The newly identified roles of Infection-Associated Macropinosomes

Salmonella is a bacteria that triggers its entry into non-phagocytic epithelial cells. For this, Salmonella injects a cocktail of bacterial proteins into the host cell cytosol leading to the remodeling of the actin cytoskeleton and the formation of host membrane protrusions called ruffles. Some ruffles enclose the extracellular bacterium, fuse, and pinch-off from the plasma membrane, allowing the pathogen to enter its host cells within a membrane-bound compartment called the Salmonella-Containing Vacuole (SCV). Concomitantly, other ruffles engulf extracellular fluid leading to the formation of large vesicles called infection-associated macropinosomes (IAMs*) at the infection site.

During my PhD in the Enninga Lab at Institut Pasteur (Paris), I studied the interaction between SCV and IAMs. Altogether, my PhD work established the following:

• The SCV is a tight compartment, distinct from IAMs.

• A few minutes after formation, SCV and IAMs fuse together, which results in an increase in the SCV size.

• IAMs can be isolated for mass spectrometry analysis using magnetic beads.

• The SNAREs SNAP25, VAMP2, STX4, and STX12 promote SCV-IAM fusion.

• The SCV growth is compensated by the formation of tubules emanating from the SCV and leading to SCV shrinkage.

• The equilibrium between the SCV expansion and shrinkage determines the SCV stability: while favoring shrinkage over expansion promotes the SCV rupture and the liberation of Salmonella inside the host cytosol, the fusion of IAMs with the SCV promotes the establishment of the Salmonella vacuolar niche.

Associated publications:

• Fredlund J#, Santos JC#, Stévenin V#, Weiner A, Rechav K, Mallet A, Krijnse-Locker J, Elbaum M, and Enninga J. 2018. The early entry of Salmonella in a distinct tight compartment revealed at high temporal and ultrastructural resolution. Cellular Microbiology. 20(4):e12816. doi: 10.1111/cmi.12816. (.pdf) (#equal contribution)

• Stévenin V, Chang YY, Le Toquin Y, Duchateau M, Gianetto QG, Luk CH, Salles A, Sohst V, Matondo M, Reiling N, Enninga J. 2019. Dynamic growth and shrinkage of the Salmonella-containing vacuole determines the intracellular pathogen niche. Cell Reports. 29(12):3958-3973.e7. doi: 10.1016/j.celrep.2019.11.049. (.pdf)

• Chang YY, Stévenin V, Duchateau M, Giai Gianetto Q, Hourdel V, Rodrigues CD, Matondo M, Reiling N, Enninga J. 2020. Shigella hijacks the exocyst to cluster macropinosomes for efficient vacuolar escape. PLoS Pathogens. 16(8):e1008822. doi: 10.1371/journal.ppp1008822. (.pdf)

• Chang YY, Enninga J, Stévenin V. 2021. New methods to decrypt emerging macropinosome functions during the host-pathogen crosstalk. Cellular Microbiology. 23(7):e13342. doi: 10.1111/cmi.13342. (.pdf)

• Stévenin V, Giai Gianetto Q, Duchateau M, Matondo M, Enninga J, Chang YY. 2021. Purification of infection-associated macropinosomes by magnetic isolation for proteomic characterization. Nature Protocols. 16(11):5220-5249. doi: 10.1038/s41596-021-00610-5. (.pdf)

• Schink K, Tan KW, Spangenberg H, Martorana D, Sneeggen M, Stévenin V, Enninga J, Campsteijn C, Raiborg C, Stenmark H. 2021. The phosphoinositide coincidence detector Phafin2 promotes macropinocytosis by coordinating actin organisation at forming macropinosomes. Nature Communication. 12(1):6577. doi: 10.1038/s41467-021-26775-x. (.pdf)

Recently featured in these reviews:

• Fang Z, Méresse S. Endomembrane remodeling and dynamics in Salmonella infection. 2021 Microb Cell. 27;9(2):24-41. doi: 10.15698/mic2022.02.769

• Petit TJP, Lebreton A. Adaptations of intracellular bacteria to vacuolar or cytosolic niches. 2022. Trends Microbiol. 30(8):736-748. doi: 10.1016/j.tim.2022.01.015.

• Gutierrez MG, Enninga J. Intracellular niche switching as host subversion strategy of bacterial pathogens. 2022 Curr Opin Cell Biol. 76:102081. doi: 10.1016/j.ceb.2022.102081.

• Luk CH, Enninga J, Valenzuela C. Fit to dwell in many places – The growing diversity of intracellular Salmonella niches. Front. Cell. Infect. Microbiol. 12:989451. doi: 10.3389/fcimb.2022.989451

*Funny fact: we picked the name "IAM" to distinguish these compartments from classical macropinosomes. We also found this name pretty cool, as a reference to the famous French band from the 90s'.