scholarly journals Antigenic Restimulation of Virus-Specific Memory CD8+ T Cells Requires Days of Lytic Protein Accumulation for Maximal Cytotoxic Capacity

2020 ◽  
Vol 94 (23) ◽  
Author(s):  
Stephen A. Migueles ◽  
Daniel C. Rogan ◽  
Noah V. Gavil ◽  
Elizabeth P. Kelly ◽  
Sushila A. Toulmin ◽  
...  
Keyword(s):  
T Cells ◽  
Protein Expression ◽  
Posttranscriptional Regulation ◽  
Granzyme B ◽  
Protein Accumulation ◽  
Mrna Levels ◽  
Memory Cells ◽  
Virus Infections ◽  
Prolonged Stimulation ◽  
Specific Memory

ABSTRACT In various infections or vaccinations of mice or humans, reports of the persistence and the requirements for restimulation of the cytotoxic mediators granzyme B (GrB) and perforin (PRF) in CD8+ T cells have yielded disparate results. In this study, we examined the kinetics of PRF and GrB mRNA and protein expression after stimulation and associated changes in cytotoxic capacity in virus-specific memory cells in detail. In patients with controlled HIV or cleared respiratory syncytial virus (RSV) or influenza virus infections, all virus-specific CD8+ T cells expressed low PRF levels without restimulation. Following stimulation, they displayed similarly delayed kinetics for lytic protein expression, with significant increases occurring by days 1 to 3 before peaking on days 4 to 6. These increases were strongly correlated with, but were not dependent upon, proliferation. Incremental changes in PRF and GrB percent expression and mean fluorescence intensity (MFI) were highly correlated with increases in HIV-specific cytotoxicity. mRNA levels in HIV-specific CD8+ T-cells exhibited delayed kinetics after stimulation as with protein expression, peaking on day 5. In contrast to GrB, PRF mRNA transcripts were little changed over 5 days of stimulation (94-fold versus 2.8-fold, respectively), consistent with posttranscriptional regulation. Changes in expression of some microRNAs, including miR-17, miR-150, and miR-155, suggested that microRNAs might play a significant role in regulation of PRF expression. Therefore, under conditions of extremely low or absent antigen levels, memory virus-specific CD8+ T cells require prolonged stimulation over days to achieve maximal lytic protein expression and cytotoxic capacity. IMPORTANCE Antigen-specific CD8+ T cells play a major role in controlling most virus infections, primarily by perforin (PRF)- and granzyme B (GrB)-mediated apoptosis. There is considerable controversy regarding whether PRF is constitutively expressed, rapidly increased similarly to a cytokine, or delayed in its expression with more prolonged stimulation in virus-specific memory CD8+ T cells. In this study, the degree of cytotoxic capacity of virus-specific memory CD8+ T cells was directly proportional to the content of lytic molecules, which required antigenic stimulation over several days for maximal levels. This appeared to be modulated by increases in GrB transcription and microRNA-mediated posttranscriptional regulation of PRF expression. Clarifying the requirements for maximal cytotoxic capacity is critical to understanding how viral clearance might be mediated by memory cells and what functions should be induced by vaccines and immunotherapies.

scholarly journals The route of priming influences the ability of respiratory virus–specific memory CD8+ T cells to be activated by residual antigen

2010 ◽  
Vol 207 (6) ◽  
pp. 1153-1160 ◽  
Author(s):  
Shiki Takamura ◽  
Alan D. Roberts ◽  
Dawn M. Jelley-Gibbs ◽  
Susan T. Wittmer ◽  
Jacob E. Kohlmeier ◽  
...  
Keyword(s):  
T Cells ◽  
Virus Infection ◽  
Cd8 T Cells ◽  
Respiratory Virus ◽  
Virus Infections ◽  
Memory Cd8 T Cells ◽  
Virus Clearance ◽  
Specific Memory ◽  
Respiratory Virus Infections ◽  
Lung Airways

