The Journal of Experimental Medicine

Flomerfelt, Francis A.; El Kassar, Nahed; Gurunathan, Chandra; Chua, Kevin S.; League, Stacy C.; Schmitz, Sabrina; Gershon, Timothy R.; Kapoor, Veena; Yan, Xiao-Yi; Schwartz, Ronald H.; Gress, Ronald E.

doi: 10.1084/jem.20092759pmid: 20937703

Niche availability provided by stromal cells is critical to thymus function. Thymi with diminished function contain fewer stromal cells, whereas thymi with robust function contain proliferating stromal cell populations. Here, we show that the thymus, brain, and testes–associated gene ( Tbata ; also known as SPATIAL) regulates thymic epithelial cell (TEC) proliferation and thymus size. Tbata is expressed in thymic stromal cells and interacts with the enzyme Uba3, thereby inhibiting the Nedd8 pathway and cell proliferation. Thymi from aged Tbata -deficient mice are larger and contain more dividing TECs than wild-type littermate controls. In addition, thymic reconstitution after bone marrow transplantation occurred more rapidly in Rag2 −/− Tbata −/− mice than in Rag2 −/− Tbata +/+ littermate controls. These findings suggest that Tbata modulates thymus function by regulating stromal cell proliferation via the Nedd8 pathway. Footnotes Abbreviations used: AppBp1 amyloid protein precursor binding protein 1 BMT BM transplantation EGL external germinal layer mTEC medullary TEC Nedd8 neural precursor cell–expressed developmentally down-regulated 8 Tbata thymus, brain, and testes–associated gene TEC thymic epithelial cell Ttv Tbata transcript variant Submitted: 24 December 2009 Accepted: 7 September 2010 This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms ). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/ ).

Pareek, Tej K.; Lam, Eric; Zheng, Xiaojing; Askew, David; Kulkarni, Ashok B.; Chance, Mark R.; Huang, Alex Y.; Cooke, Kenneth R.; Letterio, John J.

doi: 10.1084/jem.20100876pmid: 20937706

Cyclin-dependent kinase 5 (Cdk5) is a ubiquitously expressed serine/threonine kinase. However, a requirement for Cdk5 has been demonstrated only in postmitotic neurons where there is abundant expression of its activating partners p35 and/or p39. Although hyperactivation of the Cdk5–p35 complex has been found in a variety of inflammatory neurodegenerative disorders, the potential contribution of nonneuronal Cdk5–p35 activity has not been explored in this context. We describe a previously unknown function of the Cdk5–p35 complex in T cells that is required for induction of experimental autoimmune encephalomyelitis (EAE). T cell receptor (TCR) stimulation leads to a rapid induction of Cdk5–p35 expression that is required for T lymphocyte activation. Chimeric mice lacking Cdk5 gene expression in hematopoietic tissues (Cdk5 −/−C ) are resistant to induction of EAE, and adoptive transfer of either Cdk5 −/−C or p35 −/− encephalitogenic lymphocytes fails to transfer disease. Moreover, our data reveal a novel mechanism involving Cdk5-mediated phosphorylation of the actin modulator coronin 1a on threonine 418. Cdk5-deficient lymphocytes lack this posttranslational modification of coronin 1a and exhibit defective TCR-induced actin polarization and reduced migration toward CCL-19. These data define a distinct role for Cdk5 in lymphocyte biology and suggest that inhibition of this kinase may be beneficial in the treatment of T cell–mediated inflammatory disorders. Footnotes Abbreviations used: ANOVA analysis of variance Cdk5 cyclin-dependent kinase 5 CNS central nervous system EAE experimental autoimmune encephalomyelitis IS immune synapse MOG myelin oligodendrocyte glycoprotein MPM-2 mitotic protein monoclonal 2 MS multiple sclerosis NF-H neurofilament H PTX pertussis toxin Submitted: 3 May 2010 Accepted: 20 September 2010 This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms ). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/ ).

