Multidomain proapoptotic BAX and BAK, once activated, permeabilize mitochondria to trigger apoptosis, whereas antiapoptotic BCL-2 members preserve mitochondrial integrity. The BH3-only molecules (BH3s) promote apoptosis by either activating BAX-BAK or inactivating antiapoptotic members. Here, we present biochemical and genetic evidence that NOXA is a bona fide activator BH3. Using combinatorial gain-of-function and loss-of-function approaches in Bid−/−Bim−/−Puma−/−Noxa−/− and Bax−/−Bak−/− cells, we have constructed an interconnected hierarchical model that accommodates and explains how the intricate interplays between the BCL-2 members dictate cellular survival versus death. BID, BIM, PUMA and NOXA directly induce stepwise, bimodal activation of BAX-BAK. BCL-2, BCL-XL and MCL-1 inhibit both modes of BAX-BAK activation by sequestering activator BH3s and “BH3-exposed” monomers of BAX-BAK, respectively. Furthermore, autoactivation of BAX and BAK can occur independently of activator BH3s through downregulation of BCL-2, BCL-XL and MCL-1. Our studies lay a foundation on targeting the BCL-2 family for treating diseases with dysregulated apoptosis.

Neurofibromatosis type 1 associates with multiple neoplasms, and the Schwann cell tumor neurofibroma is the most prevalent. A hallmark feature of neurofibroma is mast cell infiltration, which is recruited by chemoattractant stem cell factor (SCF) and has been suggested to sustain neurofibroma tumorigenesis. In the present study, we use new, genetically engineered Scf mice to decipher the contributions of tumor-derived SCF and mast cells to neurofibroma development. We demonstrate that mast cell infiltration is dependent on SCF from tumor Schwann cells. However, removal of mast cells by depleting the main SCF source only slightly affects neurofibroma progression. Other inflammation signatures show that all neurofibromas are associated with high levels of macrophages regardless of Scf status. These findings suggest an active inflammation in neurofibromas and partly explain why mast cell removal alone is not sufficient to relieve tumor burden in this experimental neurofibroma model. Furthermore, we show that plexiform neurofibromas are highly associated with injury-prone spinal nerves that are close to flexible vertebras. In summary, our study details the role of inflammation in neurofibromagenesis. Our data indicate that prevention of inflammation and possibly also nerve injury at the observed tumor locations are therapeutic approaches for neurofibroma prophylaxis and that such treatment should be explored.

Summary Neurofibromatosis type 1 is a tumor-predisposing genetic disorder. Plexiform neurofibromas are common NF1 tumors carrying a risk of malignant transformation, which is typically fatal. Little is known about mechanisms mediating initiation and identity of specific cell-type that gives rise to neurofibromas. Using cell-lineage tracing, we identify a population of GAP43+ PLP+ precursors in embryonic nerve roots as the cells of origin for these tumors and report a non-germline model of neurofibroma for preclinical drug screening to identify effective therapies. The identity of tumor cell-of-origin and facility for isolation and expansion provides fertile ground for continued analysis to define intrinsic and extrinsic factors critical for neurofibromagenesis. It also provides unique approaches to develop therapies to prevent neurofibroma formation in NF1 patients.

Hair differentiates from follicle stem cells through progenitor cells in the matrix. In contrast to stem cells in the bulge, the identities of the progenitors and the mechanisms by which they regulate hair shaft components are poorly understood. Hair is also pigmented by melanocytes in the follicle. However, the niche that regulates follicular melanocytes is not well characterized. Here, we report the identification of hair shaft progenitors in the matrix that are differentiated from follicular epithelial cells expressing transcription factor KROX20. Depletion of Krox20 lineage cells results in arrest of hair growth, confirming the critical role of KROX20 + cells as antecedents of structural cells found in hair. Expression of stem cell factor (SCF) by these cells is necessary for the maintenance of differentiated melanocytes and for hair pigmentation. Our findings reveal the identities of hair matrix progenitors that regulate hair growth and pigmentation, partly by creating an SCF-dependent niche for follicular melanocytes.

The innate immune response to Pneumocystis infection is not well understood. In this study, normal C57BL/6 mouse alveolar macrophages were found to respond to Pneumocystis murina organisms through Toll-like receptor 2 (TLR2), leading to the nuclear translocation of NF-B and the production of proinflammatory cytokine tumor necrosis factor alpha (TNF-␣) and chemokine macrophage inflammatory protein 2 (MIP-2). P. murina stimulation of normal alveolar macrophages from C57BL/6 mice resulted in increased TLR2 transcription but not increased TLR4 transcription. In gain-of-function studies with HEK293 cells expressing TLR2 or TLR4, only TLR2 was found to stimulate an NF-B response to P. murina. TNF-␣ and MIP-2 production in response to P. murina by mouse alveolar macrophages was inhibited by a monoclonal antibody that specifically blocked the ligand-binding ability of TLR2. Alveolar macrophages from TLR2 knockout (TLR2 ؊/؊ ) mice showed little increase in TNF-␣ and MIP-2 mRNA levels upon P. murina stimulation. An in vivo study showed that TLR2 ؊/؊ mice challenged with P. murina had reduced cytokine responses. These results indicate that TLR2 plays a major role in the innate immune response to P. murina.Pneumocystis is an opportunistic fungal pathogen of immunocompromised hosts. Pneumocystis organisms that infect humans are now called Pneumocystis jiroveci (41) and are a common cause of pneumonia in patients with AIDS (37). Pneumocystis carinii and Pneumocystis murina are those species that infect rats and mice, respectively (17,41). Although the use of highly active antiretroviral therapy has decreased the incidence of Pneumocystis pneumonia (PcP), the morbidity and mortality rates of PcP remain high (30). Patients who receive long-term immunosuppressive medications or who have genetic immunodeficiency are also susceptible to Pneumocystis infection.Effective immune responses of the host are required to control PcP. Alveolar macrophages play a critical role in the innate immune response to many microbes. Experimental evidence indicates that alveolar macrophages contribute to the host immune response to Pneumocystis organisms by directly ingesting or killing them and producing inflammatory cytokines and chemokines such as tumor necrosis factor alpha (TNF-␣) and macrophage inflammatory protein 2 (MIP-2) (14, 15, 54). The exact mechanism by which Pneumocystis stimulates alveolar macrophages to produce cytokines and chemokines is largely unknown.Microbes induce immune responses of host cells by interacting with certain cell surface receptors, termed pattern recognition receptors. Toll-like receptors (TLRs) have been identified as important pattern recognition receptors (28, 34) and are a large family with at least 11 members. Each TLR has a specific ligand(s) (44). TLR4 and its coreceptor, MD2, recognize lipopolysaccharides (LPSs) from gram-negative bacteria as well as many other ligands such as glucuronoxylomannan found in the polysaccharide capsule of Cryptococcus neoformans (38) and glycoinositolphospholipids from Trypano...