After respiratory virus infections, memory CD8+ T cells are maintained in the lung airways by a process of continual recruitment. Previous studies have suggested that this process is controlled, at least in the initial weeks after virus clearance, by residual antigen in the lung-draining mediastinal lymph nodes (MLNs). We used mouse models of influenza and parainfluenza virus infection to show that intranasally (i.n.) primed memory CD8+ T cells possess a unique ability to be reactivated by residual antigen in the MLN compared with intraperitoneally (i.p.) primed CD8+ T cells, resulting in the preferential recruitment of i.n.-primed memory CD8+ T cells to the lung airways. Furthermore, we demonstrate that the inability of i.p.-primed memory CD8+ T cells to access residual antigen can be corrected by a subsequent i.n. virus infection. Thus, two independent factors, initial CD8+ T cell priming in the MLN and prolonged presentation of residual antigen in the MLN, are required to maintain large numbers of antigen-specific memory CD8+ T cells in the lung airways.

scholarly journals HIV-1 Viremia Prevents the Establishment of Interleukin 2–producing HIV-specific Memory CD4+ T Cells Endowed with Proliferative Capacity

2003 ◽  
Vol 198 (12) ◽  
pp. 1909-1922 ◽  
Author(s):  
Souheil-Antoine Younes ◽  
Bader Yassine-Diab ◽  
Alain R. Dumont ◽  
Mohamed-Rachid Boulassel ◽  
Zvi Grossman ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd4 T Cells ◽  
Cd4 T Cell ◽  
Memory Cells ◽  
Cell Responses ◽  
Specific Memory ◽  
Ifn Γ ◽  
Memory Cd4 T Cells

CD4+ T cell responses are associated with disease control in chronic viral infections. We analyzed human immunodeficiency virus (HIV)-specific responses in ten aviremic and eight viremic patients treated during primary HIV-1 infection and for up to 6 yr thereafter. Using a highly sensitive 5-(and-6)-carboxyfluorescein diacetate-succinimidyl ester–based proliferation assay, we observed that proliferative Gag and Nef peptide-specific CD4+ T cell responses were 30-fold higher in the aviremic patients. Two subsets of HIV-specific memory CD4+ T cells were identified in aviremic patients, CD45RA− CCR7+ central memory cells (Tcm) producing exclusively interleukin (IL)-2, and CD45RA− CCR7− effector memory cells (Tem) that produced both IL-2 and interferon (IFN)-γ. In contrast, in viremic, therapy-failing patients, we found significant frequencies of Tem that unexpectedly produced exclusively IFN-γ. Longitudinal analysis of HIV epitope–specific CD4+ T cells revealed that only cells that had the capacity to produce IL-2 persisted as long-term memory cells. In viremic patients the presence of IFN-γ–producing cells was restricted to periods of elevated viremia. These findings suggest that long-term CD4+ T cell memory depends on IL-2–producing CD4+ T cells and that IFN-γ only–producing cells are short lived. Our data favor a model whereby competent HIV-specific Tcm continuously arise in small numbers but under persistent antigenemia are rapidly induced to differentiate into IFN-γ only–producing cells that lack self-renewal capacity.

scholarly journals CD4 memory T cells survive and proliferate but fail to differentiate in the absence of CD40

2006 ◽  
Vol 203 (4) ◽  
pp. 897-906 ◽  
Author(s):  
Megan MacLeod ◽  
Mark J. Kwakkenbos ◽  
Alison Crawford ◽  
Sheila Brown ◽  
Brigitta Stockinger ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Memory T Cells ◽  
Cell Receptor ◽  
Memory Cells ◽  
Cell Stage ◽  
Effector Cells ◽  
Cell Responses ◽  
Histocompatibility Complex ◽  
Specific Memory

Secondary T cell responses are enhanced because of an expansion in numbers of antigen-specific (memory) cells. Using major histocompatibility complex class II tetramers we have tracked peptide-specific endogenous (non–T cell receptor transgenic) CD4 memory T cells in normal and in costimulation-deficient mice. CD4 memory T cells were detectable after immunization for more than 200 days, although decay was apparent. Memory cells generated in CD40 knockout mice by immunization with peptide-pulsed wild-type dendritic cells survived in the absence of CD40 and proliferated when boosted with peptide (plus adjuvant) in a CD40-independent fashion. However, differentiation of the memory cells into cytokine-producing effector cells did not occur in the absence of CD40. The data indicate that memory cells can be generated without passing through the effector cell stage.

