Ehrlichia chaffeensis modulates numerous host cell processes, including gene transcription to promote
infection of the mononuclear phagocyte. Modulation of these host cell processes is directed through E. chaffeensis effectors, including TRP120. We previously reported that TRP120 moonlights as a HECT E3 Ub
ligase that ubiquitinates host cell transcription and fate regulators (PCGF5 and FBW7) to promote
infection. In this study, we identified a novel TRP120 substrate and examined the relationship between TRP120 and α-
enolase (ENO1), a metalloenzyme that catalyzes glycolytic pathway substrate
dehydration. Immunofluorescence microscopy and coimmunoprecipitation demonstrated interaction between ENO1 and TRP120, and ubiquitination of ENO-1 by TRP120 was detected in vivo and in vitro. Further, ENO-1 degradation was observed during
infection and was inhibited by the proteasomal inhibitor
bortezomib. A direct role of TRP120 Ub
ligase activity in ENO-1 degradation was demonstrated and confirmed by ectopic expression of TRP120 HECT Ub
ligase catalytic site mutant.
siRNA knockdown of ENO-1 coincided with increased E. chaffeensis
infection and ENO-1 knockdown disrupted glycolytic flux by decreasing the levels of
pyruvate and
lactate that may contribute to changes in host cell metabolism that promote
infection. In addition, we elucidated a functional role of TRP120 auto-ubiquitination as an activating event that facilitates the recruitment of the UbcH5
E2 ubiquitin-conjugating enzyme. This investigation further expands the repertoire of TRP120 substrates and extends the potential role of TRP120 Ub
ligase in
infection to include metabolic reprogramming.