Emeritus Faculty, Acad Council, Biology
Director, PhD Program in Immunology, Stanford University (1988 - 1994)
Chair, Faculty Senate, Stanford University (1993 - 1994)
Chair, Department of Biological Sciences, Stanford University (1994 - 1997)
Associate Dean of Research, Stanford University (1998 - 2000)
Vice Provost for Faculty Development and Diversity, Stanford University (2000 - 2010)
Director, Stanford Immunology (2011 - 2017)
Honors & Awards
Founder's Prize, Texas Instruments Foundation (1984)
Hoagland Prize for Undergraduate Teaching, Stanford University (1987)
Duca Family University Fellow in Undergraduate Education, Stanford University (2002)
Immunology Program Faculty Mentor Award, Stanford University (2003)
Dr. Nancy Chang Professorship in Humanities and Sciences, Stanford University (2004)
PhD, The Johns Hopkins University, Biology/Immunology (1974)
Current Research and Scholarly Interests
Dr. Jones' research has focused on genetic, molecular, and cellular mechanisms that regulate innate and adaptive immune responses. As immune responses can be harmful, they are highly regulated in their occurrence, magnitude, and duration. Her current work focuses on innate immune and inflammatory responses, triggered by conserved microbial components. Her research group discovered and is characterizing a novel mechanism that regulates innate responses of mammalian macrophages, dendritic cells, and other cells to microbial pathogens, resulting in generation of anti-microbial responses and production of cytokines and other proteins that contribute to innate, inflammatory, and adaptive immune responses. Dr. Jones' group discovered that the signaling pathways activated by the binding of microbial components to Toll-like receptors (TLR), leading to activation of the transcription factor NF-kB as well as MAPK pathways, is negatively-regulated by the protein phosphatase calcineurin. This inhibitory role of calcineurin, which helps to keep signaling downstream of TLR off in resting macrophages and other cells, is opposite to calcineurin's activating role in T and B lymphocytes following activation by antigen. Reflecting calcineurin's inhibitory role in macrophages, the signaling pathway downstream of TLR is activated by calcineurin inhibitors, such as cyclosporine A and FK506, that have long been used as immunosuppressants to block undesired T cell immune responses, such as those mediating organ transplant rejection.
To understand the physiological role of calcineurin in regulating innate immune responses in vivo, the Jones lab has explored the effect of inhibiting calcineurin in mice, using two experimental models. In the first model, mice received multiple injections of the calcineurin inhibitor FK506. In the second model, transgenic mice were created using the Cre-lox system that are deficient in the expression of calcineurin in myeloid cells. In both models mice were tested to see whether the resulting reduction in calcineurin activity led the activation of innate responses or alternatively, as has been observed with multiple exposures to microbial components such as bacterial lipopolysaccharide (LPS), to de-sensitization (a reduction in responsiveness referred to as LPS tolerance). In both models of reduced calcineurin activity, mice did not show upregulation of innate immune responses. In contrast, macrophages from these mice showed reduced responses to innate stimuli in vitro, and when challenged with a lethal dose of LPS (a model for septic shock) these mice showed partial protection. Thus, chronic inhibition of calcineurin activity leads to the induction of negative feedback pathways that limit the potential harmful effects of innate immune and inflammatory responses. These findings suggest that people chronically-treated with calcineurin inhibitor immunosuppressants, such as organ transplant recipients, may be suppressed in their innate as well as adaptive immune responses, perhaps contributing to the well-known increase in their susceptibility to infection.
Independent Studies (11)
- Directed Reading in Biology
BIO 198 (Aut, Win, Spr, Sum)
- Directed Reading in Immunology
IMMUNOL 299 (Aut, Win, Spr, Sum)
- Early Clinical Experience in Immunology
IMMUNOL 280 (Aut, Win, Spr, Sum)
- Graduate Research
BIO 300 (Aut)
- Graduate Research
IMMUNOL 399 (Aut, Win, Spr, Sum)
- Out-of-Department Directed Reading
BIO 198X (Sum)
- Out-of-Department Undergraduate Research
BIO 199X (Win, Sum)
- Teaching Practicum in Biology
BIO 290 (Aut, Spr)
- Teaching in Immunology
IMMUNOL 290 (Aut, Win, Spr, Sum)
- Undergraduate Research
BIO 199 (Sum)
- Undergraduate Research
IMMUNOL 199 (Aut, Win, Spr, Sum)
- Directed Reading in Biology
Prior Year Courses
- Molecular and Cellular Immunology
BIO 230 (Aut)
- Molecular and Cellular Immunology
BIO 230 (Aut)
- Molecular and Cellular Immunology Literature Review
BIO 230A (Aut)
- Molecular and Cellular Immunology
Graduate and Fellowship Programs
Biology (School of Humanities and Sciences) (Phd Program)
The early history of Stanford Immunology.
2014; 58 (2-3): 164-178
From its 1960 beginnings in a pair of windowless Genetics Department laboratories under the Stanford Medical School Dean's Office to its current broad-based program, which joins faculty members from departments across the Medical School, the Stanford Immunology Program has played a central role in shaping both basic and clinical immunology thinking. In this article, we tell the story of the beginnings of this odyssey in a reminiscence-based format that brings the flavor of the time in the words of people who lived and built the history.
View details for DOI 10.1007/s12026-014-8518-z
View details for PubMedID 24804901
Introduction to the special issue on Stanford Immunology.
