Programmable protein delivery with a bacterial contractile injection system

Abstract

Endosymbiotic bacteria have evolved intricate delivery systems that enable these organisms to interface with host biology. One example, the extracellular contractile injection systems (eCISs), are syringe-like macromolecular complexes that inject protein payloads into eukaryotic cells by driving a spike through the cellular membrane. Recently, eCISs have been found to target mouse cells1–3, raising the possibility that these systems could be harnessed for therapeutic protein delivery. However, whether eCISs can function in human cells remains unknown, and the mechanism by which these systems recognize target cells is poorly understood. Here we show that target selection by the Photorhabdus virulence cassette (PVC)—an eCIS from the entomopathogenic bacterium Photorhabdus asymbiotica—is mediated by specific recognition of a target receptor by a distal binding element of the PVC tail fibre. Furthermore, using in silico structure-guided engineering of the tail fibre, we show that PVCs can be reprogrammed to target organisms not natively targeted by these systems—including human cells and mice—with efficiencies approaching 100%. Finally, we show that PVCs can load diverse protein payloads, including Cas9, base editors and toxins, and can functionally deliver them into human cells. Our results demonstrate that PVCs are programmable protein delivery devices with possible applications in gene therapy, cancer therapy and biocontrol.

Proximity-Based Modalities for Biology and Medicine

“…The potential antigenicity and in vivo delivery challenge of macromolecule-based modalities may also be problematic, yet this remains an intense area of development. 95 To overcome these hurdles, lysosome-based degraders can leverage experiences from PROTAC degraders, such as investigating the structure−activity relationship, in-depth mechanism investigation and developing assays to detect the…”

“…Their degradation kinetics and potencies are not comparable with those of most well-developed PROTACs and MGs, which may be attributed to the lack of catalytic character or the decomposition of the proximity agents in the acidic and enzymatic lysosome. The potential antigenicity and in vivo delivery challenge of macromolecule-based modalities may also be problematic, yet this remains an intense area of development . To overcome these hurdles, lysosome-based degraders can leverage experiences from PROTAC degraders, such as investigating the structure–activity relationship, in-depth mechanism investigation and developing assays to detect the lysosome function and to monitor each step of the degradation pathway.…”

Revolutionary delivery system enables precise protein transportation

“…Inspired by previous reports regarding CISs, recently, a team led by Professor Feng Zhang at the Broad Institute developed a redesigned protein delivery system; the corresponding results have been published in Nature [5] . Therein, extracellular contractile injection systems (eCISs), syringe‐like nanomachines mimicking bacteriophage tails that can transport payloads independently and extracellularly, served as a new tool to solve a long‐standing problem, that is, how to deliver therapeutic molecules to specific types of human cells precisely and efficiently (Figure 1).…”

“…[4] Inspired by previous reports regarding CISs, recently, a team led by Professor Feng Zhang at the Broad Institute developed a redesigned protein delivery system; the corresponding results have been published in Nature. [5] Therein, extracellular contractile injection systems (eCISs), syringelike nanomachines mimicking bacteriophage tails that can transport payloads independently and extracellularly, served as a new tool to solve a long-standing problem, that is, how to deliver therapeutic molecules to specific types of human cells precisely and efficiently (Figure 1). The structural composition of eCISs originating from the Photorhabdus virulence cassette (PVC) is such that the tail fibers on the outside of one end recognize specific receptors on the cell surface and anchor to host cells; thus, in their study, the researchers speculated that modifying the structure of these tail fibers may enable them to recognize different cells.…”

Information storage across a microbial community using universal RNA memory

“…Nucleic acids acquired through gene transfer can alter cell-cell interactions (1,2), facilitate niche expansion (3), drive bacterial resistance to phage (4), and control gene stability (5). Gene transfer is also critical to microbial domestication for synthetic biology (6)(7)(8) and a challenge for the safe application of such technologies (9). Because gene transfer can occur across species from different genera, phyla, and even kingdoms (10,11), there is a need to understand how it varies across communities.…”