Ascertaining cells' synaptic connections and RNA expression simultaneously with barcoded rabies virus libraries.

Brain function depends on synaptic connections between specific neuron types, yet systematic descriptions of synaptic networks and their molecular properties are not readily available. Here, we introduce SBARRO (Synaptic Barcode Analysis by Retrograde Rabies ReadOut), a method that uses single-cell RNA sequencing to reveal directional, monosynaptic relationships based on the paths of a barcoded rabies virus from its "starter" postsynaptic cell to that cell's presynaptic partners. Thousands of these partner relationships can be ascertained in a single experiment, alongside genome-wide RNAs. We use SBARRO to describe synaptic networks formed by diverse mouse brain cell types in vitro, finding that different cell types have presynaptic networks with differences in average size and cell type composition. Patterns of RNA expression suggest that functioning synapses are critical for rabies virus uptake. By tracking individual rabies clones across cells, SBARRO offers new opportunities to map the synaptic organization of neural circuits.

RNAi screen reveals a role for PACSIN2 and caveolins during bacterial cell-to-cell spread.

Listeria monocytogenes is a human bacterial pathogen that disseminates through host tissues using a process called cell-to-cell spread. This critical yet understudied virulence strategy resembles a vesicular form of intercellular trafficking that allows L. monocytogenes to move between host cells without escaping the cell. Interestingly, eukaryotic cells can also directly exchange cellular components via intercellular communication pathways (e.g., trans-endocytosis) using cell-cell adhesion, membrane trafficking, and membrane remodeling proteins. Therefore, we hypothesized that L. monocytogenes would hijack these types of host proteins during spread. Using a focused RNA interference screen, we identified 22 host genes that are important for L. monocytogenes spread. We then found that caveolins (CAV1 and CAV2) and the membrane sculpting F-BAR protein PACSIN2 promote L. monocytogenes protrusion engulfment during spread, and that PACSIN2 specifically localizes to protrusions. Overall, our study demonstrates that host intercellular communication pathways may be coopted during bacterial spread and that specific trafficking and membrane remodeling proteins promote bacterial protrusion resolution.