Beatriz Montaner

Asymmetric Segregation of Polarized Antigen on B Cell Division Shapes Presentation Capacity

Antigen Polarity in B Cell Differentiation Communication received through cell contact is critical for the differentiation of specialized effector cell populations during the immune response. For example, B lymphocytes acquire antigen that they present to helper T lymphocytes. T lymphocytes, in turn, provide key differentiation signals to B lymphocytes. In order to learn more about this process, Thaunat et al. (p. 475; see the Perspective by Dustin and Meyer-Hermann) used multiphoton microscopy and imaging flow cytometry to visualize the localization of antigen in B lymphocytes during an immune response. Antigen acquired by B lymphocytes exhibited a polarized distribution that was sustained over several rounds of cell division. This produced a population of activated B lymphocytes that contained very low levels of antigen. Daughter cells that received more antigen were better able to stimulate T cells. Because cues received through T lymphocyte interactions are likely to influence B lymphocyte fate decisions, unequal distribution of antigen in dividing B lymphocytes may influence their differentiation. Antigen distribution across activated B cells influences B-T lymphocyte interactions. During the activation of humoral immune responses, B cells acquire antigen for subsequent presentation to cognate T cells. Here we show that after mouse B cells accumulate antigen, it is maintained in a polarized distribution for extended periods in vivo. Using high-throughput imaging flow cytometry, we observed that this polarization is preserved during B cell division, promoting asymmetric antigen segregation among progeny. Antigen inheritance correlates with the ability of progeny to activate T cells: Daughter cells receiving larger antigen stores exhibit a prolonged capacity to present antigen, which renders them more effective in competing for T cell help. The generation of progeny with differential capacities for antigen presentation may have implications for somatic hypermutation and class switching during affinity maturation and as B cells commit to effector cell fates.

Reactive oxygen-mediated damage to a human DNA replication and repair protein

Ultraviolet A (UVA) makes up more than 90% of incident terrestrial ultraviolet radiation. Unlike shorter wavelength UVB, which damages DNA directly, UVA is absorbed poorly by DNA and is therefore considered to be less hazardous. Organ transplant patients treated with the immunosuppressant azathioprine frequently develop skin cancer. Their DNA contains 6‐thioguanine—a base analogue that generates DNA‐damaging singlet oxygen (1O2) when exposed to UVA. Here, we show that this 1O2 damages proliferating cell nuclear antigen (PCNA), the homotrimeric DNA polymerase sliding clamp. It causes covalent oxidative crosslinking between the PCNA subunits through a histidine residue in the intersubunit domain. Crosslinking also occurs after treatment with higher—although still moderate—doses of UVA alone or with chemical oxidants. Chronic accumulation of oxidized proteins is linked to neurodegenerative disorders and ageing. Our findings identify oxidative damage to an important DNA replication and repair protein as a previously unrecognized hazard of acute oxidative stress.

Inflammation-induced disruption of SCS macrophages impairs B cell responses to secondary infection

Bacterial infection breaks the lymph node barrier During infections, lymph nodes are command central. Fragments from invading pathogens enter lymph nodes through the lymph. There, specialized cells called subcapsular sinus (SCS) macrophages capture these antigens and use them to initiate humoral immunity. Despite being such important players, Gaya et al. report that in mice, infection throws these organized sentinels into disarray (see the Perspective by Buzsaki). Disrupting SCS macrophages had important consequences: Bacterially infected mice could not respond as efficiently to a subsequent viral infection. Science, this issue p. 667; see also p. 612 Infection disrupts the organization of macrophages in the lymph node and impairs immunity to secondary infections. [Also see Perspective by Hickman] The layer of macrophages at the subcapsular sinus (SCS) captures pathogens entering the lymph node, preventing their global dissemination and triggering an immune response. However, how infection affects SCS macrophages remains largely unexplored. Here we show that infection and inflammation disrupt the organization of SCS macrophages in a manner that involves the migration of mature dendritic cells to the lymph node. This disrupted organization reduces the capacity of SCS macrophages to retain and present antigen in a subsequent secondary infection, resulting in diminished B cell responses. Thus, the SCS macrophage layer may act as a sensor or valve during infection to temporarily shut down the lymph node to further antigenic challenge. This shutdown may increase an organism’s susceptibility to secondary infections.

