococcus (GAS) with the ability to bind important mammalian proteins, including plasmin(ogen). Although several biological properties of SDH are suggestive of its possible role in GAS virulence, its direct role in GAS pathogenesis has not been ascertained because it is essential for GAS survival. Thus, it has remained enigmatic as to “how and why” SDH/GAPDH is exported onto the bacterial surface. The present investigation highlights “why” SDH is exported onto the GAS surface. Differential microarray-based genome-wide transcript abundance analysis was carried out using a specific mutant, which was created by inserting a hydrophobic tail at the C-terminal end of SDH (M1-SDHHBtail) and thus preventing its exportation onto the GAS surface. This analysis revealed downregulation of the majority of genes involved in GAS virulence and genes belonging to carbohydrate and amino acid metabolism and upregulation of those related to lipid metabolism. The complete attenuation of this mutant for virulence in the mouse model and the decreased and increased virulence of the wild-type and mutant strains postcomplementation with SDHHBtail and SDH, respectively, indicated that the SDH surface export indeed regulates GAS virulence. M1-SDHHBtail also displayed unaltered growth patterns, increased intracellular ATP concentration and Hpr double phosphorylation, and significantly reduced pH tolerance, streptolysin S, and SpeB activities. These phenotypic and physiological changes observed in the mutant despite the unaltered expression levels of established transcriptional regulators further highlight the fact that SDH interfaces with many regulators and its surface exportation is essential for GAS virulence. Streptococcal surface dehydrogenase (SDH), a classical anchorless cytoplasmically localized glycolytic enzyme, is exported onto the group A Streptococcus (GAS) surface through a hitherto unknown mechanism(s). It has not been known why GAS or other prokaryotes should export this protein onto the surface. By genetic manipulations, we created a novel GAS mutant strain expressing SDH with a 12-amino-acid hydrophobic tail at its C-terminal end and thus were able to prevent its surface exportation without altering its enzymatic activity or growth pattern. Interestingly, the mutant was completely attenuated for virulence in a mouse peritonitis model. The global gene expression profiles of this mutant reveal that the surface exportation of SDH is mandatory to maintain GAS virulence. The ability of GAS as a successful pathogen to localize SDH in the cytoplasm as well as on the surface is physiologically relevant and dynamically obligatory to fine-tune the functions of many transcriptional regulators and also to exploit its virulence properties for infection.