Publications by authors named "Lisa A Miosge"

20 Publications

  • Page 1 of 1

A Point Mutation in IKAROS ZF1 Causes a B Cell Deficiency in Mice.

J Immunol 2021 Apr 3;206(7):1505-1514. Epub 2021 Mar 3.

Department of Immunology and Infectious Disease, John Curtin School of Medical Research, The Australian National University, Canberra, Australian Capital Territory 2601, Australia;

IKZF1 (IKAROS) is essential for normal lymphopoiesis in both humans and mice. Previous mouse models have demonstrated the dual role for IKZF1 in both B and T cell development and have indicated differential requirements of each zinc finger. Furthermore, mutations in are known to cause common variable immunodeficiency in patients characterized by a loss of B cells and reduced Ab production. Through -ethyl--nitrosourea mutagenesis, we have discovered a novel mutant mouse with a missense mutation (L132P) in zinc finger 1 (ZF1) located in the DNA binding domain. Unlike other previously reported murine mutations, this L132P point mutation ( ) conserves overall protein expression and has a B cell-specific phenotype with no effect on T cell development, indicating that ZF1 is not required for T cells. Mice have reduced Ab responses to immunization and show a progressive loss of serum Igs compared with wild-type littermates. IKZF1 overexpressed in NIH3T3 or HEK293T cells failed to localize to pericentromeric heterochromatin and bind target DNA sequences. Coexpression of wild-type and mutant IKZF1, however, allows for localization to pericentromeric heterochromatin and binding to DNA indicating a haploinsufficient mechanism of action for IKZF1 Furthermore, mice have late onset defective Ig production, similar to what is observed in common variable immunodeficiency patients. RNA sequencing revealed a total loss of expression in follicular B cells, suggesting a possible functional link for the humoral immune response defects observed in mice.
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http://dx.doi.org/10.4049/jimmunol.1901464DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7987828PMC
April 2021

NINJ1 mediates plasma membrane rupture during lytic cell death.

Nature 2021 Mar 20;591(7848):131-136. Epub 2021 Jan 20.

Department of Physiological Chemistry, Genentech Inc., South San Francisco, CA, USA.

Plasma membrane rupture (PMR) is the final cataclysmic event in lytic cell death. PMR releases intracellular molecules known as damage-associated molecular patterns (DAMPs) that propagate the inflammatory response. The underlying mechanism of PMR, however, is unknown. Here we show that the cell-surface NINJ1 protein, which contains two transmembrane regions, has an essential role in the induction of PMR. A forward-genetic screen of randomly mutagenized mice linked NINJ1 to PMR. Ninj1 macrophages exhibited impaired PMR in response to diverse inducers of pyroptotic, necrotic and apoptotic cell death, and were unable to release numerous intracellular proteins including HMGB1 (a known DAMP) and LDH (a standard measure of PMR). Ninj1 macrophages died, but with a distinctive and persistent ballooned morphology, attributable to defective disintegration of bubble-like herniations. Ninj1 mice were more susceptible than wild-type mice to infection with Citrobacter rodentium, which suggests a role for PMR in anti-bacterial host defence. Mechanistically, NINJ1 used an evolutionarily conserved extracellular domain for oligomerization and subsequent PMR. The discovery of NINJ1 as a mediator of PMR overturns the long-held idea that cell death-related PMR is a passive event.
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http://dx.doi.org/10.1038/s41586-021-03218-7DOI Listing
March 2021

Nfkb2 variants reveal a p100-degradation threshold that defines autoimmune susceptibility.

J Exp Med 2021 Feb;218(2)

Infection and Immunity Program, Monash Biomedicine Discovery Institute and Department of Biochemistry and Molecular Biology, Monash University, Clayton, Australia.

NF-κB2/p100 (p100) is an inhibitor of κB (IκB) protein that is partially degraded to produce the NF-κB2/p52 (p52) transcription factor. Heterozygous NFKB2 mutations cause a human syndrome of immunodeficiency and autoimmunity, but whether autoimmunity arises from insufficiency of p52 or IκB function of mutated p100 is unclear. Here, we studied mice bearing mutations in the p100 degron, a domain that harbors most of the clinically recognized mutations and is required for signal-dependent p100 degradation. Distinct mutations caused graded increases in p100-degradation resistance. Severe p100-degradation resistance, due to inheritance of one highly degradation-resistant allele or two subclinical alleles, caused thymic medullary hypoplasia and autoimmune disease, whereas the absence of p100 and p52 did not. We inferred a similar mechanism occurs in humans, as the T cell receptor repertoires of affected humans and mice contained a hydrophobic signature of increased self-reactivity. Autoimmunity in autosomal dominant NFKB2 syndrome arises largely from defects in nonhematopoietic cells caused by the IκB function of degradation-resistant p100.
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http://dx.doi.org/10.1084/jem.20200476DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7595743PMC
February 2021

Functional rare and low frequency variants in BLK and BANK1 contribute to human lupus.

