Identification of the YopE and YopH domains required for secretion and internalization into the cytosol of macrophages, using the cyaA gene fusion approach
Identification of the YopE and YopH domains required for secretion and internalization into the cytosol of macrophages, using the cyaA gene fusion approach
复制标题
DOI：
10.1073/pnas.92.26.11998
复制
发表时间：
1995-12-19
影响因子：
11.1
通讯作者：
Cornelis, GR
中科院分区：
文献类型：
--
作者：
Sory, MP;Boland, A;Cornelis, GR
致病性耶尔森氏菌通过IU型分泌机制分泌一组称为Yops的抗宿主蛋白。在感染培养的上皮细胞后，细胞外假结核耶尔森菌和小肠结肠炎耶尔森菌将细胞毒素YopE转运穿过宿主细胞质膜。一些证据表明酪氨酸磷酸酶YopH遵循相同的途径。我们用重组Y. YopE和YopH的融合蛋白很容易被小肠结肠炎耶尔森氏菌分泌。分泌所需的N端结构域不超过YopE的15个残基和YopH的17个残基。通过cAMP的产生揭示，内化到真核细胞中仅需要YopE的N-末端50个氨基酸残基和YopH的N-末端71个氨基酸残基。因此，YopE和YopH是由分泌结构域、易位结构域和效应结构域组成的模块蛋白。YopE和YopH跨宿主细胞膜的易位也依赖于同一细菌分泌YopB和YopD。环化酶融合的方法可以很容易地扩展到研究入侵性细菌病原体分泌的其他蛋白质的命运。
Pathogenic yersiniae secrete a set of antihost proteins, called Yops, by a type IU secretion mechanism. Upon infection of cultured epithelial tells, extracellular Yersinia pseudotuberculosis and Yersinia enterocolitica translocate cytotoxin YopE across the host cell plasma membrane. Several lines of evidence suggest that tyrosine phosphatase YopH follows the same pathway. We analyzed internalization of YopE and YopH into murine PU5-1.8 macrophages by using recombinant Y. enterocolitica producing truncated YopE and YopH proteins fused to a calmodulin-dependent adenylate cyclase, The YopE-cyclase and YopH-cyclase hybrids were readily secreted by Y. enterocolitica, The N-terminal domain required for secretion was not longer than 15 residues of YopE and 17 residues of YopH. Internalization into eukaryotic cells, revealed by cAMP production, only required the N-terminal 50 amino acid residues of YopE and the N-terminal 71 amino acid residues of YopH. YopE and YopH are thus modular proteins composed of a secretion domain, a translocation domain, and an effector domain, Translocation of YopE and YopH across host cell's membranes was also dependent on the secretion of YopB and YopD by the same bacterium. The cyclase fusion approach could be readily extended to study the fate of other proteins secreted by invasive bacterial pathogens.