Dissecting the Antibody Cleavage System of Mycoplasmas – DACSyMy

Mycoplasmas are parasitic bacteria that are able to infect the majority of livestock animals, including cattle, goat, sheep, swine and poultry. The infected animal exhibit reduced production yields (loss of weight, limited milk and egg production), causing important economic losses to the farming industry. Current treatment and prevention strategies are considered relatively inefficient, and our ability to combat these diseases is hampered by our limited knowledge of mycoplasma pathogenicity. Only a small number of virulence factors have been characterized and data are relatively scarce on the mechanisms deployed by these bacteria to evade the host immune system.
Addressing this lack of knowledge is essential to foster the development of new and more efficient prophylaxis and treatment strategies. Recently, our group has discovered a novel mycoplasma virulence factor, based on two surface proteins, termed MiB (Mycoplasma Immunoglobulin Binding) and MiP (Mycoplasma Immunoglobulin Protease). MiB and MiP act in coordination to capture and inactivate the host antibodies by cleaving off their VH domains.
Interestingly, the genes encoding MiB and MiP are part of a large operon containing multiple variants of MiB and MiP, and seven genes encoding an atypical F1-like X0 ATPase whose function is currently unknown. This “MiB-MiP-ATPase” operon is highly conserved throughout the genus Mycoplasma, and is found in almost all the species that infect livestock animals.
Given the “antibody killer” role of MiB-MiP, and the widespread distribution of the “MiB-MiP-ATPase” operon, we proposed that this system could be key for the ability of mycoplasmas to evade the host immunity. The data we have gathered to this day have allowed us to propose a basic functional model, but we lack insight on several key aspects of this system. In particular, we are asking three main scientific questions:
1. What is the exact molecular mechanism of the antibody cleavage?
2. The F1-like X0 ATPase is genetically linked to MiB-MiP, but is it involved in the antibody cleavage?
3. Why are there multiple variants of MiB and MiP in a genome, and are they functionally different?
To answer these questions, we propose to dissect this system using three complementary approaches:
i. Biochemical and functional characterization of all the proteins encoded in the operon
ii. Resolution of the 3D structure of MiB, MiP and the F1-like X0 ATPase by crystallography and cryoEM.
iii. Analysis of the MiB-MiP-ATPase system in cellulo by super-resolution microscopy
The results of the DACSyMy project will shed a new light on the relationship of mycoplasmas with their hosts, and allow us to better understand how these bacteria escape the immune system. On the long term, we anticipate that this improved fundamental knowledge of mycoplasma virulence will pave the way for the development of new prevention and treatment strategies against mycoplasma diseases in farm animals