2014; 58 (2-3): 161-163
View details for DOI 10.1007/s12026-014-8520-5
View details for PubMedID 24798554
Calcineurin inactivation leads to decreased responsiveness to LPS in macrophages and dendritic cells and protects against LPS-induced toxicity in vivo
2009; 15 (2): 109-120
Microbial components such as lipopolysaccharide (LPS) bind to Toll-like receptors (TLRs) and activate innate and inflammatory responses. Responses to LPS and other microbial components are limited by the activation of negative feedback mechanisms that reduce responsiveness to subsequent LPS exposure, often termed LPS tolerance. Our laboratory has previously shown that calcineurin, a phosphatase known for its activation of T cells via NFAT, negatively regulates the TLR pathway in macrophages; consequently, calcineurin inhibitors (FK506 and cyclosporin A) mimic TLR ligands in activating the TLR pathway, NF-KB, and associated innate and inflammatory responses. This study investigated the physiological consequences of calcineurin inactivation for LPS-induced inflammatory responses in vitro and in vivo using two models: calcineurin inhibition by FK506 (tacrolimus) and myeloid cell-specific calcineurin deletion. Activation of dendritic cells and macrophages with FK506 in vitro was shown to induce a state of reduced responsiveness to LPS (i.e. a form of LPS tolerance). Similarly, macrophages from FK506-treated mice or from mice in which the calcineurin B1 (CnB1) subunit was conditionally knocked out in myeloid cells were found to have diminished LPS-induced inflammatory responses. In addition, mice with CnB1-deficient myeloid cells and mice undergoing FK506 treatment showed improved survival and recovery when challenged with high doses of systemic LPS compared to controls. These results demonstrate that inactivation of calcineurin in macrophages and other myeloid cells by inhibition or deletion can induce a form of LPS tolerance and protect the host from LPS toxicity in vivo.
View details for DOI 10.1177/1753425908100928
View details for Web of Science ID 000265785600005
View details for PubMedID 19318421
Calcineurin negatively regulates TLR-Mediated activation pathways
JOURNAL OF IMMUNOLOGY
2007; 179 (7): 4598-4607
In innate immunity, microbial components stimulate macrophages to produce antimicrobial substances, cytokines, other proinflammatory mediators, and IFNs via TLRs, which trigger signaling pathways activating NF-kappaB, MAPKs, and IFN response factors. We show in this study that, in contrast to its activating role in T cells, in macrophages the protein phosphatase calcineurin negatively regulates NF-kappaB, MAPKs, and IFN response factor activation by inhibiting the TLR-mediated signaling pathways. Evidence for this novel role for calcineurin was provided by the findings that these signaling pathways are activated when calcineurin is inhibited either by the inhibitors cyclosporin A or FK506 or by small interfering RNA-targeting calcineurin, and that activation of these pathways by TLR ligands is inhibited by the overexpression of a constitutively active form of calcineurin. We further found that IkappaB-alpha degradation, MAPK activation, and TNF-alpha production by FK506 were reduced in macrophages from mice deficient in MyD88, Toll/IL-1R domain-containing adaptor-inducing IFN-beta (TRIF), TLR2, or TLR4, whereas macrophages from TLR3-deficient or TLR9 mutant mice showed the same responses to FK506 as those of wild-type cells. Biochemical studies indicate that calcineurin interacts with MyD88, TRIF, TLR2, and TLR4, but not with TLR3 or TLR9. Collectively, these results suggest that calcineurin negatively regulates TLR-mediated activation pathways in macrophages by inhibiting the adaptor proteins MyD88 and TRIF, and a subset of TLRs.
View details for Web of Science ID 000249752100036
View details for PubMedID 17878357
Inhibitor kappa B and nuclear factor kappa B in granulocyte-macrophage colony-stimulating factor antagonism of dexamethasone suppression of the macrophage response to Aspergillus fumigatus conidia
42nd Annual Meeting of the Infectious-Diseases-Society-of-America
UNIV CHICAGO PRESS. 2006: 1023–28
The dexamethasone (DEX) immunosuppressive effect on macrophage killing activity and cytokine production in response to Aspergillus fumigatus conidia is antagonized by granulocyte-macrophage colony-stimulating factor (GM-CSF). The molecular mechanism is unknown. We postulated that this antagonism is mediated by inhibitor kappaB (I kappaB) induction by DEX and is opposed by acceleration of I kappaB degradation by GM-CSF with or without conidia stimulation, with corresponding effects on translocation and activation of nuclear factor kappa B (NF-kappaB).We studied 2 types of cells, resident peritoneal macrophages from CD-1 mice and the murine macrophage RAW264.7 cell line. Cells were unstimulated or stimulated with conidia and simultaneously treated with DEX, GM-CSF, or DEX plus GM-CSF, for 2-4 hours. I kappaB degradation and NF-kappaB activation were assessed by Western blot.Macrophages stimulated with conidia alone increased NF-kappaB translocation. DEX increased I kappaB levels in cytoplasm and blocked translocation of NF-kappaB to the nucleus in unstimulated and conidia-stimulated macrophages. Conversely, GM-CSF decreased I kappaB levels. GM-CSF reversed the effect of DEX on I kappaB levels. NF-kappaB levels were minimal in DEX-treated macrophage nuclear extracts, compared with those from GM-CSF-treated and GM-CSF plus DEX-treated macrophages.GM-CSF can reverse the DEX immunosuppressive effect by enhancing I kappaB degradation and promoting NF-kappaB translocation. This would allow macrophage production of proinflammatory cytokines, facilitating resistance to aspergillosis.
