B Cell Activation: Roles of AMP-activated Protein Kinase, Liver Kinase B1, and Metabolism in Establishing Humoral Immune Responses
- Author(s): Waters, Lynnea Rae
- Advisor(s): Teitell, Michael A
- et al.
B cells are a core component of the adaptive immune system that make specific antibodies to fight off infections. When a B cell recognizes an antigen they become activated and can differentiate into antibody-secreting plasma cells in a process called the germinal center reaction. The signaling pathways involved in this activation are not well characterized. Previously, our lab discovered a pathway induced by DNA double strand breaks that signals through ATM, LKB1, and an AMPK family member to phosphorylate the transcriptional coactivator CRTC2, which leaves the nucleus allowing for plasma cell differentiation. In this dissertation, we investigated first the role of LKB1, then the role of AMPK during B cell activation. Finally, spurred by metabolic functions of AMPK, we identified key metabolic pathways that regulate B cell activation.
LKB1 is a serine/threonine kinase, a tumor suppressor, and the genetic cause of Peutz-Jeghers syndrome. B-cell specific knockout of LKB1 (BKO) resulted in spontaneous germinal center formation in the absence of antigenic stimulation and a 100 fold increase in the number of germinal center B cells. BKO mice exhibit splenomegaly driven by an increase in T cell numbers, likely driven by LKB1 knockout B cells that exhibit an increase in production of activating cytokinesand chemokines. Additionally, we show that loss of LKB1 in B cells results in activation of NF-κB, which controls the expression of IL-6, driving TFH differentiation. For this reason, we believe that Lkb1 acts as a regulator of GC entry. Contextualizing these findings with previous reports that LKB1 is also required for GC exit and plasma cell differentiation, we posit that LKB1 is a critical regulator of the GC reaction by controlling both entry and exit.
The main target of LKB1 is AMPK, which is another understudied signaling pathway in B cell activation. Here, we report surprising AMPK activation upon B cell stimulation in vitro in the absence of energy stress coupled with rapid biomass accumulation. An IP-MS substrate screen to identify AMPK targets revealed only seven canonical AMPK target proteins. Despite AMPK activation and a controlling role for LKB1 in B cell activation, knockout of AMPK did not significantly affect B cell activation, differentiation, nutrient dynamics, gene expression, or humoral immune responses. Instead, AMPK loss specifically repressed the transcriptional expression of IgD and its regulator, Zfp318.
Finally, studies of the metabolic regulator AMPK prompted dissection of the metabolic reprogramming B cells undergo upon activation. We found that B cells upregulate glucose uptake, but not glycolysis, and undergo a switch to OXPHOS. Isotopomer tracing showed that glucose was not being utilized in the TCA cycle but was instead shunted towards nucleotide and lipid biosynthesis. Finally, we showed that mitochondria undergo major remodeling upon stimulation. In summary, these findings reveal critical roles for novel signaling pathways and metabolic regulation during B cell activation.