scholarly journals Tumor-targeted delivery of childhood vaccine recall antigens by attenuated Listeria reduces pancreatic cancer

2019 ◽  
Author(s):  
Benson Chellakkan Selvanesan ◽  
Dinesh Chandra ◽  
Wilber Quispe-Tintaya ◽  
Arthee Jahangir ◽  
Ankur Patel ◽  
...  
Keyword(s):  
T Cells ◽  
Tumor Cells ◽  
T Cell Activation ◽  
Cd8 T Cells ◽  
Targeted Delivery ◽  
Cd4 T Cells ◽  
Granzyme B ◽  
Pancreatic Tumors ◽  
Specific Memory ◽  
Kpc Mice

ABSTRACTPancreatic ductal adenocarcinoma is highly metastatic, poorly immunogenic, and immune suppression prevents T cell activation. We developed a microbial-based immunotherapeutic concept for selective delivery of a highly immunogenic tetanus toxoid protein (TT856-1313) as an alternative for neoantigens, into tumor cells by attenuated Listeria monocytogenes, and reactivation of pre-existing TT-specific memory T cells (generated during childhood) to kill infected tumor cells. Thus, TT here functions as the tumor antigen. Treatment of KPC mice with Listeria-TT resulted in TT accumulation inside tumor cells, and attraction of the reactivated TT-specific memory CD4 T cells. Gemcitabine (GEM) combined with Listeria-TT significantly improved the migration of CD4 T cells into tumors and the production of perforin and granzyme B, turning cold into immunological hot tumors. The number of CD8 T cells in the KPC tumors was 3-fold lower than that of CD4 T cells. Moreover, lymph node like structures (LNS) were observed in close contact with the pancreatic tumors exhibiting CD4 T cells (and less abundantly CD8 T cells) of all treatment groups, but most frequently in KPC mice treated with Listeria-TT or Listeria-TT+GEM. Notably, the production of granzyme B (and less of perforin) was observed in the LNS of Listeria-TT+GEM only. The Listeria-TT+GEM treatment significantly reduced pancreatic tumors and metastases by 80% in Panc-02 and KPC mouse models, with minimal side effects. Our results unveil new mechanisms of Listeria and GEM improving immunotherapy for PDAC.

Mutations in CD58 and Other Immune System Related Genes in Hodgkin Lymphoma

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 1439-1439
Author(s):  
Fazlyn Reeny Abdul Razak ◽  
Arjan Diepstra ◽  
Lydia Visser ◽  
Anke Van den Berg
Keyword(s):  
Flow Cytometry ◽  
T Cells ◽  
Immune System ◽  
Protein Expression ◽  
Tumor Cells ◽  
Cell Lines ◽  
Hla Class I ◽  
Mrna Levels ◽  
Deleterious Mutations ◽  
Gm Csf

