The ubiquitin ligase CBL and Fas-associated factor 2 cooperate to regulate the innate immune response to M. tuberculosis.

As a first line of host defense, macrophages must be able to effectively sense and respond to diverse types of pathogens, and while a particular response may be beneficial in some circumstances, it can be detrimental in others. Upon infection, Mycobacterium tuberculosis (Mtb) induces proinflammatory cytokines and activates antibacterial responses. Surprisingly, Mtb also triggers antiviral responses that actually hinder the ability of macrophages to restrict Mtb growth. In Mtb-infected macrophages, the ubiquitin ligase CBL suppresses antiviral responses and preserves the antibacterial capacity of the macrophage. However, the mechanisms by which CBL regulates immune signaling are unknown. We found that CBL controls responses to multiple immune stimuli and broadly suppresses the expression of antiviral response genes. We used mass spectrometry to identify potential CBL substrates, and found, in total, over 46,000 ubiquitylated peptides in Mtb-infected macrophages, including roughly 400 peptides with CBL-dependent ubiquitylation. We then performed genetic interaction analysis of CBL and its putative substrates, and identified the Fas-associated factor 2 (FAF2) adapter protein as a key signaling molecule downstream of CBL. Together, these analyses reveal thousands of previously uncharacterized ubiquitin-mediated signaling events and identify an important new regulator of immune signaling.