Nat Commun 2019 05 17;10(1):2201. Epub 2019 May 17.

Department of Immunology and Infectious Disease, John Curtin School of Medical Research, Acton, 2601, ACT, Australia.

Systemic lupus erythematosus (SLE) is the prototypic systemic autoimmune disease. It is thought that many common variant gene loci of weak effect act additively to predispose to common autoimmune diseases, while the contribution of rare variants remains unclear. Here we describe that rare coding variants in lupus-risk genes are present in most SLE patients and healthy controls. We demonstrate the functional consequences of rare and low frequency missense variants in the interacting proteins BLK and BANK1, which are present alone, or in combination, in a substantial proportion of lupus patients. The rare variants found in patients, but not those found exclusively in controls, impair suppression of IRF5 and type-I IFN in human B cell lines and increase pathogenic lymphocytes in lupus-prone mice. Thus, rare gene variants are common in SLE and likely contribute to genetic risk.
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http://dx.doi.org/10.1038/s41467-019-10242-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6525203PMC
May 2019

DNA Hypermethylation Encroachment at CpG Island Borders in Cancer Is Predisposed by H3K4 Monomethylation Patterns.

Cancer Cell 2019 02;35(2):297-314.e8

Epigenetics Research Laboratory, Genomics and Epigenetics Division, Garvan Institute of Medical Research, 384 Victoria St, Sydney, NSW 2010, Australia; St Vincent's Clinical School, UNSW, Sydney, NSW 2010, Australia. Electronic address:

Promoter CpG islands are typically unmethylated in normal cells, but in cancer a proportion are subject to hypermethylation. Using methylome sequencing we identified CpG islands that display partial methylation encroachment across the 5' or 3' CpG island borders. CpG island methylation encroachment is widespread in prostate and breast cancer and commonly associates with gene suppression. We show that the pattern of H3K4me1 at CpG island borders in normal cells predicts the different modes of cancer CpG island hypermethylation. Notably, genetic manipulation of Kmt2d results in concordant alterations in H3K4me1 levels and CpG island border DNA methylation encroachment. Our findings suggest a role for H3K4me1 in the demarcation of CpG island methylation borders in normal cells, which become eroded in cancer.
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http://dx.doi.org/10.1016/j.ccell.2019.01.004DOI Listing
February 2019

Systems-guided forward genetic screen reveals a critical role of the replication stress response protein ETAA1 in T cell clonal expansion.

Proc Natl Acad Sci U S A 2017 06 12;114(26):E5216-E5225. Epub 2017 Jun 12.

John Curtin School of Medical Research, Australian National University, Canberra, ACT 2601, Australia;

T-cell immunity requires extremely rapid clonal proliferation of rare, antigen-specific T lymphocytes to form effector cells. Here we identify a critical role for ETAA1 in this process by surveying random germ line mutations in mice using exome sequencing and bioinformatic annotation to prioritize mutations in genes of unknown function with potential effects on the immune system, followed by breeding to homozygosity and testing for immune system phenotypes. Effector CD8 and CD4 T-cell formation following immunization, lymphocytic choriomeningitis virus (LCMV) infection, or herpes simplex virus 1 (HSV1) infection was profoundly decreased despite normal immune cell development in adult mice homozygous for two different mutations: an exon 2 skipping allele that deletes Gly78-Leu119, and a Cys166Stop truncating allele that eliminates most of the 877-aa protein. ETAA1 deficiency decreased clonal expansion cell autonomously within the responding T cells, causing no decrease in their division rate but increasing TP53-induced mRNAs and phosphorylation of H2AX, a marker of DNA replication stress induced by the ATM and ATR kinases. Homozygous ETAA1-deficient adult mice were otherwise normal, healthy, and fertile, although slightly smaller, and homozygotes were born at lower frequency than expected, consistent with partial lethality after embryonic day 12. Taken together with recently reported evidence in human cancer cell lines that ETAA1 activates ATR kinase through an exon 2-encoded domain, these findings reveal a surprisingly specific requirement for this ATR activator in adult mice restricted to rapidly dividing effector T cells. This specific requirement may provide new ways to suppress pathological T-cell responses in transplantation or autoimmunity.
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http://dx.doi.org/10.1073/pnas.1705795114DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5495275PMC
June 2017

A Novel Mutation in Nucleoporin 35 Causes Murine Degenerative Colonic Smooth Muscle Myopathy.

