Contractile injection systems (CISs) are phage tail-like nanomachines, mediating bacterial cell–cell interactions as either type VI secretion systems (T6SSs) or extracellular CISs (eCISs). Bioinformatic studies uncovered a phylogenetic group of hundreds of putative CIS gene clusters that are highly diverse and widespread; however, only four systems have been characterized. Here we studied a putative CIS gene cluster in the marine bacterium Algoriphagus machipongonensis. Using an integrative approach, we show that the system is compatible with an eCIS mode of action. Our cryo-electron microscopy structure revealed several features that differ from those seen in other CISs: a ‘cap adaptor’ located at the distal end, a ‘plug’ exposed to the tube lumen, and a ‘cage’ formed by massive extensions of the baseplate. These elements are conserved in other CISs, and our genetic tools identified that they are required for assembly, cargo loading and function. Furthermore, our atomic model highlights specific evolutionary hotspots and will serve as a framework for understanding and re−engineering CISs.

Contractile injection systems (CISs) mediate cell–cell interactions by phage tail-like structures, using two distinct modes of action: extracellular CISs are released into the medium, while type 6 secretion systems (T6SSs) are attached to the cytoplasmic membrane and function upon cell–cell contact. Here, we characterized a CIS in the multicellular cyanobacterium Anabaena, with features distinct from extracellular CISs and T6SSs. Cryo-electron tomography of focused ion beam-milled cells revealed that CISs were anchored in thylakoid membrane stacks, facing the cell periphery. Single particle cryo-electron microscopy showed that this unique in situ localization was mediated by extensions of tail fibre and baseplate components. On stress, cyanobacteria induced the formation of ghost cells, presenting thylakoid-anchored CISs to the environment. Functional assays suggest that these CISs may mediate ghost cell formation and/or interactions of ghost cells with other organisms. Collectively, these data provide a framework for understanding the evolutionary re-engineering of CISs and potential roles of these CISs in cyanobacterial programmed cell death.

Src associated in mitosis (SAM68) plays major roles in regulating RNA processing events, such as alternative splicing and mRNA translation, implicated in several developmental processes. It was previously shown that SAM68 regulates the alternative splicing of the mechanistic target of rapamycin ( mTor ), but the mechanism regulating this process remains elusive. Here, we report that SAM68 interacts with U1 small nuclear ribonucleoprotein (U1 snRNP) to promote splicing at the 5′ splice site in intron 5 of mTor . We also show that this direct interaction is mediated through U1A, a core-component of U1snRNP. SAM68 was found to bind the RRM1 domain of U1A through its C-terminal tyrosine rich region (YY domain). Deletion of the U1A-SAM68 interaction domain or mutation in SAM68-binding sites in intron 5 of mTor abrogates U1A recruitment and 5′ splice site recognition by the U1 snRNP, leading to premature intron 5 termination and polyadenylation. Taken together, our results provide the first mechanistic study by which SAM68 modulates alternative splicing decision, by affecting U1 snRNP recruitment at 5′ splice sites.

At the end of a lytic bacteriophage replication cycle in Gram-positive bacteria, peptidoglycan-degrading endolysins that cause explosive cell lysis of the host can also attack non-infected bystander cells. Here we show that in osmotically stabilized environments, Listeria monocytogenes can evade phage predation by transient conversion to a cell wall-deficient L-form state. This L-form escape is triggered by endolysins disintegrating the cell wall from without, leading to turgor-driven extrusion of wall-deficient, yet viable L-form cells. Remarkably, in the absence of phage predation, we show that L-forms can quickly revert to the walled state. These findings suggest that L-form conversion represents a population-level persistence mechanism to evade complete eradication by phage attack. Importantly, we also demonstrate phage-mediated L-form switching of the urinary tract pathogen Enterococcus faecalis in human urine, which underscores that this escape route may be widespread and has important implications for phage- and endolysin-based therapeutic interventions.

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