The K134 and R234 residues are involved in degradation of JAM1 by P. gingivalis in IHGE cells.
(A) Schematic view of the JAM1 structure and derivatives. SP, IG-LIKE, or TMD domains are indicated by gray boxes. HA-tag is shown in green. The point mutations K134H and R234H are shown in magenta. (B) IHGE cells were transiently transfected with plasmid encoding HA-inserted JAM1 or the indicated JAM1 mutants. Following 48 h of incubation, the cells were infected with P. gingivalis at an MOI of 100 for 1 h, and then analyzed by immunoblotting using the indicated antibodies. (C) IHGE cells stably expressing HA-tagged JAM1 Δ (1–133) or HA-tagged JAM1 Δ (1–133) K134H R234H were infected for 1 h with P. gingivalis WT or the Δkgp ΔrgpA ΔrgpB mutant at an MOI of 100. The cells were then fixed, stained with DAPI (cyan) and anti-HA (yellow), and analyzed by immunofluorescence microscopy. Scale bars, 10 μm.