Targeting of ELMO2 by the Shigella E3 Ligase IpaH4.5 Reveals Key Differences Between ELMO Paralogs

The IpaH class of effector proteins secreted by Shigella flexneri ubiquitinate specific immune factors, targeting them for proteasomal degradation. ELMO1-3 are key signaling molecules involved in cell motility and phagocytosis, but the distinct roles of these three proteins remain unclear. Here, we identify ELMO2 as a substrate of S. flexneri IpaH4.5 using an unbiased screening approach in human cell lysate. Using in vitro and cellular assays, we show that IpaH4.5 specifically targets ELMO2 for ubiquitination but has no activity against its paralogs ELMO1 and ELMO3. Using chimeric proteins, we show that IpaH4.5 specificity is dictated by divergence within the ELMO2 Armadillo Repeat Region, identifying this domain as a key determinant of ELMO paralog specialization. Consistent with this divergence, we find that commonly used ELMO antibodies lack paralog specificity and that ELMO2 is the predominant paralog expressed across multiple human cell lines. Further, functional assays demonstrate that IpaH4.5-mediated degradation of ELMO2 disrupts RAC activation downstream of the ELMO/DOCK complex, suggesting that this signaling pathway is perturbed during infection. We conclude that ELMO2 plays an as-yet unidentified role in counteracting bacterial infection and is therefore targeted by IpaH4.5.