Murine macrophages surface N-glycans participate in YopP-mediated immunopathogenesis of Yersinia enterocolitica infections

JUAN AGUSTÍN GARAY; JUAN EDUARDO SILVA; JOSÉ VARAS; SAMANTA FUNES; BRENDA JOFRÉ; MARÍA SILVIA DI GENARO; ROBERTO CARLOS DAVICINO

Mar del Plata

Congreso; Reunión Conjunta SAIC SAI&FAIC SAFIS 2022; 2022

Sociedades científicas SAIC, SAI&FAIC, SAFIS

Yersinia enterocolitica (Ye) outer membrane protein P (YopP) is able to manipulate macrophages (MΦ) by inducing apoptosis and suppressing signaling molecules production. Although YopP has been shown to interact in a carbohydrate-dependent manner with host proteins, its interaction with glycans remains unclear. The aim of our work is to explore the influence of MΦ surface N-glycans on the immunomodulatory effects triggered by YopP. Murine peritoneal MΦ were obtained by intraperitoneal lavage, plated at a 1x106 cells/mL and purified by incubation for 2 h at 37°C in DMEM medium in an atmosphere with 5% CO2. Then, MΦ were incubated with 5 IU/mL of PNGase F in DMEM medium without Fetal Bovine Serum for 4 h (MΦd) and cultured overnight in supplemented DMEM medium. Subsequently, MΦ were infected with Ye serotype 0:8 (pYV+, WA-314) (Ye wt) or with Ye WA-314 deficient in YopP (pYV+, WA-C pYVNalrKanr) (Ye ∆yopP) at MOI 10:1. Then, MTT, lactate dehydrogenase (LDH), Griess and Urease assays were performed. Apoptosis was evaluated by flow cytometry using Annexin V-propidium iodide, and TNF-α was measured by ELISA. Our results shows that N-deglycosylation does not affect viability. Besides, we observed no significant apoptosis in Ye wt-infected MΦd compared to uninfected counterpart, suggesting a critical role of N-glycans in the well-known triggering role of YopP on apoptosis. On the other hand, we showed that YopP has a dual effect: in MΦ inhibits NO production (p