SUMOylation, the covalent attachment of a member of the small ubiquitin-like 
modifier (SUMO) family of proteins to lysines in target substrates, is an 
essential post-translational modification in eukaryotes. Microbial manipulation 
of SUMOylation recently emerged as a key virulence strategy for viruses and 
facultative intracellular bacteria, the latter of which have only been shown to 
deploy effectors that negatively regulate SUMOylation. Here, we demonstrate that 
the obligate intracellular bacterium, Anaplasma phagocytophilum, utilizes an 
effector, AmpA (A. phagocytophilum post-translationally modified protein A) that 
becomes SUMOylated in host cells and this is important for the pathogen's 
survival. We previously discovered that AmpA (formerly APH1387) localizes to the 
A. phagocytophilum-occupied vacuolar membrane (AVM). Algorithmic prediction 
analyses denoted AmpA as a candidate for SUMOylation. We verified this 
phenomenon using a SUMO affinity matrix to precipitate both native AmpA and 
ectopically expressed green fluorescent protein (GFP)-tagged AmpA. SUMOylation 
of AmpA was lysine dependent, as SUMO affinity beads failed to precipitate a 
GFP-AmpA protein when its lysine residues were substituted with arginine. 
Ectopically expressed and endogenous AmpA were poly-SUMOylated, which was 
consistent with the observation that AmpA colocalizes with SUMO2/3 at the AVM. 
Only late during the infection cycle did AmpA colocalize with SUMO1, which 
terminally caps poly-SUMO2/3 chains. AmpA was also detected in the cytosol of 
infected host cells, further supporting its secretion and likely participation 
in interactions that aid pathogen survival. Indeed, whereas siRNA-mediated 
knockdown of Ubc9 - a necessary enzyme for SUMOylation - slightly bolstered 
A. phagocytophilum infection, pharmacologically inhibiting SUMOylation in 
infected cells significantly reduced the bacterial load. Ectopically expressed 
GFP-AmpA served as a competitive agonist against native AmpA in infected cells, 
while lysine-deficient GFP-AmpA was less effective, implying that modification 
of AmpA lysines is important for infection. Collectively, these data show that 
AmpA becomes directly SUMOylated during infection, representing a novel tactic 
for A. phagocytophilum survival.