View details for Web of Science ID 000235777000016
View details for PubMedID 16518765
Identification of an IFN-gamma responsive region in an intron of the invariant chain gene
EUROPEAN JOURNAL OF IMMUNOLOGY
2000; 30 (9): 2604-2611
The mechanism by which IFN-gamma up-regulates invariant chain mRNA in antigen-presenting cells has been under intensive investigation. This study shows that in murine monocytic cells the transcriptional up-regulation mediated by the invariant chain (Ii) upstream enhancer only accounts for part of the induction of Ii mRNA by IFN-gamma. We have identified an intronic region in the murine Ii gene that contributes to the transcriptional up-regulation by IFN-gamma. The region has S (H), X/X2 and Y boxes similar to those in MHC class II promoters and the Ii upstream enhancer. Mutations at the putative S, X and Y boxes have abolished the ability of the region to mediate Ii up-regulation by IFN-gamma. Consistent with the functions of these boxes, our findings reveal that the up-regulation of Ii transcription by IFN-gamma mediated by the intronic region is dependent on the induction of class II transactivator by IFN-gamma.
View details for Web of Science ID 000089223800020
View details for PubMedID 11009094
Calcineurin and vacuolar-type H+-ATPase modulate macrophage effector functions
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
1999; 96 (11): 6324-6329
While effector molecules produced by activated macrophages (including nitric oxide, tumor necrosis factor alpha, interleukin 1, etc.) help to eliminate pathogens, high levels of these molecules can be deleterious to the host itself. Despite their importance, the mechanisms modulating macrophage effector functions are poorly understood. This work introduces two key negative regulators that control the levels and duration of macrophage cytokine production. Vacuolar-type H+-ATPase (V-ATPase) and calcineurin (Cn) constitutively act in normal macrophages to suppress expression of inflammatory cytokines in the absence of specific activation and to inhibit macrophage cytokine responses induced by bacterial lipopolysaccharide (V-ATPase), interferon gamma (V-ATPase and Cn), and calcium (Ca2+) flux (Cn). Cn and V-ATPase modulate effector gene expression at the mRNA level by inhibiting transcription factor NF-kappaB. This negative regulation by Cn is opposite to its crucial positive role in T cells, where it activates NFAT transcription factor(s) leading to expression of interleukin 2, tumor necrosis factor alpha, and other cytokine genes. The negative effects of V-ATPase and Cn on NF-kappaB-dependent gene expression are not limited to the macrophage lineage, as similar effects have been seen with a murine fibroblast cell line and with primary astrocytes.
View details for Web of Science ID 000080527100075
View details for PubMedID 10339586
The invariant chain gene intronic enhancer shows homology to class II promoter elements
JOURNAL OF IMMUNOLOGY
1998; 161 (4): 1844-1852
Coordinate expression of MHC class II proteins and the class II-associated invariant chain (Ii) is important for proper MHC class II functioning in Ag processing and presentation. The coordinate regulation of these genes results, in part, from the sharing of transcriptional regulatory regions between MHC class II and Ii genes; the Ii has previously been shown to have an upstream enhancer closely related to the essential class II promoter elements. We report here the characterization of a second enhancer in the Ii gene, located within the first intron. This intronic enhancer is contained within a 155-bp region, enhances transcription from the Ii minimal promoter, and also contains elements that are homologous to class II promoter elements X1, X2, and Y boxes.
View details for Web of Science ID 000075345400035
View details for PubMedID 9712052
Evidence that the autoimmune antigen myelin basic protein (MBP) Ac1-9 binds towards one end of the major histocompatibility complex (MHC) cleft
JOURNAL OF EXPERIMENTAL MEDICINE
1998; 187 (9): 1505-1516
The NH2-terminal peptide of myelin basic protein (MBP) bound to the class II major histocompatibility complex (MHC) protein I-Au is an immunodominant epitope in experimental autoimmune encephalomyelitis, a murine model of multiple sclerosis. However, the MBP-I-Au complex is very unstable. To investigate this, we performed site-directed mutagenesis of the I-Au MHC protein and the MBP peptide. Biochemical, T cell activation, and molecular modeling studies of mutant complexes demonstrate that the MBP peptide's key residue for MHC binding, lysine 4, is buried in the P6 pocket of I-Au, which is predominantly hydrophobic. This implies that the MBP-I-Au complex differs from more stable complexes in two respects: (a) the peptide leaves the NH2-terminal region of the MHC peptide-binding cleft unoccupied; (b) the peptide is not anchored by typical favorable interactions between peptide side chains and MHC pockets. To test these hypotheses, a modified MBP peptide was designed based on molecular modeling, with the aim of producing strong I-Au binding. Extension of the NH2 terminus of MBP with six amino acids from the ova peptide, and replacement of the lysine side chain in the P6 pocket with an aromatic anchor, results in >1,000-fold increased binding stability. These results provide an explanation for the unusual peptide-MHC-binding kinetics of MBP, and should facilitate an understanding of why mice are not tolerant to this self-peptide- MHC complex.
View details for Web of Science ID 000073505900015
View details for PubMedID 9565642
Novel genetic regulation of T helper 1 (Th1)/Th2 cytokine production and encephalitogenicity in inbred mouse strains
JOURNAL OF EXPERIMENTAL MEDICINE
1997; 185 (3): 439-451
Development of T helper cell (Th)1 or Th2 cytokine responses is essential for effector and regulatory functions of T helper cells. We have compared cytokine profiles of myelin basic protein (MBP) Ac1-16 peptide-specific T helper cells from inbred mouse strains expressing identical k haplotype-derived MHC class II molecules B10.A and B10.BR, B10.BR T cell lines (TCL) produced Th1 cytokines (including high levels of TNF-alpha) and induced experimental autoimmune encephalomyelitis after adoptive transfer. In contrast, B10.A TCL produced Th2 cytokines (including low levels of TNF-alpha) and were poorly encephalitogenic. The contributions of the genetic origin of the T cells and the APC were explored. Serial restimulations of the B10.BR TCL with B10.A or (B10.A x B10.BR) F1 splenic antigen presenting cells (APC) during the establishment of TCL markedly reduced both Th1 cytokine production and encephalitogenicity. In addition, a single restimulation with B10. A splenic APC reduced IFN-gamma and TNF-alpha production by established Th1 MBP-specific Ak-restricted B10.BR TCL and by a Th1 KLH-specific, Ek-restricted B10.BR T cell clone. These studies suggest that B10.A and B10.BR APC differ in their ability to stimulate IFN-gamma and TNF-alpha production by mature Th1 cells and also influence their Th1/Th2 commitment in vivo. The nature of the downregulatory activity of B10.A APC on IFN-gamma and TNF-alpha production was explored. 2-hour supernatants from antigen-activated B10.A APC/TCL cultures or from B10.A APC activated by LPS had the same inhibitory effects on IFN-gamma and TNF-alpha production by B10.BR TCL. The downregulatory effects of B10.A APC are independent of TNF-alpha, IL-4, IL-10, IL-12p40, IFN-gamma, IL-13, TGF-beta, and PGE2. Thus, genetic difference(s) between B10.A and B10.BR APC appear(s) to control the production or activity of a novel soluble cytokine regulatory factor that influences Th1/Th2 commitment and controls production of IFN-gamma and TNF-alpha by mature Th1 cells.