DNA breakage and cell cycle checkpoint abrogation induced by a therapeutic thiopurine and UVA radiation

The frequency of squamous cell skin carcinoma in organ transplant patients is around 100-fold higher than normal. This dramatic example of therapy-related cancer reflects exposure to sunlight and to immunosuppressive drugs. Here, we show that the interaction between low doses of UVA, the major ultraviolet component of incident sunlight, and 6-TG, a UVA chromophore that is introduced into DNA by one of the most widely prescribed immunosuppressive drugs, causes DNA single- and double-strand breaks (DSB). S phase cells are particularly vulnerable to this DNA breakage and cells defective in rejoining of S-phase DSB are hypersensitive to the combination of low-dose UVA and DNA 6-TG. 6-TG/UVA-induced DNA lesions provoke canonical DNA damage responses involving activation of the ATM/Chk2 and ATR/Chk1 pathways and appropriate cell cycle checkpoints. Higher levels of photochemical DNA damage induce a proteasome-mediated degradation of Chk1 and checkpoint abrogation that is consistent with persistent unrepaired DNA damage. These findings indicate that the interaction between UVA and an immunosuppressant drug causes photochemical DNA lesions, including DNA breaks, and can compromise cell cycle checkpoints. These two properties could contribute to the high risk of sunlight-related skin cancer in long-term immunosuppressed patients.

Increased Vascular Permeability in the Bone Marrow Microenvironment Contributes to Disease Progression and Drug Response in Acute Myeloid Leukemia

Summary The biological and clinical behaviors of hematological malignancies can be influenced by the active crosstalk with an altered bone marrow (BM) microenvironment. In the present study, we provide a detailed picture of the BM vasculature in acute myeloid leukemia using intravital two-photon microscopy. We found several abnormalities in the vascular architecture and function in patient-derived xenografts (PDX), such as vascular leakiness and increased hypoxia. Transcriptomic analysis in endothelial cells identified nitric oxide (NO) as major mediator of this phenotype in PDX and in patient-derived biopsies. Moreover, induction chemotherapy failing to restore normal vasculature was associated with a poor prognosis. Inhibition of NO production reduced vascular permeability, preserved normal hematopoietic stem cell function, and improved treatment response in PDX.

A switch from canonical to noncanonical autophagy shapes B cell responses

Change for good In the immune system, autophagy has been implicated in the maintenance and survival of plasma and memory cells, but its role in B cells during early viral infection remains unclear. Martinez-Martin et al. investigated the role of autophagy in B cells by using a combination of innovative imaging, pharmacological agents, and genetic models. B cell activation triggered an increase in the rate of autophagy and also switched the mechanism from canonical autophagy to noncanonical pathways involving the regulator WIPI2. Genetic ablation of WIPI2 in B cells promoted noncanonical autophagy. WIPI2 restrains noncanonical autophagy upon B cell activation through a mechanism involving mitochondrial status. Thus, the switch from canonical to noncanonical autophagy regulates B cell differentiation and fate during viral infection. Science, this issue p. 641 An unusual form of autophagy is triggered after immunological B cell activation and tunes B cell responses to viral infection. Autophagy is important in a variety of cellular and pathophysiological situations; however, its role in immune responses remains elusive. Here, we show that among B cells, germinal center (GC) cells exhibited the highest rate of autophagy during viral infection. In contrast to mechanistic target of rapamycin complex 1–dependent canonical autophagy, GC B cell autophagy occurred predominantly through a noncanonical pathway. B cell stimulation was sufficient to down-regulate canonical autophagy transiently while triggering noncanonical autophagy. Genetic ablation of WD repeat domain, phosphoinositide–interacting protein 2 in B cells alone enhanced this noncanonical autophagy, resulting in changes of mitochondrial homeostasis and alterations in GC and antibody-secreting cells. Thus, B cell activation prompts a temporal switch from canonical to noncanonical autophagy that is important in controlling B cell differentiation and fate.