Fang, Min; Roscoe, Felicia; Sigal, Luis J.

doi: 10.1084/jem.20100282pmid: 20876312

Although it is well known that aged hosts are generally more susceptible to viral diseases than the young, specific dysfunctions of the immune system directly responsible for this increased susceptibility have yet to be identified. We show that mice genetically resistant to mousepox (the mouse parallel of human smallpox) lose resistance at mid-age. Surprisingly, this loss of resistance is not a result of intrinsically defective T cell responses. Instead, the primary reason for the loss of resistance results from a decreased number of total and mature natural killer (NK) cells in the blood and an intrinsic impairment in their ability to migrate to the lymph node draining the site of infection, which is essential to curb systemic virus spread. Hence, our work links the age-dependent increase in susceptibility to a viral disease to a specific defect of NK cells, opening the possibility of exploring treatments to improve NK cell function in the aged with the goal of enhancing their resistance to viral diseases. Footnotes Abbreviations used: Ab antibody D-LN draining LN dpi days post infection ECTV Ectromelia virus GzB granzyme B ND-LN non–D-LN OPV Orthopoxvirus VACV vaccinia virus Submitted: 9 February 2010 Accepted: 1 September 2010 This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms ). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/ ).

Weiss, Jonathan M.; Ridnour, Lisa A.; Back, Tim; Hussain, S. Perwez; He, Peijun; Maciag, Anna E.; Keefer, Larry K.; Murphy, William J.; Harris, Curtis C.; Wink, David A.; Wiltrout, Robert H.

doi: 10.1084/jem.20100670pmid: 20921282

Using an orthotopic model of renal cell carcinoma, we showed previously that IL-2/anti-CD40 immunotherapy resulted in synergistic anti-tumor responses, whereas IL-2 or α-CD40 alone mediated partial transient anti-tumor effects. We now show that treatment of tumor-bearing mice with IL-2/α-CD40, but not IL-2 or α-CD40, induced significant nitric oxide synthase (NOS) 2 expression in tumor-associated macrophages. In control-treated mice (low NO), NOS2 inhibition reduced tumor burden. However, during immunotherapy (high NO), NOS2 inhibition or macrophage depletion reversed the ability of IL-2/α-CD40 treatment to reduce lung metastases but had no effect on primary tumor burden. Furthermore, IL-2/α-CD40 induced the IFN-γ– and NO-dependent decrease in matrix metalloproteinase (MMP) expression and activity, concomitant with increases in tissue inhibitor of metalloproteinase (TIMP) 1 and E-cadherin expression within tumors. Finally, treatment of tumor-bearing mice with the NO donor JS-K significantly reduced metastases. These data differentiate the mechanism for primary anti-tumor effects of IL-2/α-CD40 immunotherapy, which are independent of NO, from the NO-dependent inhibition of metastases. Furthermore, reduced MMP9 activity implicates M1-polarized macrophages within the tumor microenvironment as critical components of therapeutic response. Our data demonstrate the mechanistic basis for IL-2/α-CD40–mediated control of metastases and suggest that the context-dependent application of NO donors may hold promise for prevention of metastatic disease. Footnotes Abbreviations used: GKO IFN-γ knockout HPRT hypoxanthine phosphoribosyltransferase L-NAME N ω -nitro- l -arginine methyl ester MDSC myeloid-derived suppressor cell MMP matrix metalloproteinase NOS NO synthase RCC renal cell carcinoma TIMP tissue inhibitor of metalloproteinase VC vehicle control Submitted: 6 April 2010 Accepted: 15 September 2010 This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms ). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/ ).

Abboud, Nareen; Chow, Siu-Kei; Saylor, Carolyn; Janda, Alena; Ravetch, Jeffery V.; Scharff, Matthew D.; Casadevall, Arturo

doi: 10.1084/jem.20100995pmid: 20921285

One important function of humoral immunity is toxin neutralization. The current view posits that neutralization results from antibody-mediated interference with the binding of toxins to their targets, a phenomenon viewed as dependent only on antibody specificity. To investigate the role of antibody constant region function in toxin neutralization, we generated IgG2a and IgG2b variants of the Bacillus anthracis protective antigen–binding IgG1 monoclonal antibody (mAb) 19D9. These antibodies express identical variable regions and display the same specificity. The efficacy of antibody-mediated neutralization was IgG2a > IgG2b > IgG1, and neutralization activity required competent Fcγ receptor (FcγR). The IgG2a mAb prevented lethal toxin cell killing and mitogen-activated protein kinase/extracellular signal-regulated kinase kinase cleavage more efficiently than the IgG1 mAb. Passive immunization with IgG1 and IgG2a mAb protected wild-type mice, but not FcγR-deficient mice, against B. anthracis infection. These results establish that constant region isotype influences toxin neutralization efficacy of certain antibodies through a mechanism that requires engagement of FcγR. These findings highlight a new parameter for evaluating vaccine responses and the possibility of harnessing optimal FcγR interactions in the design of passive immunization strategies. Footnotes Abbreviations used: BMM BM-derived macrophage FcR Fc receptor LeTx lethal toxin MAPKK mitogen-activated protein kinase kinase MEK MAPK/ERK kinase PA protective antigen Submitted: 19 May 2010 Accepted: 9 September 2010 This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms ). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/ ).

