ICOS costimulation is indispensable for the differentiation of T follicular regulatory cells.

ICOS is a T-cell costimulatory receptor critical for Tfh cell generation and function. However, the role of ICOS in Tfr cell differentiation remains unclear. Using Foxp3-Cre-mediated ICOS knockout (ICOS FC) mice, we show that ICOS deficiency in Treg-lineage cells drastically reduces the number of Tfr cells during GC reactions but has a minimal impact on conventional Treg cells. Single-cell transcriptome analysis of Foxp3+ cells at an early stage of the GC reaction suggests that ICOS normally inhibits Klf2 expression to promote follicular features including Bcl6 up-regulation. Furthermore, ICOS costimulation promotes nuclear localization of NFAT2, a known driver of CXCR5 expression. Notably, ICOS FC mice had an unaltered overall GC B-cell output but showed signs of expanded autoreactive B cells along with elevated autoantibody titers. Thus, our study demonstrates that ICOS costimulation is critical for Tfr cell differentiation and highlights the importance of Tfr cells in maintaining humoral immune tolerance during GC reactions.

Hippo Signal Transduction Mechanisms in T Cell Immunity.

Hippo signaling pathways are evolutionarily conserved signal transduction mechanisms mainly involved in organ size control, tissue regeneration, and tumor suppression. However, in mammals, the primary role of Hippo signaling seems to be regulation of immunity. As such, humans with null mutations in STK4 (mammalian homologue of Drosophila Hippo; also known as MST1) suffer from recurrent infections and autoimmune symptoms. Although dysregulated T cell homeostasis and functions have been identified in MST1-deficient human patients and mouse models, detailed cellular and molecular bases of the immune dysfunction remain to be elucidated. Although the canonical Hippo signaling pathway involves transcriptional co-activator Yes-associated protein (YAP) or transcriptional coactivator with PDZ motif (TAZ), the major Hippo downstream signaling pathways in T cells are YAP/TAZ-independent and they widely differ between T cell subsets. Here we will review Hippo signaling mechanisms in T cell immunity and describe their implications for immune defects found in MST1-deficient patients and animals. Further, we propose that mutual inhibition of Mst and Akt kinases and their opposing roles on the stability and function of forkhead box O and ß-catenin may explain various immune defects discovered in mutant mice lacking Hippo signaling components. Understanding these diverse Hippo signaling pathways and their interplay with other evolutionarily-conserved signaling components in T cells may uncover molecular targets relevant to vaccination, autoimmune diseases, and cancer immunotherapies.

Progression of AITL-like tumors in mice is driven by Tfh signature proteins and T-B cross talk.

Angioimmunoblastic T-cell lymphoma (AITL) is an aggressive peripheral T-cell lymphoma driven by a pool of neoplastic cells originating from T follicular helper (Tfh) cells and concomitant expansion of B cells. Conventional chemotherapies for AITL have shown limited efficacy, and as such, there is a need for improved therapeutic options. Because AITL originates from Tfh cells, we hypothesized that AITL tumors continue to rely on essential Tfh components and intimate T-cell-B-cell (T-B) interactions. Using a spontaneous AITL mouse model (Roquinsan/+ mice), we found that acute loss of Bcl6 activity in growing tumors drastically reduced tumor size, demonstrating that AITL-like tumors critically depend on the Tfh lineage-defining transcription factor Bcl6. Because Bcl6 can upregulate expression of signaling lymphocytic activation molecule-associated protein (SAP), which is known to promote T-B conjugation, we next targeted the SAP-encoding Sh2d1a gene. We observed that Sh2d1a deletion from CD4+ T cells in fully developed tumors also led to tumor regression. Further, we provide evidence that tumor progression depends on T-B cross talk facilitated by SAP and high-affinity LFA-1. In our study, AITL-like tumors relied heavily on molecular pathways that support Tfh cell identity and T-B collaboration, revealing potential therapeutic targets for AITL.

Inducible T-cell co-stimulator: Signaling mechanisms in T follicular helper cells and beyond.

Human patients with homozygous null mutations in the ICOS gene suffer from recurrent infections due to humoral immune defects. Studies on human patients and mouse models have shown that inducible T-cell co-stimulator (ICOS)-deficient individuals cannot form T follicular helper (Tfh) cells, a group of CD4 T cells that migrate into B cell follicles and facilitate germinal center (GC) reactions. ICOS-induced phosphoinositide 3-kinase signaling pathways have been shown to play critical roles in Tfh programming, migration of Tfh cells into the GC, and delivery of T cell help during Tfh-GC B cell conjugation. These processes are also assisted by ICOS-mediated intracellular calcium mobilization and TANK-binding kinase 1 signaling. However, ICOS signaling also has stimulatory roles in T regulatory cells and innate lymphoid cells (ILCs), providing another layer of complexity. In this review, we discuss cell-type-specific signaling mechanisms utilized by ICOS in Tfh cells, T regulatory cells, and ILCs. Whenever relevant, we compare the roles and signaling pathways of ICOS and CD28. Understanding ICOS signal transduction mechanisms used by distinct immune subsets at different stages of immune responses or disease progression may help improve vaccination protocols, treat autoimmune diseases, and enhance cancer immunotherapy.