View details for Web of Science ID A1997WG59900008
View details for PubMedID 9053444
Early biochemical signals arise from low affinity TCR-ligand reactions at the cell-cell interface
JOURNAL OF EXPERIMENTAL MEDICINE
1996; 184 (2): 777-782
The kinetics of acid release by a mixture of T cells and antigen presenting cells were measured with a microphysiometer during a brief exposure to antigenic peptides. We find that some of the early biochemical events that lead to cellular proliferation cause a specific increase in the rate of acid release. The duration of this increase in acid release reflects the life-time of the peptide-MHC complexes. Peptides that form long-lived complexes produce a response that is stable for more than an hour. Serial TCR engagement is suggested by the observation that the amplitude of this stable response can be rapidly shifted up or down with additional agonist peptide or with antibodies that block T cell receptor binding. Cells briefly exposed to a peptide that forms short-lived peptide-MHC complexes produce a response that decays rapidly as peptide is washed away. A quantitative analysis of the kinetics of this decay in acidification demonstrates that intercellular TCR-ligand reactions are rapid, reversible, and of low apparent affinity with < 20% of peptide-MHC ligand bound to a TCR at any one time. These results demonstrate that the fraction of peptide-MHC ligands bound to TCRs at the cell-cell interface is no higher than anticipated from the affinities observed in solution for isolated TCRs and ligands.
View details for Web of Science ID A1996VC33700052
View details for PubMedID 8760833
INACTIVATION OF E(ALPHA) AND E(BETA) EXPRESSION IN INBRED AND WILD MICE BY MULTIPLE DISTINCT MUTATIONS, SOME OF WHICH PREDATE SPECIATION WITHIN MUS SPECIES
1995; 7 (9): 1459-1471
The H-2 MHC of mice encodes two functional class II heterodimeric proteins: A alpha A beta (A) and E alpha E beta (E). While failure to express the A protein has not been reported, a significant proportion of of H-2 haplotypes in both inbred and wild mice do not express E proteins. We and others have previously characterized the molecular basis for defective E expression in haplotypes from Mus domesticus (b, f, q, s, from inbred strains) and M. castaneus (w17, wild-derived) species, identifying six distinct defects in the genes for E alpha or E beta. In this report we have extended these studies to other E- haplotypes, including several from t-haplotype-bearing M. domesticus mice (w29, w57, w302) and one derived from the Asian species M. bactrianus (w301). Analyses at the protein, RNA and DNA levels were employed to identify the defects in the genes for Ea and Eb. At least one new defect was identified that prevents E beta expression in a t-associated H-2 haplotypes (w57), bringing the number of distinct mutations causing the E- phenotype to seven. Another t-associated haplotype, w302, was found to share the same E beta defect with mice of the inbred q haplotype and of the w17 haplotype from M. castaneus, while its Ea gene contains the deletion carried also by the inbred b and s haplotypes and by a number of wild haplotypes. The mutations in the Ea and Eb genes of the w301 haplotype from M. bactrianus were found to be identical to those of the inbred f haplotype. This indicates that the origin of the mutations in the Eb genes of the q, w17 and w302 haplotypes and in the Ea and Eb genes of the f and w301 haplotypes, predated speciation within Mus, thought to have occurred approximately 0.35-1 million years ago. Their maintenance in mouse populations suggests that in certain conditions the failure to express E alpha E beta proteins may be advantageous and selected for.
View details for Web of Science ID A1995RV02100010
View details for PubMedID 7495754
INTERACTIONS AMONG POLYMORPHIC AND CONSERVED RESIDUES IN MHC CLASS-II PROTEINS AFFECT MHC-PEPTIDE CONFORMATION AND T-CELL RECOGNITION
1995; 7 (5): 747-761
The structural basis for MHC-restricted T cell recognition of the N-terminal peptide of myelin basic protein (MBP Ac1-11) presented by two mouse class II alleles, Ak and Au, was examined, focusing on the roles of A beta chain polymorphic residues 38 beta (in the beta sheet) and 61 beta (in the alpha helix) in controlling the responses of panels of Ak- and Au-restricted T cell hybridomas. Despite the conservative nature of the substitutions at 38 beta (k = Val, u = Leu) and 61 beta (k = Trp, u = Tyr), transfectants expressing Ak or Au proteins carrying allelic substitutions at 38 beta and/or 61 beta gave dramatically reduced T cell responses. The modest reduction in peptide binding detected using a biotinylated MBP peptide analog appear insufficient to explain the reduced responses, suggesting that changes at 38,61 beta create conformational changes in the MHC-peptide complex. The impact of allelic substitutions at 38,61 beta on T cell responses is also modulated by other residues differing between Ak and Au. To explore the structural basis for these phenomena, protein models were developed of the Ak, Au and 38,61 beta mutant proteins using self-consistent ensemble optimization methodologies. Substitutions of the alternative allelic residue at 38 beta and/or 61 beta, which are in van der Waals contact, change the configuration of this region of the peptide-binding groove, and potentially might affect the conformation of the bound peptide and its hydrogen-bonding to residue 61 beta. The models predict that this region of the groove is markedly altered by allelic differences at A beta residue 9 beta (k = His, u = Val) which determine the position of the side-chain of Tyr30 beta, adjacent to residues 38 beta and 61 beta. Thus, interactions among polymorphic and conserved residues control the antigen presentations functions of MHC class II proteins.