Abstract Hodgkin Lymphoma (HL) is a B cell derived malignancy characterized by a minority of tumor cells, known as Hodgkin Reed-Sternberg (HRS) cells. The background is composed of a wide variety of inflammatory cells with T cells representing the largest population. Chemokines and cytokines produced by HRS cells and by the infiltrating cells shape the environment and provide proliferative and survival signals to the HRS cells. Despite this critical dependence on the microenvironment, HRS cells also need to apply mechanisms to escape from both antigen-dependent and innate immune responses. HRS cells have evolved multiple mechanisms to evade cytotoxic T cell (CTL) and natural killer (NK) cell mediated anti-tumor responses. These mechanisms include secretion of immune-suppressive factors (IL10, TGFβ and others), recruitment of regulatory and helper T cells, expression of PDL1 and CD95 and loss of HLA expression. Recent publications show that mutations in immune system related genes might represent a mechanism of HRS cells to evade detection by immune cells. The aim of this study was to validate whole exome sequencing results of seven HL cell lines focusing on immune system associated genes. We previously showed that B2M mutations affect the ATG start codon in L428 (heterozygous) and DEV (homozygous) cells. B2M mRNA levels were reduced in both cell lines as compared to L1236, whereas HLA-A, HLA-B and HLA-C mRNA levels were in the same range. Consistent with these findings we observed no membranous B2M and HLA class I expression by flow cytometry in the two cell lines with mutated B2M genes. In primary diagnostic HL tissue we showed lack of membranous B2M in 51% of the cases. We now studied two additional genes in more detail. CD58 gene mutations were observed in KMH2 and DEV cells. By manual inspection of the alignments using the Integrative Genomics Viewer (IGV), we also noticed a lack of reads of exons 1, 2 and 3 in SUPHD1. Heterozygous mutations and homozygous loss of exons 1-3 were confirmed for all three cell lines. CD58 mRNA levels were low or absent in SUPHD1 and KMH2 cells and normal in DEV. CD58 protein expression as determined by flow, western blot and IHC was low or absent in all 3 mutated HL cell lines in comparison to four cell lines with wild type CD58. Tumor cells of 36 primary HL cases with good treatment outcome showed a strong CD58 expression in all cases. As HL cell lines are derived from end stage HL patients, we next studied CD58 expression in relapsed HL patients. No or weak CD58 staining was observed in HRS cells in 6 out of 45 patients who experienced a relapse. Our results indicate that mutations in CD58 and loss of CD58 expression are common in HL derived cell lines and that loss of CD58 expression in tumor cells is restricted to relapsed HL patients. Heterozygous CSF2RB mutations in KMH2, SUPHD1, DEV and L1236 were validated by RNA-seq and Sanger-seq. As CSF2RB encodes the common β chain (CD131) shared by the interleukin-3 (IL-3), granulocytic macrophage colony-stimulating factor (GM-CSF) and IL-5 receptors, we also measured the expression of these 3 α chain receptors. We observed the same expression pattern between CD131 and CD116 (GM-CSF α receptor chain) in HL cell lines by flow cytometry suggesting that these mutations mainly affect the GM-CSF receptor. In conclusion, we show that mutations of immune system genes are common in HL. Deleterious mutations in B2M explain the lack of HLA class I expression, indicating that this genetic alteration is responsible for defective antigen presentation. Deleterious mutations or deletions of CD58 exons result in loss of CD58 protein expression. This will lead to loss of binding to CD2 expressed on T cells and will result in a defect in T cell adhesion and activation. Overall these results indicate that mutations are likely to contribute to the immune escape mechanisms applied by the HRS cells. Disclosures No relevant conflicts of interest to declare.

scholarly journals Complex Memory T-Cell Phenotypes Revealed by Coexpression of CD62L and CCR7

2005 ◽  
Vol 79 (7) ◽  
pp. 4510-4513 ◽  
Author(s):  
Heike Unsoeld ◽  
Hanspeter Pircher
Keyword(s):  
T Cells ◽  
Lymphocytic Choriomeningitis ◽  
Effector Memory ◽  
Memory Cells ◽  
Memory Cd8 T Cells ◽  
Memory T Cell ◽  
Specific Memory ◽  
Major Shift ◽  
T Cell Phenotypes ◽  
Cell Phenotypes

ABSTRACT Antigen-experienced T cells have been divided into CD62L+ CCR7+ central memory (TCM) and CD62L− CCR7− effector memory (TEM) cells. Here, we examined coexpression of CD62L and CCR7 in lymphocytic choriomeningitis virus-specific memory CD8 T cells from both lymphoid and nonlymphoid tissues. Three main points emerged: firstly, memory cells frequently expressed a mixed CD62L− CCR7+ phenotype that differed from the phenotypes of classical TEM and TCM cells; secondly, TCM cells were not restricted to lymphoid organs but were also present in significant numbers in nonlymphoid tissues; and thirdly, a major shift from a TCM to TEM phenotype was found in memory cells that had been stimulated repetitively with antigen.