Hippo Pathway Kinase Mst1 Is Required for Long-Lived Humoral Immunity.

The protein kinase Mst1 is a key component of the evolutionarily conserved Hippo pathway that regulates cell survival, proliferation, differentiation, and migration. In humans, Mst1 deficiency causes primary immunodeficiency. Patients with MST1-null mutations show progressive loss of naive T cells but, paradoxically, mildly elevated serum Ab titers. Nonetheless, the role of Mst1 in humoral immunity remains poorly understood. In this study, we found that early T cell-dependent IgG1 responses in young adult Mst1-deficient mice were largely intact with signs of impaired affinity maturation. However, the established Ag-specific IgG1 titers in Mst1-deficient mice decayed more readily because of a loss of Ag-specific but not the overall bone marrow plasma cells. Despite the impaired affinity and longevity of Ag-specific Abs, Mst1-deficient mice produced plasma cells displaying apparently normal maturation markers with intact migratory and secretory capacities. Intriguingly, in immunized Mst1-deficient mice, T follicular helper cells were hyperactive, expressing higher levels of IL-21, IL-4, and surface CD40L. Accordingly, germinal center B cells progressed more rapidly into the plasma cell lineage, presumably forgoing rigorous affinity maturation processes. Importantly, Mst1-deficient mice had elevated levels of CD138+Blimp1+ splenic plasma cell populations, yet the size of the bone marrow plasma cell population remained normal. Thus, overproduced low-affinity plasma cells from dysregulated germinal centers seem to underlie humoral immune defects in Mst1-deficiency. Our findings imply that vaccination of Mst1-deficient human patients, even at the early stage of life, may fail to establish long-lived high-affinity humoral immunity and that prophylactic Ab replacement therapy can be beneficial to the patients.

ICOS Signaling Controls Induction and Maintenance of Collagen-Induced Arthritis.

ICOS is a key costimulatory receptor facilitating differentiation and function of follicular helper T cells and inflammatory T cells. Rheumatoid arthritis patients were shown to have elevated levels of ICOS+ T cells in the synovial fluid, suggesting a potential role of ICOS-mediated T cell costimulation in autoimmune joint inflammation. In this study, using ICOS knockout and knockin mouse models, we found that ICOS signaling is required for the induction and maintenance of collagen-induced arthritis (CIA), a murine model of rheumatoid arthritis. For the initiation of CIA, the Tyr181-based SH2-binding motif of ICOS that is known to activate PI3K was critical for Ab production and expansion of inflammatory T cells. Furthermore, we found that Tyr181-dependent ICOS signaling is important for maintenance of CIA in an Ab-independent manner. Importantly, we found that a small molecule inhibitor of glycolysis, 3-bromopyruvate, ameliorates established CIA, suggesting an overlap between ICOS signaling, PI3K signaling, and glucose metabolism. Thus, we identified ICOS as a key costimulatory pathway that controls induction and maintenance of CIA and provide evidence that T cell glycolytic pathways can be potential therapeutic targets for rheumatoid arthritis.

MYSM1-dependent checkpoints in B cell lineage differentiation and B cell-mediated immune response.

MYSM1 is a chromatin-binding histone deubiquitinase. MYSM1 mutations in humans result in lymphopenia whereas loss of Mysm1 in mice causes severe hematopoietic abnormalities, including an early arrest in B cell development. However, it remains unknown whether MYSM1 is required at later checkpoints in B cell development or for B cell-mediated immune responses. We analyzed conditional mouse models Mysm1fl/flTg.mb1-cre, Mysm1fl/flTg.CD19-cre, and Mysm1fl/flTg.CD21-cre with inactivation of Mysm1 at prepro-B, pre-B, and follicular B cell stages of development. We show that loss of Mysm1 at the prepro-B cell stage in Mysm1fl/flTg.mb1-cre mice results in impaired B cell differentiation, with an ∼2-fold reduction in B cell numbers in the lymphoid organs. Mysm1fl/flTg.mb1-cre B cells also showed increased expression of activation markers and impaired survival and proliferation. In contrast, Mysm1 was largely dispensable from the pre-B cell stage onward, with Mysm1fl/flTg.CD19-cre and Mysm1fl/flTg.CD21-cre mice showing no alterations in B cell numbers and largely normal responses to stimulation. MYSM1, therefore, has an essential role in B cell lineage specification but is dispensable at later stages of development. Importantly, MYSM1 activity at the prepro-B cell stage of development is important for the normal programming of B cell responses to stimulation once they complete their maturation process.