View details for Web of Science ID A1995QZ06400005
View details for PubMedID 7547702
THE MINIMAL POLYMORPHISM OF CLASS-II E-ALPHA CHAINS IS NOT DUE TO THE FUNCTIONAL NEUTRALITY OF MUTATIONS
1994; 40 (1): 9-20
Given the extensive allelic amino acid sequence polymorphism present in the first domain of A alpha, A beta, and E beta chains and its profound effects on class II function, the minimal polymorphism in the mouse E alpha chain (and in its human homologue DR alpha) is paradox. Two possible explanations for the lack of polymorphism in E alpha are: (1) the E alpha chain plays such a uniquely critical structural/functional role in antigen presentation, T-cell activation, repertoire selection, and/or pairing with E beta or other proteins for expression that it cannot vary, and mutations are selected against; (2) the E alpha chain plays a less significant role than the outer domains of other major histocompatibility complex (MHC) proteins in determining the interactions with processed peptides or with T-cell receptor (TCR), so there is no selective pressure to maintain new mutations. To explore this question we compared the ability of transfectants expressing wild type (wt) E alpha E beta d and mutant E alpha wt E beta d proteins to present peptides and bacterial superantigens to T-cell hybridomas. Mutations at the E alpha amino acid positions 31, 52, and 65&66, to residues that represent allelic alternatives in A alpha chains, significantly reduced activation of peptide-specific T hybridomas, and mutations at 71 sometimes enhanced T-cell stimulation. None of the E alpha mutations reduced, and some enhanced, superantigen stimulation of T-cell hybridomas. These results argue against the hypothesis that E alpha chains are minimally polymorphic because mutations in E alpha are functionally neutral.
View details for Web of Science ID A1994NN36200002
View details for PubMedID 8206532
EVIDENCE THAT THE MECHANISM OF PRENATAL GERM-CELL DEATH IN THE MOUSE IS APOPTOSIS
EXPERIMENTAL CELL RESEARCH
1993; 209 (2): 238-247
Using fluorescence-activated cell sorting combined with fluorescence microscopy the mechanism of embryonic germ cell death in the mouse has been shown to be apoptosis. Primordial germ cells (PGCs) from embryos at specific developmental stages have been analyzed, and cells with apoptotic morphology have been isolated by cell sorting. In the female, apoptotic oogonia at Day 13 and apoptotic oocytes at Days 15 and 17 were found. In the male, apoptotic cells were seen on Day 13 through Day 17. Apoptotic germ cells were not detected at Day 12 (combined male and female PGCs). Examination of sorted cells by fluorescence microscopy and by light microscopic analysis after alkaline phosphatase staining confirmed that the cells are apoptotic germ cells. Electron microscopy further confirmed that cells showing the morphological characteristics of apoptosis are present.
View details for Web of Science ID A1993MM46500010
View details for PubMedID 8262141
CHANGES IN PROTOONCOGENE EXPRESSION CORRELATED WITH GENERAL AND SEX-SPECIFIC DIFFERENTIATION IN MURINE PRIMORDIAL GERM-CELLS
MECHANISMS OF DEVELOPMENT
1993; 42 (1-2): 49-58
Primordial germ cells (PGCs) of the mouse undergo key developmental transitions during embryonic days 12-15. On day 12 they complete migration into the gonads. They cease mitotic proliferation on day 13 and subsequently enter sex-specific pathways of development. The molecular mechanisms controlling these transitions are poorly understood, yet they are crucial to production of normal gametes later in life. We have used the polymerase chain reaction (PCR) to directly compare levels of expression of several protooncogenes proposed to be involved in control of cell proliferation and differentiation in proliferating and differentiating PCGs of both sexes over a 4 day time course. We report here that mRNA levels for nuclear protooncogenes c-myc, c-fos, and c-jun increase dramatically in both sexes from little or no detectable expression on day 12 to high expression on days 13-15. We observe c-kit message on day 12 in combined PGCs of both sexes, in female but not male PGCs on day 13, and in both sexes on day 14, c-kit mRNA is undetectable on day 15 in either sex, c-mos is not expressed at detectable levels on day 12 in either sex, but increases gradually in female PGCs to very high levels on day 15. In male PGCs, c-mos is expressed at high levels on days 13-15. Our results are consistent with a role for protooncogenes c-myc, c-fos and c-jun in mediating the initial differentiation of PGCs of both sexes that occurs upon colonization of the gonad. Because c-kit and c-mos are expressed differentially in male and female day 13-15 germ cells, they may play roles in initiating or mediating progress along the sex-specific pathways of development that PGCs embark upon at this time.