Miletic, Ana V.; Anzelon-Mills, Amy N.; Mills, David M.; Omori, Sidne A.; Pedersen, Irene M.; Shin, Dong-Mi; Ravetch, Jeffrey V.; Bolland, Silvia; Morse, Herbert C.; Rickert, Robert C.

doi: 10.1084/jem.20091962pmid: 20956547

The inositol phosphatases phosphatase and tensin homologue (PTEN) and Src homology 2 domain–containing inositol phosphatase (SHIP) negatively regulate phosphatidylinositol-3-kinase (PI3K)–mediated growth, survival, and proliferation of hematopoietic cells. Although deletion of PTEN in mouse T cells results in lethal T cell lymphomas, we find that animals lacking PTEN or SHIP in B cells show no evidence of malignancy. However, concomitant deletion of PTEN and SHIP (bPTEN/SHIP −/− ) results in spontaneous and lethal mature B cell neoplasms consistent with marginal zone lymphoma or, less frequently, follicular or centroblastic lymphoma. bPTEN/SHIP −/− B cells exhibit enhanced survival and express more MCL1 and less Bim. These cells also express low amounts of p27 kip1 and high amounts of cyclin D3 and thus appear poised to undergo proliferative expansion. Unlike normal B cells, bPTEN/SHIP −/− B cells proliferate to the prosurvival factor B cell activating factor (BAFF). Interestingly, although BAFF availability may promote lymphoma progression, we demonstrate that BAFF is not required for the expansion of transferred bPTEN/SHIP −/− B cells. This study reveals that PTEN and SHIP act cooperatively to suppress B cell lymphoma and provides the first direct evidence that SHIP is a tumor suppressor. As such, assessment of both PTEN and SHIP function are relevant to understanding the etiology of human B cell malignancies that exhibit augmented activation of the PI3K pathway. Footnotes Abbreviations used: APRIL a proliferation-inducing ligand BAFF B cell activating factor BCR B cell receptor DLBCL diffuse large B cell lymphoma FL follicular B cell lymphoma GC germinal center MCL mantle cell lymphoma MZ marginal zone MZL MZ lymphoma PBL peripheral blood lymphocyte PI3K phosphatidylinositol-3-kinase PIP 3 phosphatidylinositol-(3,4,5)-trisphosphate PTEN phosphatase and tensin homologue SHIP Src homology 2 domain–containing inositol phosphatase Submitted: 9 September 2009 Accepted: 16 September 2010 This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms ). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/ ).

Jayaraman, Pushpa; Sada-Ovalle, Isabel; Beladi, Sarah; Anderson, Ana C.; Dardalhon, Valerie; Hotta, Chie; Kuchroo, Vijay K.; Behar, Samuel M.

doi: 10.1084/jem.20100687pmid: 20937702

T cell immunoglobulin and mucin domain 3 (Tim3) is a negative regulatory molecule that inhibits effector T H 1-type responses. Such inhibitory signals prevent unintended tissue inflammation, but can be detrimental if they lead to premature T cell exhaustion. Although the role of Tim3 in autoimmunity has been extensively studied, whether Tim3 regulates antimicrobial immunity has not been explored. Here, we show that Tim3 expressed on T H 1 cells interacts with its ligand, galectin-9 (Gal9), which is expressed by Mycobacterium tuberculosis –infected macrophages to restrict intracellular bacterial growth. Tim3–Gal9 interaction leads to macrophage activation and stimulates bactericidal activity by inducing caspase-1–dependent IL-1β secretion. We propose that the T H 1 cell surface molecule Tim3 has evolved to inhibit growth of intracellular pathogens via its ligand Gal9, which in turn inhibits expansion of effector T H 1 cells to prevent further tissue inflammation. Footnotes Abbreviations used: ANOVA analysis of variance Gal9 galectin-9 HCV hepatitis C virus HuIgG human Ig-γ IL-1RA IL-1R antagonist iNOS inducible NO synthase LDH lactate dehydrogenase L-NIL L-N6-(1-Iminoethyl)lysine dihydrochloride MOI multiplicity of infection Mtb Mycobacterium tuberculosis NO nitric oxide TIM-3 T cell Ig and mucin domain 3 Tim3-Ig Tim3-Ig fusion protein TLR Toll-like receptor Submitted: 8 April 2010 Accepted: 24 September 2010 This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms ). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/ ).

