IglF mediates type VI secretion system spike assembly and promotes <i>Francisella</i> virulence.

Abstract

Type VI secretion systems (T6SSs) are widely distributed among Gram-negative bacteria, where they mostly act to promote bacterial warfare. Bacteria from the <i>Francisella</i> genus possess T6SSs that phylogenetically diverge from all other T6SSs and constitute the T6SSii subtype. <i>Francisella</i> tularensis, the agent of tularemia, relies on its T6SS to secrete effectors into host cells. Despite the key role of this nanomachine in <i>Francisella</i> virulence, the structure of T6SSii and the mechanism underlying its assembly are still poorly understood. Here, using <i>Francisella novicida</i>, we focused on understanding the structure and assembly of the spike, the most apical T6SS complex coupling effector delivery and membrane-puncturing activity. We solved the structure of the protein of unknown function, IglF, in complex with the N-terminal domain of IglG, the T6SSii PAAR protein. Interaction between IglF and IglG enabled the assembly of a mature T6SS spike complex both in <i>Francisella</i> and in a heterologous expression system. In contrast, disrupting IglF:IglG interactions prevented assembly of the PAAR protein with the central spike complex and invalidated T6SS assembly, as visualized by monitoring T6SS dynamics or secretion. Accordingly, IglF:IglG interactions were required for <i>F. novicida</i> virulence in vitro and in a mouse model of tularemia. Altogether, our findings shed light on the assembly mechanism of the <i>Francisella</i> T6SSii spike complex and its importance in virulence.