Initiation of Antiviral B Cell Immunity Relies on Innate Signals from Spatially Positioned NKT Cells

Summary B cells constitute an essential line of defense from pathogenic infections through the generation of class-switched antibody-secreting cells (ASCs) in germinal centers. Although this process is known to be regulated by follicular helper T (TfH) cells, the mechanism by which B cells initially seed germinal center reactions remains elusive. We found that NKT cells, a population of innate-like T lymphocytes, are critical for the induction of B cell immunity upon viral infection. The positioning of NKT cells at the interfollicular areas of lymph nodes facilitates both their direct priming by resident macrophages and the localized delivery of innate signals to antigen-experienced B cells. Indeed, NKT cells secrete an early wave of IL-4 and constitute up to 70% of the total IL-4-producing cells during the initial stages of infection. Importantly, the requirement of this innate immunity arm appears to be evolutionarily conserved because early NKT and IL-4 gene signatures also positively correlate with the levels of neutralizing antibodies in Zika-virus-infected macaques. In conclusion, our data support a model wherein a pre-TfH wave of IL-4 secreted by interfollicular NKT cells triggers the seeding of germinal center cells and serves as an innate link between viral infection and B cell immunity.

The Small Rho GTPase TC10 Modulates B Cell Immune Responses

Rho family GTPases regulate diverse cellular events, such as cell motility, polarity, and vesicle traffic. Although a wealth of data exists on the canonical Rho GTPases RhoA, Rac1, and Cdc42, several other family members remain poorly studied. In B cells, we recently demonstrated a critical role for Cdc42 in plasma cell differentiation. In this study, we focus on a close homolog of Cdc42, TC10 (also known as RhoQ), and investigate its physiological role in B cells. By generating a TC10-deficient mouse model, we show that despite reduced total B cell numbers, B cell development in these mice occurs normally through distinct developmental stages. Upon immunization, IgM levels were reduced and, upon viral infection, germinal center responses were defective in TC10-deficient mice. BCR signaling was mildly affected, whereas cell migration remained normal in TC10-deficient B cells. Furthermore, by generating a TC10/Cdc42 double knockout mouse model, we found that TC10 can compensate for the lack of Cdc42 in TLR-induced cell activation and proliferation, so the two proteins play partly redundant roles. Taken together, by combining in vivo and in vitro analysis using TC10-deficient mice, we define the poorly studied Rho GTPase TC10 as an immunomodulatory molecule playing a role in physiological B cell responses.

Dynamic reorganisation of intermediate filaments coordinates early B-cell activation

This study examines the role of vimentin, a type III intermediate filament, in B-cell function using a combination of in vitro and in vivo assays, including super-resolution microscopic techniques. During B-cell activation, the dynamic reorganisation of the cytoskeleton is crucial for multiple cellular responses, such as receptor signalling, cell spreading, antigen internalisation, intracellular trafficking, and antigen presentation. However, the role of intermediate filaments (IFs), which represent a major component of the mammalian cytoskeleton, is not well defined. Here, by using multiple super-resolution microscopy techniques, including direct stochastic optical reconstruction microscopy, we show that IFs in B cells undergo drastic reorganisation immediately upon antigen stimulation and that this reorganisation requires actin and microtubules. Although the loss of vimentin in B cells did not impair B-cell development, receptor signalling, and differentiation, vimentin-deficient B cells exhibit altered positioning of antigen-containing and lysosomal associated membrane protein 1 (LAMP1+) compartments, implying that vimentin may play a role in the fine-tuning of intracellular trafficking. Indeed, vimentin-deficient B cells exhibit impaired antigen presentation and delayed antibody responses in vivo. Thus, our study presents a new perspective on the role of IFs in B-cell activation.