Am J Pathol 2016 09 16;186(9):2254-61. Epub 2016 Jul 16.

Department of Anatomy and Neuroscience, University of Melbourne, Parkville, Victoria, Australia.

Chronic intestinal pseudo-obstruction (CIPO) is a rare but life-threatening disease characterized by severe intestinal dysmotility. Histopathologic studies in CIPO patients have identified several different mechanisms that appear to be involved in the dysmotility, including defects in neurons, smooth muscle, or interstitial cells of Cajal. Currently there are few mouse models of the various forms of CIPO. We generated a mouse with a point mutation in the RNA recognition motif of the Nup35 gene, which encodes a component of the nuclear pore complex. Nup35 mutants developed a severe megacolon and exhibited a reduced lifespan. Histopathologic examination revealed a degenerative myopathy that developed after birth and specifically affected smooth muscle in the colon; smooth muscle in the small bowel and the bladder were not affected. Furthermore, no defects were found in enteric neurons or interstitial cells of Cajal. Nup35 mice are likely to be a valuable model for the subtype of CIPO characterized by degenerative myopathy. Our study also raises the possibility that Nup35 polymorphisms could contribute to some cases of CIPO.
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http://dx.doi.org/10.1016/j.ajpath.2016.04.016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5012464PMC
September 2016

Comparison of predicted and actual consequences of missense mutations.

Proc Natl Acad Sci U S A 2015 Sep 12;112(37):E5189-98. Epub 2015 Aug 12.

Immunogenomics Laboratory, John Curtin School of Medical Research, Australian National University, Canberra City, ACT 2601, Australia;

Each person's genome sequence has thousands of missense variants. Practical interpretation of their functional significance must rely on computational inferences in the absence of exhaustive experimental measurements. Here we analyzed the efficacy of these inferences in 33 de novo missense mutations revealed by sequencing in first-generation progeny of N-ethyl-N-nitrosourea-treated mice, involving 23 essential immune system genes. PolyPhen2, SIFT, MutationAssessor, Panther, CADD, and Condel were used to predict each mutation's functional importance, whereas the actual effect was measured by breeding and testing homozygotes for the expected in vivo loss-of-function phenotype. Only 20% of mutations predicted to be deleterious by PolyPhen2 (and 15% by CADD) showed a discernible phenotype in individual homozygotes. Half of all possible missense mutations in the same 23 immune genes were predicted to be deleterious, and most of these appear to become subject to purifying selection because few persist between separate mouse substrains, rodents, or primates. Because defects in immune genes could be phenotypically masked in vivo by compensation and environment, we compared inferences by the same tools with the in vitro phenotype of all 2,314 possible missense variants in TP53; 42% of mutations predicted by PolyPhen2 to be deleterious (and 45% by CADD) had little measurable consequence for TP53-promoted transcription. We conclude that for de novo or low-frequency missense mutations found by genome sequencing, half those inferred as deleterious correspond to nearly neutral mutations that have little impact on the clinical phenotype of individual cases but will nevertheless become subject to purifying selection.
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http://dx.doi.org/10.1073/pnas.1511585112DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4577149PMC
September 2015

Delayed control of herpes simplex virus infection and impaired CD4(+) T-cell migration to the skin in mouse models of DOCK8 deficiency.

Immunol Cell Biol 2015 Jul 17;93(6):517-21. Epub 2015 Mar 17.

Research School of Biology, Australian National University, Canberra, Australian Capital Territory, Australia.