scholarly journals CD8+ T-Cell Dysfunction due to Cytolytic Granule Deficiency in Persistent Friend Retrovirus Infection

2005 ◽  
Vol 79 (16) ◽  
pp. 10619-10626 ◽  
Author(s):  
Gennadiy Zelinskyy ◽  
Shelly J. Robertson ◽  
Simone Schimmer ◽  
Ronald J. Messer ◽  
Kim J. Hasenkrug ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Granzyme B ◽  
Chronic Phase ◽  
Mrna Levels ◽  
Cell Dysfunction ◽  
Granzyme A ◽  
T Cell Dysfunction ◽  
In Vivo Cytotoxicity

ABSTRACT Virus-specific CD8+ T cells are critical for the control of acute Friend virus (FV) infections, but are rendered impotent by CD4+ regulatory T cells during the chronic phase of infection. The current study examines this CD8+ T-cell dysfunction by analyzing the production and release of cytolytic molecules by CD8+ T cells. CD8+ T cells with an activated phenotype (CD43+) from acutely infected mice produced all three key components of lytic granules: perforin, granzyme A, and granzyme B. Furthermore, they displayed evidence of recent degranulation and in vivo cytotoxicity. In contrast, activated CD8+ T cells from chronically infected mice were deficient in cytolytic molecules and showed little evidence of recent degranulation and poor in vivo cytotoxicity. Evidence from tetramer-positive CD8+ T cells with known virus specificity confirmed the findings from the activated subset of CD8+ T cells. Interestingly, perforin and granzyme A mRNA levels were not significantly reduced during chronic infection, indicating control at a posttranscriptional level. Granzyme B deficiency was associated with a significant decrease in mRNA levels, but posttranscriptional control also appeared to contribute to deficiency. These results demonstrate a broad impairment of cytotoxic CD8+ T-cell effector function during chronic retroviral infection and explain the inability of virus-specific CD8+ T cells to eliminate persistent virus.

Differential Regulation of Granzyme B and C Expression in Murine Cytotoxic T and NK Cells.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2291-2291
Author(s):  
Sheng F. Cai ◽  
Todd A. Fehniger ◽  
Xuezhi Dai ◽  
Xuefang Cao ◽  
Timothy J. Ley
Keyword(s):  
T Cells ◽  
Nk Cells ◽  
Cd8 T Cells ◽  
Nk Cell ◽  
Granzyme B ◽  
Transcriptional Level ◽  
Cytotoxic Lymphocytes ◽  
Mrna Levels ◽  
C Protein ◽  
Granzyme A

