Cell surface ADP-ribosyltransferase regulates lymphocyte function-associated molecule-1 (LFA-1) function in T cells

Post-translational modifications are known to regulate intracellular signaling proteins. However, their role in extracellular domains of cell surface proteins remains poorly understood. Prior research has shown that intracellular modifications influence protein function, but no prior work had resolved how these processes might affect extracellular domains. This gap motivated recent studies on cell surface enzymes like mono-ADP-ribosyltransferase (ADPRT). No prior work had resolved the specific targets of ADPRT on immune cell surfaces. The role of ADPRT in T cell signaling was previously unknown. This uncertainty drove investigations into whether ADPRT modifies adhesion molecules like LFA-1. No prior work had resolved how ADPRT affects LFA-1 epitopes or signaling. The connection between ADPRT and T cell cytolytic activity was also unexplored.

The aim of this study was to determine whether cell surface ADPRT modifies LFA-1 and how this affects T cell function. LFA-1 is a key adhesion molecule involved in T cell interactions. The researchers sought to identify if ADPRT modifies both alpha and beta chains of LFA-1. They also aimed to assess how ADPRT activity influences LFA-1 signaling. The study focused on whether ADPRT inhibition affects T cell adhesion and proliferation. The researchers wanted to clarify if ADPRT regulates LFA-1 through ADP-ribose modification. They also sought to determine if ADPRT activity is necessary for LFA-1 epitope recognition. The specific problem addressed was the unknown mechanism of LFA-1 regulation by ADPRT.

The study used T cells expressing cell surface ADPRT and compared them to cells lacking ADPRT. Researchers applied phospholipase C to remove ADPRT from the cell surface. They used phosphodiesterase inhibitors like ADP-ribose to block ADPRT activity. The team labeled ADP-ribose modifications on LFA-1 alpha and beta chains. They assessed epitope recognition using alpha- and beta-chain-specific antibodies. Inositol phosphate generation was measured after antibody cross-linking of LFA-1. Homotypic cell adhesion was tested using anti-LFA-1 antibodies. Transfection experiments introduced ADPRT into cells lacking it to confirm its role.

LFA-1 was identified as a major target of ADPRT modification. Both alpha and beta chains of LFA-1 were ADP-ribosylated on extracellular domains. Labeling persisted longer on the alpha-chain than the beta-chain. Phosphodiesterase inhibitors suppressed label loss from the beta-chain. Modification of LFA-1 caused loss of epitopes recognized by chain-specific antibodies. Inositol phosphate generation after LFA-1 cross-linking was significantly inhibited. Anti-LFA-1-induced homotypic adhesion was also reduced in modified cells. Transfected cells gained ADPRT-dependent adhesion inhibition.

The authors concluded that cell surface ADPRT regulates LFA-1 functions. The enzyme modifies both chains of LFA-1 but with different persistence. ADPRT activity suppresses epitope recognition by chain-specific antibodies. Inositol phosphate generation is inhibited following LFA-1 modification. Homotypic cell adhesion is reduced when LFA-1 is modified by ADPRT. Cells lacking ADPRT do not show these effects. Transfection with ADPRT restored adhesion inhibition. These findings support the role of ADPRT in modulating T cell adhesion and signaling.