T4 requires two proteins: holin, T (lesion formation and lysis timing) and
endolysin, E (cell wall degradation) to lyse the host at the end of its life cycle. E is a
cytoplasmic protein that sequestered away from its substrate, but the inner membrane
lesion formed by T allows E to gain access to the cell wall. T4 exhibits lysis inhibition
(LIN), a phenomenon in which a second T4 infection occurs ≤ 3 min after primary
infection results a delay in lysis. Mutations that abolish LIN mapped to several genes
but only rV encoding the holin, T, and rI, encoding the antiholin, RI, are required for
LIN in all hosts which support T4 replication. Antiholin RI inhibits T-mediated lysis by
direct interaction with the holin. T has at least one transmembrane domain with its Nterminus
(TNTD) in the cytoplasm and C-terminus in the periplasm (TCTD). In contrast,
the N-terminus of RI (RINTD) is predicted to function as a cleavable signal sequence
allowing the secretion of the RI C-terminal domain (RICTD) into the periplasm. Most of
RI mutations which abolish LIN occur in the RICTD, suggesting RI inhibits T-mediated
lysis by interacting with T via RICTD. Topological analysis of RI and T showed that fusion of PhoA signal sequence (ssPhoA) to RICTD is necessary and sufficient for LIN
and ssPhoAΦTCTD interferes with RI-mediated LIN, indicating T and RI interact via
periplasmic C-terminal domains.
In T4 infection, LIN is observed only when superinfection takes place, indicating
either the antiholin or the LIN signal must be unstable. Both RI and RINTDΦPhoA are
localized to both the inner membrane and the periplasm suggesting that the RINTD is a
Signal-Anchor-Release (SAR) domain. Protein stability studies indicated that the SAR
domain is the proteolytic determinant of RI, and DegP is the protease that is responsible
for RI degradation.
To date, how TNTD participates in lysis and LIN is not known. Modifications and
deletion of the N-terminus of T change the lysis time, indicating this domain is involved
the in timing of lysis. GFP fusion to holin T allowed microscopic visualization of
fluorescent patches on the membrane at the time of lysis.
| Identifer | oai:union.ndltd.org:tamu.edu/oai:repository.tamu.edu:1969.1/ETD-TAMU-1241 |
| Date | 15 May 2009 |
| Creators | Tran, Tram Anh Thi |
| Contributors | Young, Ryland |
| Source Sets | Texas A and M University |
| Language | en_US |
| Detected Language | English |
| Type | Book, Thesis, Electronic Dissertation, text |
| Format | electronic, application/pdf, born digital |
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