Guarding the vacuole: how Legionella T4SS substrate SdhA maintains vacuole integrity.
Anand, Ila.
2019
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Legionella pneumophila
is a Gram-negative, intracellular pathogen that targets amoebal species in aquatic
environments and alveolar macrophages in humans. To sustain intracellular replication,
L. pneumophila uses a type IV secretion system (T4SS) that translocates bacterial
protein substrates into the host cytoplasm. These T4SS substrates are involved in the
biogenesis of a membrane-bound ... read morecompartment known as the Legionella containing vacuole
(LCV). Our knowledge of LCV membrane integrity has been greatly enhanced by studying the
T4SS substrate SdhA. The SdhA substrate plays an important role in maintaining LCV
integrity, as a sdhA mutant exhibits a defect in vacuole integrity and intracellular
growth. However, the specific mechanism by which SdhA maintains vacuole integrity is not
well understood. To understand how SdhA maintains vacuole integrity during infection, we
performed high-throughput RNAi screens against host membrane trafficking genes to
identify factors that antagonize vacuole integrity in the absence of SdhA. Host proteins
that emerged from the screen were involved in endocytic and recycling pathways. We
focused our study on three Rab GTPases and found that depletion of Rab5 isoforms, Rab11b
isoform, or Rab8b isoform partially rescued vacuole integrity and intracellular growth
of Legionella in the absence of SdhA. Additionally, depletion of downstream effectors,
EEA1, Rab11FIP1, or VAMP3 rescued vacuole integrity and intracellular growth of the sdhA
mutant. Localization analyses revealed EEA1 and Rab11FIP1 colocalize with the sdhA
mutant at a higher frequency than the WT vacuole. These findings provide insights into
the host pathways that SdhA is regulating to maintain vacuole integrity during
infection.
Thesis (Ph.D.)--Tufts University, 2019.
Submitted to the Dept. of Molecular Microbiology.
Advisor: Ralph Isberg.
Committee: John Leong, Bree Aldridge, Shumin Tan, and Jonathan Kagan.
Keyword: Microbiology.read less - ID:
- k643bd56w
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