絲氨酸蛋白酶Rv2569c通過切割e -鈣粘蛋白破壞上皮屏障，促進結核分枝桿菌的傳播

2024年5月，中國農業科學院哈爾濱獸醫研究所動物疫病預防控制國家重點實驗室(State Key Laboratory for Animal Disease Control and Prevention, Division of Bacterial Diseases, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin, PR China)劉思國老師研究團隊在《PLoS Pathog.》上發表論文：

“Serine protease Rv2569c facilitates transmission of Mycobacterium tuberculosis via disrupting the epithelial barrier by cleaving E-cadherin”

“絲氨酸蛋白酶Rv2569c通過切割e -鈣粘蛋白破壞上皮屏障，促進結核分枝桿菌的傳播”

Abstract：

Epithelial cells function as the primary line of defense against invading pathogens. However,

bacterial pathogens possess the ability to compromise this barrier and facilitate the transmi

gration of bacteria. Nonetheless, the specific molecular mechanism employed by Mycobacterium tuberculosis (M.tb) in this process is not fully understood. Here, we investigated the role of Rv2569c in M.tb translocation by assessing its ability to cleave E-cadherin, a crucial component of cell-cell adhesion junctions that are disrupted during bacterial invasion. By utilizing recombinant Rv2569c expressed in Escherichia coli and subsequently purified through affinity chromatography, we demonstrated that Rv2569c exhibited cell wall–associated serine protease activity. Furthermore, Rv2569c was capable of degrading a range of protein substrates, including casein, fibrinogen, fibronectin, and E-cadherin. We also deter

mined that the optimal conditions for the protease activity of Rv2569c occurred at a temperature of 37?C and a pH of 9.0, in the presence of MgCl2. To investigate the function of Rv2569c in M.tb, a deletion mutant of Rv2569c and its complemented strains were generated and used to infect A549 cells and mice. The results of the A549-cell infection experiments revealed that Rv2569c had the ability to cleave E-cadherin and facilitate the transmigration of M.tb through polarized A549 epithelial cell layers. Furthermore, in vivo infection assays demonstrated that Rv2569c could disrupt E-cadherin, enhance the colonization of M.tb, and induce pathological damage in the lungs of C57BL/6 mice. Collectively,

these results strongly suggest that M.tb employs the serine protease Rv2569c to disrupt epithelial defenses and facilitate its systemic dissemination by crossing the epithelial barrier.

摘要：

上皮細胞是抵御病原體入侵的主要防線。然而，細菌病原體具有破壞這一屏障并促進細菌遷移的能力。然而，結核分枝桿菌(M.tb)在這一過程中所采用的具體分子機制尚不完全清楚。在這里，科研人員通過評估Rv2569c切割e -鈣粘蛋白的能力來研究Rv2569c在結核分枝桿菌易位中的作用，e -鈣粘蛋白是細胞-細胞粘附連接的重要組成部分，在細菌入侵期間被破壞。利用在大腸桿菌中表達并通過親和層析純化的重組Rv2569c，科研人員證明了Rv2569c具有細胞壁相關絲氨酸蛋白酶活性。此外，Rv2569c能夠降解一系列蛋白質底物，包括酪蛋白、纖維蛋白原、纖維連接蛋白和e -鈣粘蛋白。科研人員還確定了Rv2569c蛋白酶活性的**條件是溫度為37℃，pH為9.0,MgCl2存在。為了研究Rv2569c在結核分枝桿菌中的功能，科研人員制備了Rv2569c的缺失突變體及其補充菌株，并將其用于感染A549細胞和小鼠。A549細胞感染實驗結果顯示，Rv2569c具有裂解E-cadherin的能力，促進M.tb通過極化的A549上皮細胞層遷移。此外，體內感染實驗表明，Rv2569c可以破壞E-cadherin，增強M.tb的定植，并在C57BL/6小鼠的肺部引起病理損傷。總之，這些結果強烈表明結核分枝桿菌利用絲氨酸蛋白酶Rv2569c破壞上皮防御，并通過跨越上皮屏障促進其全身傳播。

該論文中，A549細胞體外培養是使用Ausbian特級胎牛血清完成的。欲了解或購買Ausbian特級胎牛血清可以聯系北京締一生物400-166-8600.