Bench to Bedside: Rucks Laboratory uncovers elements of Chlamydial pathogenesis

‘Bench to Bedside’ is a new recurring segment exploring the basic science research conducted in infectious diseases right here at UNMC. This week, in recognition of STI Awareness Month, we feature the Rucks laboratory which studies chlamydial biology through the lens of infection. Special thanks to Natalie Sturd (left) and Dr. Rucks (right) for providing this content and sharing their work.

In case you missed it, see our previous Microbe Monday post on C. trachomatis for a primer on the pathogen behind the work discussed here.

What are some outstanding questions in the field of chlamydial biology and pathogenesis?

Until about a decade ago, C. trachomatis was a genetically intractable organism, meaning methods used by other researchers for studying bacteria were not available in our field. In basic science research, genetic manipulation is one of the major approaches we take when we want to study a pathogen. However, in the last 10 years, significant advances have been developed that allow transformation7-10. Though we are still playing “catch up” with research on organisms that have been genetically manipulated for several decades, advances in the chlamydial genetic toolbox have led to an explosion in our ability to interrogate the function of chlamydial proteins and effectors. Further, we can turn our attention to some questions in the field that have since remained unresolved. For example, it is not fully understood how C. trachomatis regulates alternating between its EB and RB forms. Also, with regards to the Inc proteins that were mentioned in this week’s Microbe Monday post, many of their individual functions haven’t been elucidated. Apart from their individual functions, Inc proteins have the potential to interact with each other, and their cooperative functions (and how this might expand their individual functions) aren’t currently appreciated. From the host perspective, how C. trachomatis can avoid detection and clearance by the host immune system during asymptomatic, chronic infections is also an active research topic.

What is our research focus in the Rucks lab? Any exciting new directions?

Broadly speaking, the focus of our lab is understanding the host-pathogen interactions of C. trachomatis, how these interactions are regulated, and identifying their impact on host signaling and/or trafficking pathways. As such, we must both examine 1) the microbiology of C. trachomatis and 2) what’s happening with the cell biology of the host cell. In the Rucks lab, there are several areas of active research, including 1) studying how Chlamydia manipulates host vesicle trafficking, 2) understanding proteomic turnover of Inc proteins, and 3) dissecting how Inc-host protein interactions change the interactomes and signaling pathways in host cells. Regarding that last point, a prior study in our lab used a proximity labeling system using APEX2 to identify host proteins found in the vicinity of the inclusion membrane11. This allowed us to cast a wide net to identify proteins recruited to the inclusion. Some of our exciting directions are following up on specific host proteins to investigate how Chlamydia might be recruiting them and altering host pathways. 

One of these studies that we are particularly excited about involves follow-up studies designed to better understand the function of two host proteins, LRRF1 and Flightless 1 (FLII/FLI1), that were identified in the proximity labeling study. Both proteins have been shown to be involved in regulating the host’s immune response. LRRF1 has been shown to be involved in innate immune signaling and FLII is a prominent regulator of actin dynamics, fibrosis, and wound healing. Knowing how these biological functions might relate to the reproductive tract pathology of chlamydial infections, we are particularly enthusiastic about investigating the outcome of the localization of these proteins to the inclusion membrane. Additionally, there are increasingly sophisticated co-culture tissue culture models of chlamydial infection, which are designed to better replicate the microenvironment of chlamydial infection.  We’re currently using a co-culture system in our lab, combining epithelial cells, fibroblasts, and different innate immune cells. 

In a separate line of research, we’re particularly curious about the cooperative functions of Inc proteins and how these might be temporally controlled throughout the chlamydial developmental cycle. Currently, in the field, it is typical to examine a single Inc at a single timepoint post-infection. This approach is reflective of the difficulty in studying an obligate intracellular pathogen. Improved genetic tools are helping us ask questions that we feel are more relevant to Chlamydia pathogenesis, as a successful infection is not likely beholden to a single Inc at a single timepoint post-infection. To test this question, we’ve been developing a whole cadre of new chlamydial strains that allow for the simultaneous knockdown of multiple Incs and the complementation of all or single Inc proteins.

So, while we may not be the ones treating chlamydial infections in the clinic, we are no less dedicated to improving reproductive health outcomes—we are simply coming at it from a different (very molecular) perspective!

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