| Project/Area Number |
17590382
|
|---|
| Research Category |
Grant-in-Aid for Scientific Research (C)
|
| Allocation Type | Single-year Grants |
|---|
| Section | 一般 |
|---|
| Research Field |
Bacteriology (including Mycology)
|
|---|
| Research Institution | Chiba University (2006)
University of Tsukuba (2005) |
|---|
Principal Investigator |
SHIMIZU Takeshi Chiba University, Graducate School of Medicine, Assistant Professor, 大学院医学研究院, 講師 (70312840)
|
|---|
| Co-Investigator(Kenkyū-buntansha) |
HAMABATA Takashi International Medical Center of Japan, Research Institute, Division Chief, 感染症制御研究部, 室長 (40311427)
|
|---|
| Project Period (FY) |
2005 – 2006
|
| Project Status |
Completed (Fiscal Year 2006)
|
| Budget Amount *help |
¥3,700,000 (Direct Cost: ¥3,700,000)
Fiscal Year 2006: ¥1,600,000 (Direct Cost: ¥1,600,000)
Fiscal Year 2005: ¥2,100,000 (Direct Cost: ¥2,100,000)
|
|---|
| Keywords | Enterohemorrhagic Escherichia coli / Shiga toxins / Secretion / Virulence / Gb3 / Bacterial infection |
|---|
| Research Abstract |
Shiga toxins (Stxs) produced by enterohemorrhagic Escherichia coli (EHEC) include shiga toxin 1 (Stx1) as well as shiga toxin 2 (Stx2). Stx1 is cell-associated, whereas Stx2 is localized to the culture supernatant. We have analyzed the secretion of Stx2 by generating histidine-tagged StxB (StxBH). Although neither Stx1BH nor Stx2BH was secreted in StxBH-overexpressed EHEC, Stx2BH-overexpressed EHEC showed inhibited Stx2 secretion. On the other hand, Stx1BH-overexpressed EHEC showed no alteration of Stx2 secretion. B subunit chimeras of Stx1 and Stx2 were used to identify the specific residue of Stx2B that the Stx2 secretory system recognizes. Alteration of the serine 31 residue to an asparagine residue (S31N) in Stx2BH enabled the recovery of Stx2 secretion. On the other hand, alteration of the asparagine 32 residue to a serine residue (N32S) in Stx1BH caused partial secretion of a point-mutated histidine-tagged B subunit in EHEC. Based on the evidence, it appeared possible that this residue might contain secretion-related information for Stx2 secretion. To investigate this hypothesis, we constructed isogenic mutant strains, Stx1 (B subunit, N32S)-producing EHEC and Stx2 (B subunit, S31N)-producing EHEC. Although the mutant Stx2 was cell-associated in isogenic mutant EHEC, the mutant Stx1 was not extracellular. However, when we used plasmids for the expression of the mutant holotexins, the overexpressed mutant Stx1 was found in the supernatant fraction and the overexpressed mutant Stx2 was found in the cell-associated fraction in the mutant holotexin gene-transformed EHEC. These results indicate that the serine 31 residue of the B subunit of Stx2 contains secretion-related information.
|