Wang, Yui-Hsi; Voo, Kui Shin; Liu, Bo; Chen, Chun-Yu; Uygungil, Burcin; Spoede, William; Bernstein, Jonathan A.; Huston, David P.; Liu, Yong-Jun

doi: 10.1084/jem.20101376pmid: 20921287

The inflammatory cytokine interleukin (IL)-17 is involved in the pathogenesis of allergic diseases. However, the identity and functions of IL-17–producing T cells during the pathogenesis of allergic diseases remain unclear. Here, we report a novel subset of T H 2 memory/effector cells that coexpress the transcription factors GATA3 and RORγt and coproduce T H 17 and T H 2 cytokines. Classical T H 2 memory/effector cells had the potential to produce IL-17 after stimulation with proinflammatory cytokines IL-1β, IL-6, and IL-21. The number of IL-17-T H 2 cells was significantly increased in blood of patients with atopic asthma. In a mouse model of allergic lung diseases, IL-17–producing CD4 + T H 2 cells were induced in the inflamed lung and persisted as the dominant IL-17–producing T cell population during the chronic stage of asthma. Treating cultured bronchial epithelial cells with IL-17 plus T H 2 cytokines induced strong up-regulation of chemokine eotaxin-3 , Il8, Mip1b , and Groa gene expression. Compared with classical T H 17 and T H 2 cells, antigen-specific IL-17–producing T H 2 cells induced a profound influx of heterogeneous inflammatory leukocytes and exacerbated asthma. Our findings highlight the plasticity of T H 2 memory cells and suggest that IL-17–producing T H 2 cells may represent the key pathogenic T H 2 cells promoting the exacerbation of allergic asthma. Footnotes Abbreviations used: BAL bronchoalveolar lavage BALF BAL fluid IRF4 IFN regulatory factor 4 RORγt retinoic acid–related orphan receptor Submitted: 7 July 2010 Accepted: 1 September 2010 This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms ). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/ ).

Xu, Yingjie; Bismar, Tarek A.; Su, Jie; Xu, Bin; Kristiansen, Glen; Varga, Zsuzsanna; Teng, Lianghong; Ingber, Donald E.; Mammoto, Akiko; Kumar, Rakesh; Alaoui-Jamali, Moulay A.

doi: 10.1084/jem.20100433pmid: 20937704

The actin cross-linking protein filamin A (FLNa) functions as a scaffolding protein and couples cell cytoskeleton to extracellular matrix and integrin receptor signaling. In this study, we report that FLNa suppresses invasion of breast cancer cells and regulates focal adhesion (FA) turnover. Two large progression tissue microarrays from breast cancer patients revealed a significant decrease of FLNa levels in tissues from invasive breast cancer compared with benign disease and in lymph node–positive compared with lymph node–negative breast cancer. In breast cancer cells and orthotopic mouse breast cancer models, down-regulation of FLNa stimulated cancer cell migration, invasion, and metastasis formation. Time-lapse microscopy and biochemical assays after FLNa silencing and rescue with wild-type or mutant protein resistant to calpain cleavage revealed that FLNa regulates FA disassembly at the leading edge of motile cells. Moreover, FLNa down-regulation enhanced calpain activity through the mitogen-activated protein kinase–extracellular signal-regulated kinase cascade and stimulated the cleavage of FA proteins. These results document a regulation of FA dynamics by FLNa in breast cancer cells. Footnotes Abbreviations used: EGF epidermal growth factor EGFP enhanced GFP ERK extracellular signal-regulated kinase EYFP enhanced YFP FA focal adhesion FAK FA kinase MAPK mitogen-activated protein kinase MBP myelin basic protein PSR pSuper retro shRNA short hairpin RNA siRNA small interfering RNA TMA tissue microarray Submitted: 3 March 2010 Accepted: 15 September 2010 This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms ). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/ ).

Byun, Minji; Abhyankar, Avinash; Lelarge, Virginie; Plancoulaine, Sabine; Palanduz, Ayse; Telhan, Leyla; Boisson, Bertrand; Picard, Capucine; Dewell, Scott; Zhao, Connie; Jouanguy, Emmanuelle; Feske, Stefan; Abel, Laurent; Casanova, Jean-Laurent

doi: 10.1084/jem.20101597pmid: 20876309