View details for Web of Science ID A1993LK60600004
View details for PubMedID 8369223
TEMPORAL-ORDER OF DNA-REPLICATION IN THE H-2 MAJOR HISTOCOMPATIBILITY COMPLEX OF THE MOUSE
MOLECULAR AND CELLULAR BIOLOGY
1992; 12 (11): 5174-5188
As an approach to mapping replicons in an extended chromosomal region, the temporal order of DNA replication was analyzed in the murine major histocompatibility gene complex (MHC). Replicating DNA from T-lymphoma and myelomonocyte cell lines was density labeled with bromodeoxyuridine and extracted from cells which had been fractionated into different stages of S phase by centrifugal elutriation. The replicating DNA from each fraction of S phase was separated from nonreplicating DNA on density gradients, blotted, and hybridized with 34 specific MHC probes. The earliest replication occurred in the vicinity of transcribed genes K, HAM1 and HAM2, RD, B144, D, L, T18, and T3. The temporal order of replication of groups of DNA segments suggests the location of five or six replicons within the H-2 complex, some of which appear to be either unidirectional or markedly asymmetric. The rates of replication through each of these apparent replicons appear to be similar. The TL region of the S49.1 T-lymphoma cells, which contains at least three transcribed genes, replicates earlier than the inactive TL region of WEHI-3 myelomonocytic cells. These results provide further evidence of a relationship between transcription and the initiation of DNA replication in mammalian cells. The mouse MHC examined in this study is the largest chromosomal region (> 2,000 kb) measured for timing of replication to date.
View details for Web of Science ID A1992JT84500039
View details for PubMedID 1406689
COMBINED EFFECTS OF TUMOR NECROSIS FACTOR-ALPHA, PROSTAGLANDIN-E2, AND CORTICOSTERONE ON INDUCED IA EXPRESSION ON MURINE MACROPHAGES
JOURNAL OF IMMUNOLOGY
1990; 145 (4): 1167-1175
Ia expression is an important marker of macrophage functional capacity. IFN-gamma induces Ia expression on perhaps all murine macrophages, whereas IL-4, granulocyte-macrophage CSF, and CSF-1 induce Ia on restricted sets of macrophages. Inhibitors of expression include PGE2, glucocorticoids, and IFN-beta. TNF has been found to augment Ia expression on several macrophage lineage cell lines but to inhibit expression on murine peritoneal macrophages. Our study shows that TNF can have opposite effects on Ia expression (induced by IFN-gamma) on thioglycollate-elicited peritoneal macrophages, depending on the length of time cells are treated and on the presence of other modulators. In particular, TNF augmented early expression induced by IFN-gamma but inhibited later expression. And although TNF synergized with PGE2 to markedly inhibit Ia induction on these cells, it partially antagonized the inhibition by corticosterone and IFN-beta. TNF and PGE2 also synergized to inhibit Ia expression induced on bone marrow-derived and splenic macrophages by either IFN-gamma or IL-4. In contrast to their effect on Ia expression, TNF and PGE2 had opposite effects on expression of gamma 2a FcR in macrophages. TNF blocked the increase in FcR expression due to any combination of PGE2, IFN-gamma, and IFN-beta. However, TNF and PGE2 both increased expression of gamma 2a FcR on WEHI-3 cells. If the different effects of TNF reflect the differentiation states of macrophages, its effects on Ia and FcR expression may vary with the progression of an immune response.
View details for Web of Science ID A1990DT22800018
View details for PubMedID 2116476
TRANSCRIPTIONAL CONTROL OF THE INVARIANT CHAIN GENE INVOLVES PROMOTER AND ENHANCER ELEMENTS COMMON TO AND DISTINCT FROM MAJOR HISTOCOMPATIBILITY COMPLEX CLASS-II GENES
MOLECULAR AND CELLULAR BIOLOGY
1990; 10 (8): 3906-3916
The invariant chain (Ii) is a glycoprotein coexpressed with the major histocompatibility complex (MHC) class II antigens. Although Ii is encoded by a single gene unlinked to the MHC gene complex, Ii and MHC class II appear to have similar patterns of tissue specific expression and generally are coordinately regulated by cytokines. Here we present evidence that transcription of the murine Ii gene is controlled by multiple cis-acting elements. The 5' regulatory region of the Ii gene appears to be combined of conserved class II regulatory elements with promoter elements commonly found in other eucaryotic genes. A region containing characteristic class II promoter elements (H box, X box, and a modified Y box) serves as an upstream enhancer in the Ii gene and might contribute to the coexpression of MHC class II and Ii genes. A series of positive control elements, the kappa B element, Sp1-binding site, and CCAAT box, are present in the Ii promoter and apparently serve distinct regulatory functions. The kappa B site in the Ii gene is a cell type-specific element, contributing to expression in a B-cell line but not in a fibroblast cell line, and the Sp1 site is required by the H-X-Y' enhancer element to stimulate promoter activity. In addition, an Ii enhancer in the first intron that specifically stimulates its own promoter has been identified. Our results suggest that a sequence match between enhancers and certain promoter elements is critical.
View details for Web of Science ID A1990DP92000006
View details for PubMedID 2115116
CHARACTERIZATION OF THE MOLECULAR DEFECTS IN THE MOUSE E-BETA-F AND E-BETA-Q GENES - IMPLICATIONS FOR THE ORIGIN OF MHC POLYMORPHISM
JOURNAL OF IMMUNOLOGY
1990; 144 (5): 1957-1964
The E beta f and E beta q genes have been isolated and sequenced to investigate the molecular basis for their defective expression. A previous study from this laboratory, which characterized the expression of these genes at the RNA level, showed both genes to have defects in posttranscriptional RNA processing. In this paper, the defect in the E beta q gene from the inbred mouse strain B10.G (Mus musculus domesticus) is shown to be a single base insertion in the RNA donor splice site of the first intron. This identical mutation was described previously for the E beta gene of the H-2w17 haplotype, which was derived from the Asian house mouse subspecies Mus musculus castaneus. Although it has been estimated that M. m. domesticus and M. m. castaneus separated from each other more than one million years ago, comparisons of genomic sequences reveal that the nonexpressed E beta q and E beta w17 alleles have not diverged significantly from one another; they are identical in their protein coding regions and have only minor differences elsewhere. In contrast, sequence comparisons of A beta q and A beta w17 show that these two expressed alleles differ by multiple amino acids. These findings provide evidence that selection, acting on expressed MHC proteins, plays a role in accumulation and maintenance of MHC polymorphism. The defective E beta f gene from the inbred strain B10.M has also been isolated. Sequence analysis has identified a mutation in the same RNA donor splice site as E beta q and E beta w17; however, in this gene the mutation is a single base substitution at position 5.