Abstract Cytotoxic lymphocytes, which include CD4+ and CD8+ T cells as well as NK cells, use the granule exocytosis pathway to kill virus-infected and tumor cells. Previous studies have characterized the expression of many genes in this pathway (e.g. perforin, granzyme A, granzyme B, etc.), although no reagent has been developed to measure granzyme C protein at the single-cell level. The murine granzyme C gene, orthologous to human granzyme H, lies 24.2Kb directly downstream from granzyme B in the granzyme B gene cluster. Recombinant granzyme C rapidly induces target cell death in a manner distinct from granzyme A- or B-induced death. To further understand the regulation and function of murine granzyme C, we developed a granzyme C-specific monoclonal antibody and used flow cytometry to examine the expression of granzyme C in resting and activated murine cytotoxic lymphocyte compartments. Naive CD4+ and CD8+ T cells express little or no granzyme C (0.1 + 0.1% of cells are positive). After activation of splenocytes with plate-bound CD3 and CD28 agonistic antibodies for 4 days, a small percentage of CD4+ (2.3 + 1.0% positive) and CD8+ (6.6 + 0.3% positive) T cells express granzyme C. However, only 24 hours later, almost all CD4+ (96.7 + 2.7%) and CD8+ (98.7 + 1.4%) T cells express granzyme C. Granzyme B co-staining revealed that granzyme B is detectable in both CD4+ and CD8+ T cells at least 48 hours before granzyme C is expressed. Furthermore, we employed a fully mismatched GVHD mouse model in order to examine T cell expression of granzymes B and C in vivo; similarly, granzyme B expression preceded granzyme C by at least 48 hours. In addition, CD4+Foxp3+ regulatory T cells also expressed granzyme C in this model. Murine NK cell granzyme C mRNA expression was assessed using Affymetrix microarrays (MOE430v2) at rest and following IL-15 activation. Resting NK cells express minimal granzyme C mRNA (mean gzmC probe set signal intensity + SD [n=3]): 356 + 202, which increased after IL-15 activation as follows: 4,180 + 1,106 (day 1), 60,788 + 8,455 (day 2), 167,448 + 26,398 (day 4), and 178,451+14,037 (day 6). Utilizing our granzyme C-specific mAb, we showed that very few resting murine NK cells express granzyme C protein (2.3 + 0.4% positive). Consistent with the mRNA data, the percentage of granzyme C-expressing NK cells was substantially increased after 3 days of IL-15 activation (88.8 + 5.2% positive). In contrast, granzyme B mRNA levels are high in resting NK cells (41,208 + 2,534), and increase only incrementally with IL-15 treatment (Fehniger et al., Immunity 2007 Jun;26(6):798–811). Granzyme B protein expression is controlled at a post-transcriptional level in NK cells, whereas granzyme C expression is transcriptionally regulated. Taken together, our findings show that the mouse granzyme B and C genes, despite being only 24.2Kb apart, are activated in cytotoxic lymphocytes with different kinetics; moreover, granzyme B and C protein abundance is controlled by completely different mechanisms in NK cells. These data suggest that the granzyme genes are differentially regulated in lymphocyte compartments, and that this regulation may be relevant for how cytotoxic lymphocytes function.

BACH2 Directly Regulates Expression of Foxp3 in UCB CD4+ T-Cells

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 4760-4760
Author(s):  
Mathew L. Lesniewski ◽  
Peter Haviernik ◽  
R. Patrick Weitzel ◽  
Laura Fanning ◽  
Marcie Finney ◽  
...  
Keyword(s):  
T Cells ◽  
Protein Expression ◽  
Cd4 T Cells ◽  
Foxp3 Expression ◽  
Mrna Levels ◽  
Western Blots ◽  
Foxp3 Mrna ◽  
Foxp3 Gene ◽  
Zip Family ◽  
Exon 1