DOCK8 deficiency in humans and mice leads to multiple defects in immune cell numbers and function. Patients with this immunodeficiency have a high morbidity and mortality, and are distinguished by chronic cutaneous viral infections, including those caused by herpes simplex virus (HSV). The underlying mechanism of the specific susceptibility to these chronic cutaneous viral infections is currently unknown, largely because the effect of DOCK8 deficiency has not been studied in suitable models. A better understanding of these mechanisms is required to underpin the development of more specific therapies. Here we show that DOCK8-deficient mice have poor control of primary cutaneous herpes simplex lesions and this is associated with increased virus loads. Furthermore, DOCK8-deficient mice showed a lack of CD4(+) T-cell infiltration into HSV-infected skin.
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http://dx.doi.org/10.1038/icb.2015.32DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4496291PMC
July 2015

Quantitative reduction of the TCR adapter protein SLP-76 unbalances immunity and immune regulation.

J Immunol 2015 Mar 6;194(6):2587-95. Epub 2015 Feb 6.

Department of Immunology, John Curtin School of Medical Research, Australian National University, Canberra, Australian Capital Territory 2601, Australia;

Gene variants that disrupt TCR signaling can cause severe immune deficiency, yet less disruptive variants are sometimes associated with immune pathology. Null mutations of the gene encoding the scaffold protein Src homology 2 domain-containing leukocyte protein of 76 kDa (SLP-76), for example, cause an arrest of T cell positive selection, whereas a synthetic membrane-targeted allele allows limited positive selection but is associated with proinflammatory cytokine production and autoantibodies. Whether these and other enigmatic outcomes are due to a biochemical uncoupling of tolerogenic signaling, or simply a quantitative reduction of protein activity, remains to be determined. In this study we describe a splice variant of Lcp2 that reduced the amount of wild-type SLP-76 protein by ~90%, disrupting immunogenic and tolerogenic pathways to different degrees. Mutant mice produced excessive amounts of proinflammatory cytokines, autoantibodies, and IgE, revealing that simple quantitative reductions of SLP-76 were sufficient to trigger immune dysregulation. This allele reveals a dose-sensitive threshold for SLP-76 in the balance of immunity and immune dysregulation, a common disturbance of atypical clinical immune deficiencies.
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http://dx.doi.org/10.4049/jimmunol.1400326DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4355390PMC
March 2015

Zinc-finger protein ZFP318 is essential for expression of IgD, the alternatively spliced Igh product made by mature B lymphocytes.

Proc Natl Acad Sci U S A 2014 Mar 10;111(12):4513-8. Epub 2014 Mar 10.

Department of Immunology and Australian Phenomics Facility, John Curtin School of Medical Research, Australian National University, Canberra, ACT 2601, Australia.

IgD and IgM are produced by alternative splicing of long primary RNA transcripts from the Ig heavy chain (Igh) locus and serve as the receptors for antigen on naïve mature B lymphocytes. IgM is made selectively in immature B cells, whereas IgD is coexpressed with IgM when the cells mature into follicular or marginal zone B cells, but the transacting factors responsible for this regulated change in splicing have remained elusive. Here, we use a genetic screen in mice to identify ZFP318, a nuclear protein with two U1-type zinc fingers found in RNA-binding proteins and no known role in the immune system, as a critical factor for IgD expression. A point mutation in an evolutionarily conserved lysine-rich domain encoded by the alternatively spliced Zfp318 exon 10 abolished IgD expression on marginal zone B cells, decreased IgD on follicular B cells, and increased IgM, but only slightly decreased the percentage of B cells and did not decrease expression of other maturation markers CD21, CD23, or CD62L. A targeted Zfp318 null allele extinguished IgD expression on mature B cells and increased IgM. Zfp318 mRNA is developmentally regulated in parallel with IgD, with little in pro-B cells, moderate amounts in immature B cells, and high levels selectively in mature follicular B cells. These findings identify ZFP318 as a crucial factor regulating the expression of the two major antibody isotypes on the surface of most mature B cells.
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http://dx.doi.org/10.1073/pnas.1402739111DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3970522PMC
March 2014

Rasgrp1 mutation increases naive T-cell CD44 expression and drives mTOR-dependent accumulation of Helios⁺ T cells and autoantibodies.

Elife 2013 Dec 12;2:e01020. Epub 2013 Dec 12.

Department of Immunology, John Curtin School of Medical Research, The Australian National University, Canberra, Australia.