View details for Web of Science ID A1990CR42900056
View details for PubMedID 2106558
SEQUENCE ELEMENTS REQUIRED FOR ACTIVITY OF A MURINE MAJOR HISTOCOMPATIBILITY COMPLEX CLASS-II PROMOTER BIND COMMON AND CELL-TYPE-SPECIFIC NUCLEAR FACTORS
MOLECULAR AND CELLULAR BIOLOGY
1990; 10 (2): 593-604
We have examined the sequence elements and corresponding DNA-binding factors required for transient expression of the A alpha d promoter fused to the bacterial chloramphenicol acetyltransferase reporter gene in a variety of cultured cell lines. Deletion analysis demonstrated that only about 110 nucleotides of sequence 5' of the transcription start site are required for constitutive expression in the murine B-lymphoma cell line A20 or for gamma interferon-induced expression in the murine monocytic cell line WEHI-3. Linker-scanner mutation of this region indicated that at least three sequence elements are required for promoter activity. These elements correspond to the conserved sequence elements found in other human and mouse class II genes, the X box, the Y box, and the H box. Analysis of DNA-binding activity showed that the three most predominant factors present in extracts from WEHI-3, A20, or L cells (which do not express the class II genes) are actually a family of factors that bind to a fourth sequence element, overlapping the 3' end of the X-box sequence, that is homologous to the cyclic AMP-responsive enhancer element. A single common factor that binds to the Y box was detected in extracts from all cells tested, as has been seen with the Y-box elements of other class II genes. Another common factor was found that binds to the more conserved 5' region of the X-box element, although A20 extracts contained a second, distinct binding activity for this region. A common binding factor for the H-box element was detected in extracts from WEHI-3 and L cells. However, this activity was absent in A20 cell extracts. Instead, two different H-box-binding activities were detected, suggesting that different components are involved in class II gene expression in B cells and macrophages. Finally, gamma interferon treatment did not significantly alter the DNA-binding activity in WEHI-3 cells for any of the sequence elements shown to be required for induced chloramphenicol acetyltransferase expression.
View details for Web of Science ID A1990CL19700020
View details for PubMedID 2105455
EVOLUTION OF CLASS-II GENES - ROLE OF SELECTION IN BOTH THE MAINTENANCE OF POLYMORPHISM AND THE RETENTION OF NONEXPRESSED ALLELES
1990; 9 (3): 200-211
View details for Web of Science ID A1990DT08900005
View details for PubMedID 2172414
POLYMORPHIC RESIDUES ON THE I-A-BETA CHAIN MODULATE THE STIMULATION OF T-CELL CLONES SPECIFIC FOR THE N-TERMINAL PEPTIDE OF THE AUTO-ANTIGEN MYELIN BASIC-PROTEIN
JOURNAL OF IMMUNOLOGY
1989; 143 (7): 2083-2093
The effect of polymorphic residues on the A alpha A beta molecule on T cell recognition of the N-terminal nonapeptide of myelin basic protein (R1-9) was determined. Ak-restricted T cell clones recognizing R1-9 were isolated. The peptide-Ia specificities of these clones were determined by testing the response to 1) a panel of peptide analogs of R1-11, 2) splenic APC from mice expressing MHC molecules from serologically distinct haplotypes, and 3) L cell transfectants expressing mutant/recombinant A beta cDNA containing combinations of polymorphic nucleotide sequences from the k and u alleles. Comparisons were made between the Ak-restricted clones and a previously characterized panel of Au-restricted clones. Certain Ak-restricted clones were able to recognize MBP peptide analogs that were not recognized by any of the Au-restricted clones. The Au-restricted T cell clones did not cross-react with R1-9 presented in the context of Ak, whereas the majority of the Ak-restricted clones responded to R1-9 presented in the context of Au. This nonreciprocal cross-reactivity was also reflected in the relative responses of the two sets of T cell clones to the interchange of u- and k-derived residues in the A beta chain. Residues in regions corresponding both the alpha-helical or beta-sheet portions of the hypothetical Ia three-dimensional structure were involved. The results suggest that overall specificity of the T cell clones is the summation of numerous distinct subspecificities for different regions of the peptide-Ia ligand. These results indicate that there can be striking differences in T cell specificity for an autoantigenic epitope, even in the context of A alpha A beta molecules from very closely related haplotypes.
View details for Web of Science ID A1989AR48600001
View details for PubMedID 2476496
THE ROLE OF POLYMORPHIC I-AK BETA-CHAIN RESIDUES IN PRESENTATION OF A PEPTIDE FROM MYELIN BASIC-PROTEIN
JOURNAL OF EXPERIMENTAL MEDICINE
1989; 169 (6): 2239-2244
Proteins encoded by genes in the MHC are highly polymorphic. For class II proteins the highest level of polymorphism is found in distinct regions of variability, notably in the membrane-distal domains. To investigate the role of such residues in antigen presentation, we have tested cells transfected with wild-type or mutant I-Ak beta chains for their ability to present the NH2-terminal peptide of myelin basic protein to a panel of T cell clones. We were unable to detect a gross effect on peptide binding, in that all of the mutant cell lines presented antigen to at least one of the cloned T cells. However, the results imply that the more NH2-terminal residues, particularly 12 and 14, are involved in peptide interactions. Mutations at these residues presented antigen only at high antigen concentrations. Furthermore, residues of the more COOH-terminal regions appear to determine TCR interactions. Mutations in the predicted alpha-helical regions of the beta chain affected antigen presentation without abolishing peptide binding.
