Double Tubular Contractile Structure of the Type VI Secretion System Displays Striking Flexibility and Elasticity.

Antimicrobial treatment can induce many bacterial pathogens to enter a cell wall-deficient state that contributes to persistent infections. The effect of this physiological state on the assembly of transenvelope-anchored organelles is not well understood. The type VI secretion system (T6SS) is a widespread molecular weapon for interspecies interactions and virulence, comprising a long double tubular structure and a transenvelope/baseplate complex. Here, we report that cell wall-deficient spheroplasts assembled highly flexible and elastic T6SS structures forming U, O, or S shapes. Upon contacting the inner membrane, the T6SS tubes did not contract but rather continued to grow along the membrane. Such deformation likely results from continual addition of sheath/tube subunits at the distal end. Induction of TagA repressed curved sheath formation. Curved sheaths could also contract and deliver T6SS substrates and were readily disassembled by the ClpV ATPase after contraction. Our data highlight the dramatic effect of cell wall deficiency on the shape of the T6SS structures and reveal the elastic nature of this double tubular contractile injection nanomachine. The cell wall is a physical scaffold that all transenvelope complexes have to cross for assembly. However, the cell wall-deficient state has been described as a common condition found in both Gram-negative and Gram-positive pathogens during persistent infections. Loss of cell wall is known to have pleiotropic physiological effects, but how membrane-anchored large cellular organelles adapt to this unique state is less completely understood. Our study examined the assembly of the T6SS in cell wall-deficient spheroplast cells. We report the elastic nature of contractile T6SS tubules under such conditions, providing key insights for understanding how large intracellular structures such as the T6SS accommodate the multifaceted changes in cell wall-deficient cells.