Characterization of l0032 and escP in the Pathogenic Island of Enterohemorrhagic Escherichia coli O157:H7

• Main Authors: Hao-Wei Chang, 張浩維
• Other Authors: Wan-Jr Syu
• Format: Others
• Language: en_US
• Published: 2014
• Online Access: http://ndltd.ncl.edu.tw/handle/51913562452288659698

碩士 === 國立陽明大學 === 微生物及免疫學研究所 === 102 === Abstract Enterohemorrhagic Escherichia coli (EHEC) employs a type III secretion system (T3SS) to translocate virulence effector proteins directly into host enterocytes, a critical process leading to infection and diarrheal diseases. The T3SS is encoded by a chromosomal island called locus of enterocyte effacement (LEE). The LEE island comprises of 41 open reading frames (ORFs) that could be divided into five major operons, Lee1 to Lee5. Functions of some LEE-encoded proteins remain under-characterized. Here, we investigated whether L0032 or EscP is involved in T3SS biogenesis and functions in EHEC. L0032 and escP are two different size genes but annotated from distinct start codons on the same location. The l0032-deletion mutant (L32) and the escP-deletion mutant (EscP) both showed translocator and effector protein secretion-defective profiles that differed from those of the wild-type strain. While doing the complementation assays, EscP could restore the T3SS phenotype of the two mutants but L0032 could not. Therefore, the protein with a full function must be translated from the upstream rare start codon (TTG) that leads to EscP while starting from the downstream ATG codon gives to partially functional L0032. Intriguingly, in the l0032-deletion, the intracellular levels of EspA, EspB and Tir seemed to be lower than that seen with the wild type. Judging from the levels of ectopically expressed constructs, which were driven by T5, no apparent difference was seen between the level in L32 and that in the wild type. Furthermore, EscP interacted neither with LEE regulators nor with chaperons/binding proteins of EspA. However, in a K-12 strain-based Lee1 promoter reporter assay, both L0032 and EscP exhibited suppressive activities. Whether these activities occur in EHEC remain to be explored.