Abstract
The type VI secretion system (T6SS) is a contractile injection apparatus that translocates a spike loaded with various effectors directly into eukaryotic and prokaryotic target cells. Such T6SS spike consists of a needle-shaped trimer of VgrG proteins topped by a conical and sharp PAAR protein that facilitates puncturing of the target membrane. T6SS-delivered effector proteins can be either fused to one of the two spike proteins or interact with either in a highly specific manner. In Agrobacterium tumefaciens the T6SS effector Tde1 is targeted to its cognate VgrG1 protein. Here, we attempted to use a VgrG shuttle to deliver a heterologous T6SS effector by directing Tde1 onto a T6SS spike in Pseudomonas aeruginosa. For this, we designed chimeras between VgrG1 from A. tumefaciens and VgrG1a from P. aeruginosa and showed that modification of the spike protein hampered T6SS functionality in the presence of the Tde1 effector complex. We provide evidence suggesting that Tde1 specifically binds to the VgrG spike in the heterologous environment and propose that there are additional requirements to allow proper effector delivery and translocation. Our work sheds light on complex aspects of the molecular mechanisms of T6SS delivery and highlights some limitations on how effectors can be translocated using this nanomachine.
Highlights
The T6SS is a versatile secretion system, injecting effector proteins into target cells which equips bacteria with the ability to establish a niche in any given polymicrobial environment or modulate host cell responses
We chose Tde1 from A. tumefaciens as the heterologous effector, firstly because the Tde1 orthologs are only found in a-proteobacteria and secondly because, its basic delivery system has previously been studied in great details (Ma et al, 2014; Bondage et al, 2016; Wu et al, 2020)
The current model states that VgrG1 from A. tumefaciens (VgrG1A) is assembled to a functional trimer and capped by the cognate PAAR protein, with one PAAR protein binding the three last VgrG1A β-strands
Summary
The T6SS is a versatile secretion system, injecting effector proteins into target cells which equips bacteria with the ability to establish a niche in any given polymicrobial environment or modulate host cell responses. The cytosolic tubular sheath attaches to the baseplate at the inner membrane and encompasses a tube composed of Hcp hexamers that is propelled out of the cell upon sheath contraction (Pukatzki et al, 2006; Leiman et al, 2009; Brunet et al, 2014). On top of the Hcp tube and residing within the baseplate complex sits the so-called T6SS spike consisting of a needle-shaped trimer of VgrG proteins and a conically-shaped PAAR protein (Shneider et al, 2013). The VgrG-PAAR spike complex has two main functions: it facilitates puncturing of target membranes while it is directly involved in carrying T6SS effectors into the target cell (Shneider et al, 2013)
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