Abstract Introduction: We have previously reported low constitutive nuclear factor of activated T cells (NFAT1) protein expression in unstimulated umbilical cord blood (UCB) CD4+ T-cells, and delayed up-regulation of NFAT1 expression during primary stimulation compared to adult primary CD4+ T-cells. Recent data has shown NFAT1 regulating Foxp3, an observed inverse expression of BACH2 and NFAT1 in UCB CD4+ T-cells, and our work demonstrating BACH2 directly regulates IL2 expression in the absence of NFAT1 protein. We sought to determine if BACH2 plays a role in regulating Foxp3 expression in UCB CD4+ T-cells. BACH2, a member of the b-Zip family, has been shown to act as a heterodimer with the bZip protein mafK, as a transcriptional inhibitor via recruitment of a histone deacetylase class II complex (HDAC II) in differentiating B-cells, and neurons. We hypothesized that BACH2 may directly regulate Foxp3 transcription by binding to putative NFAT1/AP1 binding sites within the Foxp3 proximal promoter. These sites may be vacant due to an absence of NFAT1 protein in resting UCB CD4+ T-cells. This hypothesis was tested by siRNA knockdown of BACH2 in primary UCB-derived CD4+ T-cells and then examining the level of Foxp3 expression at both the mRNA and protein level. In addition chromatin immunoprecipitation (ChIP) was performed in order to identify if BACH2 was binding to regions proximal to the Foxp3 gene. Finally Foxp3 driven luciferase plasmids were constructed and transiently transfected into primary UCB CD4+ T-cells with and without BACH2 siRNA and the relative expression determined. Methods: UCB T-cells were purified using autoMACs system. After overnight culture, T-cells were transfected with BACH2 siRNA using Amaxa nucleofector system. Both siRNA treated and control cells were incubated in media for 24 hours, and then stimulated using anti-CD3 and anti-CD28 antibodies. Aliquots of cells were collected at 24 hours and 30 hours post-stimulation for protein and total RNA isolation. The relative changes in mRNA levels for BACH2, FoxP3, were determined by Applied Biosystems Taqman real time RT-PCR system. Western blots were run to confirm results seen in the qRT-PCR. ChIP using BACH2 antibody, and PCR was used to determine putative BACH2 binding in DNA region surround exon 1 of Foxp3. Results: Loss of BACH2 expression correlates with reduced expression of (40-fold) FoxP3, mRNA in CD4+ UCB T-cells. Western blot performed on BACH2 siRNA treated, and anti-CD3/CD28 antibody stimulated UCB CD4+ T-cells; confirmed the observed loss of Foxp3 in unstimulated UCB CD4+ T-cells. Analysis of ChIP data from unstimulated UCB CD4+ T-cells showed that BACH2 and smafK bound to 2 regions flanking exon 1 of the Foxp3 gene, while results from unstimulated adult PB CD4+ T-cells showed BACH2 and smafK co-precipitated with only one region flanking exon 1 of Foxp3. Conclusions: NFAT1 protein has recently been shown to be required for the activation of Foxp3 in CD4+ T-cells, yet in UCB CD4+ T-cells NFAT1 protein expression is significantly attenuated. However, Foxp3 expression is unchanged in UCB CD4+ T-cells compared to adult PB CD4+ T-cells. UCB CD4+ T-cells BACH2 binds to two site surrounding exon 1 of Foxp3 gene, promoting the expression of Foxp3 mRNA in the absence of NFAT1 protein expression. These results suggest that expression of BACH2 in UCB CD4+ T-cells may underlie UCB immune tolerance in the unrelated allogeneic setting via regulation of FoxP3 impacting Treg development.

scholarly journals The Herpesvirus saimiri Small Nuclear RNAs Recruit AU-Rich Element-Binding Proteins but Do Not Alter Host AU-Rich Element-Containing mRNA Levels in Virally Transformed T Cells

2004 ◽  
Vol 24 (10) ◽  
pp. 4522-4533 ◽  
Author(s):  
Heidi L. Cook ◽  
Hannah E. Mischo ◽  
Joan A. Steitz
Keyword(s):  
T Cells ◽  
Posttranscriptional Regulation ◽  
Binding Proteins ◽  
Herpesvirus Saimiri ◽  
Mrna Levels ◽  
Small Nuclear Rna ◽  
Small Nuclear Rnas ◽  
Latently Infected ◽  
The Stability

ABSTRACT Herpesvirus saimiri (HVS) encodes seven Sm-class small nuclear RNAs, called HSURs (for Herpesvirus saimiri U RNAs), that are abundantly expressed in HVS-transformed, latently infected marmoset T cells but are of unknown function. HSURs 1, 2, and 5 have highly conserved 5′-end sequences containing the AUUUA pentamer characteristic of AU-rich elements (AREs) that regulate the stability of many host mRNAs, including those encoding most proto-oncogenes and cytokines. To test whether the ARE-containing HSURs act to sequester host proteins that regulate the decay of these mRNAs, we demonstrate their in vivo interaction with the ARE-binding proteins hnRNP D and HuR in HVS-transformed T cells using a new cross-linking assay. Comprehensive Northern and microarray analyses revealed, however, that the levels of endogenous ARE-containing mRNAs are not altered in T cells latently infected with HVS mutants lacking HSURs 1 and 2. HSUR 1 binds the destabilizing ARE-binding protein tristetraprolin induced following activation of HVS-transformed T cells, but even in such stimulated cells, the levels of host ARE-containing mRNAs are not altered by deletion of HSURs 1 and 2. Instead, HSUR 1 itself is degraded by an ARE-dependent pathway in HVS-transformed T cells, suggesting that HVS may take advantage of the host ARE-mediated mRNA decay pathway to regulate HSUR expression. This is the first example of posttranscriptional regulation of the expression of an Sm small nuclear RNA.

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