Lab Members

In the Liu lab, I am utilizing single particle cryo-EM analysis and cryo-electron tomography to study the conformational changes of ejection proteins during genome ejection in phages T7 and P22. By combining these state-of-the-art techniques with extensive knowledge from biochemical and genetic studies, we aim to reveal the molecular mechanisms of ejection protein remodeling during genome ejection. Our findings provide detailed structural and functional insights into phage infection.

Efficient genome ejection is a critical yet little understood step in phage infection and requires large conformational changes in the mature tail machine. Specifically, podophages possess short tails that cannot span the Gram-negative cell envelope to allow direct genome ejection into the cell cytoplasm. These phages contain ejection proteins present in the mature capsid, possibly in complex with the genomic dsDNA. As the tail machine changes conformation after productive adsorption, these ejection proteins relocate inside the host bacteria, where they spontaneously assemble, forming a conduit for DNA translocation across the entire cell envelope. By contrast, the ejection proteins of podophages T7 and P22 have different conformations inside mature phages, both assembling a trans-envelope channel during genome ejection (Hu et al, 2013; Wang et al, 2019) using distinct mechanisms.

NCBI My Bibliography

Related lab publications:

• Wang, C., Tu, J., Liu, J., and Molineux, I.J. Structural dynamics of bacteriophage P22 infection initiation revealed by cryo-electron tomography. Nat Microbiol 4, 1049-1056, 2019
• Hu, B., Margolin, W., Molineux, I.J., and Liu, J. The bacteriophage t7 virion undergoes extensive structural remodeling during infection. Science 339, 576-579, 2013

The cytoplasmic domain of MxiG interacts with MxiK and directs assembly of the sorting platform in the Shigella type III secretion system.

Shoichi Tachiyama‡, Yunjie Chang§,¶, Meenakumari Muthuramalingam ‖ , Bo Hu**, Michael L. Barta ‖ 1, Wendy L. Picking‡‡, Jun Liu§,¶2 and William D. Picking‡,‡‡3

From Fig 6: Surface rendering images of Shigella T3SS. Left) The overall architecture of T3SS is described by four main components; the cytoplasmic sorting platform (cytoplasmic side), basal body (between bacterial inner membrane, IM, and outer membrane, OM), needle (extended from the basal body), and tip complex (end of the needle). Right) Cryo-ET provided a view from IM side of T3SS. The cytoplasmic sorting platform and basal body form different numbers of symmetrical rings .

The flagellar motor is a powerful biological nanomachine that drives motility, and thus infectivity and survival, in bacteria. It is the only known molecular machine that can rotate bidirectionally – in both clockwise (CW) and counterclockwise (CCW) senses. Motor rotation relies on the passage of ions through inner membrane-embedded stator units to power the cytoplasmic switch complex (C-ring), thus generating torque. Despite decades of intensive research, the detailed mechanisms that underlie torque generation and directional switching are unclear. I joined the Liu lab to use cryo-electron tomography (cryo-ET) to study this sophisticated mechanism in the model system of the Lyme disease-causing spirochete Borrelia burgdorferi. Our high-resolution analysis of dynamic stator-C-ring interactions will reveal molecular mechanisms responsible for torque generation and rotational switching in the bacterial flagellar motor. In addition, I currently oversee the operation of our state-of-the-art cryo-focused ion beam (FIB) microscope, Aquilos2, which enables extraordinary high-throughput imaging to address diverse biological questions.

Figure: Cryo-FIB milling of the Gram-negative bacterium Proteus in swarming conditions reveals the in-situ structure of the flagellar motor. (Left) Top view of scanning electron microscopy (SEM) image of Proteus swarming. (Right top) Side view of scanning electron microscopy (SEM) image of Proteus swarming. (Right bottom) Final lamella with 4^o tilt.

The pathogenic bacterium Legionella pneumophila causes Legionnaires' disease by injecting a diverse array of proteins into host cells through the Dot/Icm type IV secretion system (T4SS). This elaborate nanomachine transports over 300 distinct protein effectors or toxins into eukaryotic host cells. My research focuses on utilizing high-throughput cryo-electron tomography (cryo-ET) in combination with other cutting-edge approaches to visualize and determine high-resolution 3D structures of this intricate nanomachine to reveal its transport mechanisms. This work holds great promise in advancing our understanding of T4SS macromolecular machines and their vital roles in various biological processes. Furthermore, knowledge gained from this study will open new avenues for the development of drugs and therapies combatting a wide range of infectious diseases.

Image (Jian Yue, PhD): Construction of the structural model of the Legionella pneumophila Dot/Icm T4SS, including outer membrane-associated core complex (OMCC), plug, and inner membrane-associated complex (IMC)