Missense variants are a major source of human genetic variation. Here we analyze a new mouse missense variant, Rasgrp1(Anaef), with an ENU-mutated EF hand in the Rasgrp1 Ras guanine nucleotide exchange factor. Rasgrp1(Anaef) mice exhibit anti-nuclear autoantibodies and gradually accumulate a CD44(hi) Helios(+) PD-1(+) CD4(+) T cell population that is dependent on B cells. Despite reduced Rasgrp1-Ras-ERK activation in vitro, thymocyte selection in Rasgrp1(Anaef) is mostly normal in vivo, although CD44 is overexpressed on naïve thymocytes and T cells in a T-cell-autonomous manner. We identify CD44 expression as a sensitive reporter of tonic mTOR-S6 kinase signaling through a novel mouse strain, chino, with a reduction-of-function mutation in Mtor. Elevated tonic mTOR-S6 signaling occurs in Rasgrp1(Anaef) naïve CD4(+) T cells. CD44 expression, CD4(+) T cell subset ratios and serum autoantibodies all returned to normal in Rasgrp1(Anaef)Mtor(chino) double-mutant mice, demonstrating that increased mTOR activity is essential for the Rasgrp1(Anaef) T cell dysregulation. DOI: http://dx.doi.org/10.7554/eLife.01020.001.
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http://dx.doi.org/10.7554/eLife.01020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3858598PMC
December 2013

Unlocking the bottleneck in forward genetics using whole-genome sequencing and identity by descent to isolate causative mutations.

PLoS Genet 2013 31;9(1):e1003219. Epub 2013 Jan 31.

Nuffield Department of Medicine and Wellcome Trust Centre for Human Genetics, Oxford University, Oxford, United Kingdom.

Forward genetics screens with N-ethyl-N-nitrosourea (ENU) provide a powerful way to illuminate gene function and generate mouse models of human disease; however, the identification of causative mutations remains a limiting step. Current strategies depend on conventional mapping, so the propagation of affected mice requires non-lethal screens; accurate tracking of phenotypes through pedigrees is complex and uncertain; out-crossing can introduce unexpected modifiers; and Sanger sequencing of candidate genes is inefficient. Here we show how these problems can be efficiently overcome using whole-genome sequencing (WGS) to detect the ENU mutations and then identify regions that are identical by descent (IBD) in multiple affected mice. In this strategy, we use a modification of the Lander-Green algorithm to isolate causative recessive and dominant mutations, even at low coverage, on a pure strain background. Analysis of the IBD regions also allows us to calculate the ENU mutation rate (1.54 mutations per Mb) and to model future strategies for genetic screens in mice. The introduction of this approach will accelerate the discovery of causal variants, permit broader and more informative lethal screens to be used, reduce animal costs, and herald a new era for ENU mutagenesis.
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http://dx.doi.org/10.1371/journal.pgen.1003219DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3561070PMC
May 2013

Deficiency of caspase recruitment domain family, member 11 (CARD11), causes profound combined immunodeficiency in human subjects.

J Allergy Clin Immunol 2013 Feb;131(2):477-85.e1

Pediatric Hematology-Oncology and Bone Marrow Transplantation, Hadassah Hebrew University Medical Center, Jerusalem, Israel.

Background: Profound combined immunodeficiency can present with normal numbers of T and B cells, and therefore the functional defect of the cellular and humoral immune response is often not recognized until the first severe clinical manifestation. Here we report a patient of consanguineous descent presenting at 13 months of age with hypogammaglobulinemia, Pneumocystis jirovecii pneumonia, and a suggestive family history.

Objective: We sought to identify the genetic alteration in a patient with combined immunodeficiency and characterize human caspase recruitment domain family, member 11 (CARD11), deficiency.

Methods: Molecular, immunologic, and functional assays were performed.

Results: The immunologic characterization revealed only subtle changes in the T-cell and natural killer cell compartment, whereas B-cell differentiation, although normal in number, was distinctively blocked at the transitional stage. Genetic evaluation revealed a homozygous deletion of exon 21 in CARD11 as the underlying defect. This deletion abrogated protein expression and activation of the canonical nuclear factor κB (NF-κB) pathway in lymphocytes after antigen receptor or phorbol 12-myristate 13-acetate stimulation, whereas CD40 signaling in B cells was preserved. The abrogated activation of the canonical NF-κB pathway was associated with severely impaired upregulation of inducible T-cell costimulator, OX40, cytokine production, proliferation of T cells, and B cell-activating factor receptor expression on B cells.

