HM host membrane, OM bacterial outer membrane, IM bacterial inner membrane. c Sequestration of the bait-SctQ fusion protein to the membrane prevents effector secretion.
Conversely, in the LITESEC-act system (bottom), the bait protein, a fusion of the smaller interaction switch domain, Zdk1, and the essential T3SS component SctQ, is tethered to the membrane anchor, a TMH fusion of the larger interaction switch domain, LOV2, in the dark, and gets released by illumination. In the LITESEC-supp system (top), the bait protein, a fusion of the smaller interaction switch domain SspB_Nano and the essential T3SS component SctQ, is tethered to the inner membrane (IM) by a membrane anchor, a fusion of a transmembrane helix (TMH) and the larger interaction switch domain, iLID, in the light, and gets released in the dark. b Different states of the bait and anchor proteins in dark and light conditions. Effector translocation by the T3SS is licensed by the functional interaction of the unbound bait-SctQ fusion with the T3SS. Left side, main substructures right side, dynamic cytosolic T3SS components. LITESEC-T3SS constitutes a new method to control protein secretion and translocation into eukaryotic host cells with unparalleled spatial and temporal resolution.Ī Schematic representation of the active T3SS injectisome (modified from ref. We demonstrate the light-regulated translocation of heterologous reporter proteins, and induction of apoptosis in cultured eukaryotic cells. The resulting system, LITESEC-T3SS (Light-induced translocation of effectors through sequestration of endogenous components of the T3SS), allows rapid, specific, and reversible activation or deactivation of the T3SS upon illumination. Using optogenetic interaction switches to control the availability of the dynamic cytosolic T3SS component SctQ, T3SS-dependent effector secretion can be regulated by light. In this study, we exploit the dynamic nature of the T3SS to govern its activity. Although the T3SS can efficiently export heterologous cargo proteins, a lack of target cell specificity currently limits its application in biotechnology and healthcare. Many bacteria employ a type III secretion system (T3SS) injectisome to translocate proteins into eukaryotic host cells.
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