Galectin-8 C-terminal carbohydrate recognition domain is responsible for antigen uptake enhancement in dendritic cells

CECILIA ARAHI PRATO; LAURA VICTORIA BORBOLLA; OSCAR CAMPETELLA; MARÍA VIRGINIA TRIBULATTI

Mar del Plata

Congreso; Reunion anual de Sociedades de Biociencias; 2022

SAI

Galectin-8 (Gal-8) is a mammalian lectin capable of stimulate adaptive immune responses by acting on both CD4+ T cells and antigen-presenting cells. Recently, we found that Gal-8-glycan interaction at the dendritic cell surface results in antigen attachment and internalization, a crucial step during the initiation of a given immune response. Since Gal-8 possesses two carbohydrate recognition domains (N-CRD and C-CRD) with distinct fine specificity, we aim to characterize this interaction by analyzing the involvement of each isolated CRD and the requirement of the “dimeric” structure. We generated single N- and C-CRD recombinant proteins as well as chimeric proteins consisting of two covalently fused N-CRD (N-N) or C-CRD (C-C). Then, bone marrow-derived dendritic cells (BMDC) were incubated in the presence of either fluorescently labeled ovalbumin (OVA) together with each single domain (C-CRD or N-CRD), the equimolar mixture of both (C-CRD + N-CRD) or each chimera (C-C or N-N). OVA internalization was determined by flow cytometry.Remarkably, the only presence of C-CRD (alone, in mixture with N-CRD or in the chimera protein) was sufficient to recapitulate the Gal-8 effect on antigen internalization, indicating that the “dimeric” structure is not required. Moreover, the C-CRD (but not the N-CRD) pre-attachment to the BMDC surface also facilitated antigen internalization, an effect that was prevented by the addition of the Gal-8 inhibitor lactose. In accordance, BMDCs pulsed with OVA in the presence of isolated C-terminal domain efficiently activated cognate CD4+ T cells, as efficiently as native Gal-8. Taken together, these findings demonstrate that only the C-terminal domain participates in Gal-8-induced antigen internalization, providing a new insight to advance in the use of Gal-8 as an adjuvant in vaccine formulations.