Conclusion: Thus in patients with CARD11 deficiency, the combination of impaired activation and especially upregulation of inducible T-cell costimulator on T cells, together with severely disturbed peripheral B-cell differentiation, apparently leads to a defective T-cell/B-cell cooperation and probably germinal center formation and clinically results in severe immunodeficiency. This report discloses the crucial and nonredundant role of canonical NF-κB activation and specifically CARD11 in the antigen-specific immune response in human subjects.
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http://dx.doi.org/10.1016/j.jaci.2012.11.050DOI Listing
February 2013

Lack of the phosphatase PTPN22 increases adhesion of murine regulatory T cells to improve their immunosuppressive function.

Sci Signal 2012 Nov 27;5(252):ra87. Epub 2012 Nov 27.

Institute for Immunology and Infection Research, The University of Edinburgh, West Mains Road, Edinburgh EH9 3JT, UK.

The cytoplasmic phosphatase PTPN22 (protein tyrosine phosphatase nonreceptor type 22) plays a key role in regulating lymphocyte homeostasis, which ensures that the total number of lymphocytes in the periphery remains relatively constant. Mutations in PTPN22 confer an increased risk of developing autoimmune diseases; however, the precise function of PTPN22 and how mutations contribute to autoimmunity remain controversial. Loss-of-function mutations in PTPN22 are associated with increased numbers of effector T cells and autoreactive B cells in humans and mice; however, the complete absence of PTPN22 in mice does not result in spontaneous autoimmunity. We found that PTPN22 was a key regulator of regulatory T cell (T(reg)) function that fine-tuned the signaling of the T cell receptor and integrins. PTPN22(-/-) T(regs) were more effective at immunosuppression than were wild-type T(regs), and they suppressed the activity of PTPN22(-/-) effector T cells, preventing autoimmunity. Compared to wild-type T(regs), PTPN22(-/-) T(regs) produced increased amounts of the immunosuppressive cytokine interleukin-10 and had enhanced adhesive properties mediated by the integrin lymphocyte function-associated antigen-1, processes that are critical for T(reg) function. This previously undiscovered role of PTPN22 in regulating integrin signaling and T(reg) function suggests that PTPN22 may be a useful therapeutic target for manipulating T(reg) function in human disease.
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http://dx.doi.org/10.1126/scisignal.2003365DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5836999PMC
November 2012

Opposing functions of the T cell receptor kinase ZAP-70 in immunity and tolerance differentially titrate in response to nucleotide substitutions.

Immunity 2007 Dec;27(6):912-26

John Curtin School of Medical Research and Australian Phenomics Facility, The Australian National University, Canberra 2601, Australia.

Null mutations that cripple T cell receptor (TCR) signaling explain rare primary immunodeficiencies, but it is not understood why more common polymorphisms that lead to subtle TCR signaling defects are paradoxically associated with autoimmunity. Here we analyzed how a series of Zap70 variants with step-wise decreases in TCR signaling impacted upon opposing TCR functions of immunity and tolerance. One Zap70 variant, murdock, moderately decreased TCR signaling and thymic selection without compromising immunological tolerance, whereas a more severe Zap70 defect, mrtless, abolished thymic-positive selection and led to immunodeficiency. Signaling capacities between these two thresholds disproportionately compromised negative selection and Foxp3(+) regulatory T cell formation, creating a cellular imbalance between immunogenic and tolerogenic functions that resulted in the excessive production of autoantibodies and immunoglobulin E (IgE). The pleiotropic functions of ZAP-70 and their differential response to graded variation provide a paradigm for understanding the complex outcomes of human genetic variation.
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http://dx.doi.org/10.1016/j.immuni.2007.11.013DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3163119PMC
December 2007

Essential role of membrane cholesterol in accelerated BCR internalization and uncoupling from NF-kappa B in B cell clonal anergy.

J Exp Med 2006 Jul 26;203(7):1773-83. Epub 2006 Jun 26.

John Curtin School of Medical Research, The Australian National University, Canberra ACT 2601, Australia.

