: First encounter with thymus-dependent antigen usually induces both a primary (1 primary) antibody response and B cell memory, thus providing a rationale for vaccination. However, the dichotomy in response to epitopes on (Tyr, Glu)-Ala--Lys [(T,G)-A--L] shows that B cell activation leading to the antibody forming cell (AFC) vs. memory pathways are separable events: the 1 primary antibody response is against GT, but upon memory recall, 20- 30% of the B cells respond to A-L, thus (T,G)-A--L-priming generated memory to A-L with no detectable early 1 primary effector phase. The long-term objectives are to understand the mechanisms that control cell fate determination (effector vs. memory) after 1 degree B cell activation. The proposed experiments will test the hypothesis that higher avidity 1 degree B cells (e. g., GT+) are activated to become AFC, whereas lower avidity B cells (e. g., A-L+) are activated only to enter the germinal center to memory pathway. V(D)J knock-in mice will be used to follow the cell fate of higher vs. lower avidity 1 primary B cells. The GT knock-in expresses a heavy chain from a 1 degree GT+ B cell. Two A-L+ knock-ins (AL-M, collectively) express mutated heavy chains from an A-L+ memory cell, whereas the AL-GL knock-in uses the same, but unmutated, heavy chain.
Aim 1 asks whether B cells differing in avidity have different cell fate outcomes after in vivo activation, and it will analyze different aspects of AFC or memory cell formation by GT, AL-M, and AL-GL 1 primary B cells using flow cytometry and adoptive transfer experiments.
Aim 2 asks whether in vitro correlates exist for differential signaling by BCRs differing in avidity, and it will analyze B cell antigen presentation ability, activation profiles, and BCR translocation to lipid rafts. If, as this hypothesis proposes, the signal threshold for memory cell generation is lower than that for AFC differentiation, then modifying strength of signal might modulate cell fate.
Aim 3 asks whether reducing CD22 negative regulation allows lower avidity cells to become AFC, and whether reducing Btk signals impedes AFC differentiation of higher avidity B cells. It will compare effector vs. memory cell generation in GT, AL-M, and AL-GL knock-ins on the CD22 -/- or .xid background. Understanding the relationships among B cell avidity, B cell activation signals, and B cell fate (effector vs. memory) will help in the design of vaccines that can elicit both protective, early 1 primary antibody and B cell memory after one priming event.