View details for Web of Science ID A1989AA89900030
View details for PubMedID 2471778
View details for PubMedCentralID PMC2189329
MOLECULAR DEFECTS IN THE NON-EXPRESSED H-2 E-ALPHA GENES OF THE F-HAPLOTYPES AND Q-HAPLOTYPES
JOURNAL OF IMMUNOLOGY
1989; 142 (8): 2936-2942
The expression and sequences of the mouse MHC class II genes, E alpha f and E alpha q, have been studied to determine the molecular basis for their defective expression. Previous work in our laboratory showed that H-2f and H-2q mice produce no detectable E alpha protein. Northern blot analysis confirms previous results showing normal amounts of E alpha f message and a 100-fold reduction in E alpha q mRNA. Despite that observation, the rates of transcription of both defective E alpha genes, measured by nuclear run-on transcription assays, are comparable to that of the normally expressed E alpha k gene. The nucleotide sequences of the E alpha f and E alpha q genes reveal mutations generating premature translation stop codons in both genes. A single base substitution has created the stop codon TGA at amino acid -2 in the E alpha f leader sequence. A nucleotide insertion at codon 64 in the second exon of the E alpha q gene results in a frame shift and a premature stop codon at amino acid residue 69 of a mature E alpha protein. The low steady state level of E alpha q mRNA may be correlated with the unusual size distribution of the RNA, possibly due to altered RNA processing.
View details for Web of Science ID A1989U213600051
View details for PubMedID 2701937
COMPLETE SEQUENCE OF THE MURINE INVARIANT CHAIN (II) GENE
NUCLEIC ACIDS RESEARCH
1989; 17 (1): 447-448
View details for Web of Science ID A1989R859800039
View details for PubMedID 2492095
DEFECTIVE E-BETA EXPRESSION IN 3 MOUSE H-2 HAPLOTYPES RESULTS FROM ABERRANT RNA SPLICING
JOURNAL OF IMMUNOLOGY
1988; 141 (10): 3647-3653
The molecular basis for the defective expression of the mouse class II E beta genes in the H-2w17, H-2q, and H-2f haplotypes has been examined. The results of nuclear run-on transcription and S1 nuclease digestion assays demonstrate that E beta transcription is normal in these haplotypes. Northern blot analyses reveal reduced amounts of E beta RNA of both normal and aberrant size in the w17 and q haplotypes; an even more reduced level of E beta RNA of normal size was detected in the f haplotype. In the preceding study, we reported that the only defect detected in the E beta w17 gene is a single nucleotide insertion in the 5' RNA splice site of the first intervening sequence. S1 nuclease analysis of E beta w17 RNA indicates that splicing at this site is aberrant. One major cryptic RNA splice site is used, leading to reduced amounts of aberrantly processed RNA. Limited use of the mutated splice site and of a second cryptic site also is detected. In all three cases, stop codons in the resulting RNA would prevent their translation. The molecular defect in E beta q appears identical to that of E beta w17. In the f haplotype, even more reduced levels of E beta RNA of both normal and aberrant sizes are found. We thus show that in the three defective E beta alleles, two distinct defects are responsible for the absence of E beta protein synthesis; both of these defects affect RNA processing.
View details for Web of Science ID A1988Q824700060
View details for PubMedID 2460544
MOLECULAR-BASIS FOR THE DEFECTIVE EXPRESSION OF THE MOUSE E-BETA-W17 GENE
JOURNAL OF IMMUNOLOGY
1988; 141 (10): 3654-3661
Four of the eleven independent H-2 haplotypes of inbred mouse strains and approximately 15% of wild mouse chromosomes 17 fail to express the E alpha E beta class II histocompatibility (Ia) Ag. These E- haplotypes are defective in the expression of the E alpha and/or the E beta chain. None of the E beta defects has previously been described at the molecular level. In this study, we report the molecular basis for the defective expression of the E beta gene from the w17 haplotype of the H-2 congenic strain B10.CAS2, derived from wild Mus musculus castaneus. Comparison of the Ew17 beta genomic sequence to those of the functional Eb beta and Ed beta genes reveals a single base insertion in the RNA donor splice site of the first intron. By DNA shuffling, we have corrected the single base mutation, and we show by FACS analysis and 2-D PAGE of immunoprecipitates that the corrected Ew17 beta is expressed in L cells when co-transfected with an Ed alpha gene. Conversely, an Eb beta gene construct containing the mutant RNA splice site from Ew17 beta is not expressed. We conclude that the single base insertion in the first RNA splice donor site is the sole molecular defect in the Ew17 beta gene.
View details for Web of Science ID A1988Q824700061
View details for PubMedID 2460545
HELPER T-CELLS SPECIFIC FOR PROTEIN ANTIGENS - ROLE OF SELF MAJOR HISTOCOMPATIBILITY COMPLEX AND IMMUNOGLOBULIN GENE-PRODUCTS
ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY
1982; 150: 53-71
View details for Web of Science ID A1982PJ77700005
View details for PubMedID 6293285
MOUSE IMMUNOGLOBULIN ALLOTYPES - CHARACTERIZATION AND USE IN CELLULAR IMMUNOLOGY
ANNALES D IMMUNOLOGIE
1974; C125 (1-2): 71-83
View details for Web of Science ID A1974R968600008
LYMPHOCYTE COMMITMENT TO IG ALLOTYPE AND CLASS
ANNALES D IMMUNOLOGIE
1974; C125 (1-2): 271-276
View details for Web of Science ID A1974R968600029