Divergent hypotheses exist to explain how signaling by the B cell receptor (BCR) is initiated after antigen binding and how it is qualitatively altered in anergic B cells to selectively uncouple from nuclear factor kappaB and c-Jun N-terminal kinase pathways while continuing to activate extracellular signal-regulated kinase and calcium-nuclear factor of activated T cell pathways. Here we find that BCRs on anergic cells are endocytosed at a very enhanced rate upon binding antigen, resulting in a large steady-state pool of intracellularly sequestered receptors that appear to be continuously cycling between surface and intracellular compartments. This endocytic mechanism is exquisitely sensitive to the lowering of plasma membrane cholesterol by methyl-beta-cyclodextrin, and, when blocked in this way, the sequestered BCRs return to the cell surface and RelA nuclear accumulation is stimulated. In contrast, when plasma membrane cholesterol is lowered and GM1 sphingolipid markers of membrane rafts are depleted in naive B cells, this does not diminish BCR signaling to calcium or RelA. These results provide a possible explanation for the signaling changes in clonal anergy and indicate that a chief function of membrane cholesterol in B cells is not to initiate BCR signaling, but instead to terminate a subset of signals by rapid receptor internalization.
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http://dx.doi.org/10.1084/jem.20060552DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2118358PMC
July 2006

Genes, pathways and checkpoints in lymphocyte development and homeostasis.

Immunol Cell Biol 2005 Aug;83(4):318-35

Immunogenomics Laboratory, Division of Immunology and Genetics, John Curtin School of Medical Research, Australian National University, Canberra, Australia.

A plethora of genes involved in murine B and T cell development have been identified, and developmental pathways within the primary lymphoid tissues have been well delineated. The generation of a functional, but non-self reacting lymphocyte repertoire results from the completion of several checkpoints during lymphocyte development and competition for survival factors in the periphery. Improved knowledge of these developmental checkpoints and homeostatic mechanisms is critical for understanding human immunodeficiency, leukaemia/lymphoma and autoimmunity, which are conditions where checkpoints and homeostasis are likely to be deregulated.
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http://dx.doi.org/10.1111/j.1440-1711.2005.01353.xDOI Listing
August 2005

Identifying the MAGUK protein Carma-1 as a central regulator of humoral immune responses and atopy by genome-wide mouse mutagenesis.

Immunity 2003 Jun;18(6):751-62

Australian Cancer Research Foundation Genetics Laboratory and Medical Genome Centre, John Curtin School of Medical Research, Australian National University, ACT 2601, Canberra, Australia.

In a genome-wide ENU mouse mutagenesis screen a recessive mouse mutation, unmodulated, was isolated with profound defects in humoral immune responses, selective deficits in B cell activation by antigen receptors and T cell costimulation by CD28, and gradual development of atopic dermatitis with hyper-IgE. Mutant B cells are specifically defective in forming connections between antigen receptors and two key signaling pathways for immunogenic responses, NF-kappaB and JNK, but signal normally to calcium, NFAT, and ERK. The mutation alters a conserved leucine in the coiled-coil domain of CARMA-1/CARD11, a member of the MAGUK protein family implicated in organizing multimolecular signaling complexes. These results define Carma-1 as a key regulator of the plasticity in antigen receptor signaling that underpins opposing mechanisms of immunity and tolerance.
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http://dx.doi.org/10.1016/s1074-7613(03)00141-9DOI Listing
June 2003

Analysis of an ethylnitrosourea-generated mouse mutation defines a cell intrinsic role of nuclear factor kappaB2 in regulating circulating B cell numbers.

J Exp Med 2002 Oct;196(8):1113-9

Australian Cancer Research Foundation Genetics Laboratory and Medical Genome Centre, John Curtin School of Medical Research, Australian National University, Canberra ACT 2601, Australia.

The number of circulating follicular B lymphocytes is normally kept within a precise range despite their dispersion through the body and daily overproduction of precursors in the bone marrow. By establishing a genome wide recessive mutation screen in C57BL/6 mice to identify critical components of immune system regulation, we identified a mutant strain with selective deficiency in recirculating B cells but not immature or peritoneal B1 cells. Analysis of mixed bone marrow chimeras established that the mutation affects a cell autonomous process within B cells that is required for their accumulation after emigrating to peripheral lymphoid organs. The defect is caused by a point mutation in the gene encoding transcription factor nuclear factor (NF)-kappaB2, terminating the encoded protein within the DNA-binding domain. These findings establish the feasibility of analyzing immune regulation by genome wide mutant screens and demonstrates an intrinsic requirement for NF-kappaB2 in regulating circulating follicular B cell numbers.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2194042PMC
http://dx.doi.org/10.1084/jem.20020959DOI Listing
October 2002