scholarly journals The Role of LAMP5 in Innate Immune Signaling Is Critical for the Survival of MLL Leukemias

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 3900-3900
Author(s):  
Gabriel Gracia-Maldonado ◽  
Jason Clark ◽  
Philippe Pierre ◽  
James C. Mulloy ◽  
Ashish R Kumar
Keyword(s):  
Signaling Pathway ◽  
Leukemia Cell ◽  
Innate Immune ◽  
Leukemia Cell Line ◽  
Therapeutic Window ◽  
Type I ◽  
Tlr Signaling ◽  
Immune Signaling ◽  
Over Expression ◽  
Innate Immune Signaling

Abstract Introduction Chromosomal translocations involving the Mixed Lineage Leukemia (MLL, also called KMT2A) gene account for around 80% of the acute lymphoid leukemia (ALL) and 35%-50% of the acute myeloid leukemia (AML) diagnosed in infants, and are associated with poor outcomes. Despite improvements of conventional chemotherapies and treatments, patients with MLL-rearranged leukemia have weak response to treatment and poor prognosis. There is thus an urgent need to understand the molecular pathogenesis of these leukemias to develop more effective therapies. Gene expression studies show that MLL-leukemias uniquely display over-expression of Lysosome-associated Membrane Protein 5 (LAMP5) in both ALL and AML In normal hematopoiesis, LAMP5 expression is restricted to non-activated plasmacytoid dendritic cells (pDC), where it has been shown to be required for transport of TLR9 from VAMP3+/LAMP2+/LAMP1- endolysosomal vesicles to the LAMP1+ late lysosomal compartments, modulating the signaling and production of Type I interferon to TNF production. Here, we tested the hypothesis that LAMP5 is required for MLL leukemia wherein it modulates innate immune signaling. Methods and Results In previous studies, we found LAMP5 expression being restricted to MLL-fusion leukemia cell lines. Additionally, in an inducible model, LAMP5 expression directly correlated with that of the MLL-fusion protein MLL-AF9. . Knockdown on LAMP5 in MLL-fusion leukemia cells inhibited their propagation both in vitro and in vivo whereas the non-MLL leukemias were unaffected. To determine the mechanisms by which LAMP5 promotes growth in MLL leukemias, first we studied the localization of LAMP5 in these cells. Using immunofluorescent confocal microscopy we detected LAMP5 mainly in LAMP2+/LAMP1+ compartments. Additionally, LAMP5 co-localized with MYD88, a known scaffold protein required for IL1R/TLR signaling. Accordingly, LAMP5-knockdown abrogated TLR signaling as evidenced by reduced activation of JNK, MAPK p38, IRF7 and NFKB. Conversely, over-expression of LAMP5 in the non-MLL leukemia cell line Kasumi-1 led to increased activation of JNK, p38, IRF7 and NFKB, indicating that the innate-immune signaling pathway is regulated by LAMP5 expression in leukemias.r Conclusions Collectively, these results demonstrate that LAMP5 is required for the survival of MLL leukemia and that it plays an important role in the activation of the IL1/TLR signaling pathway. Overall, based on our results and the limited expression in normal hematopoiesis, we propose that LAMP5 could potentially serve as a therapeutic target with a wide therapeutic-window to treat MLL-leukemias. Disclosures No relevant conflicts of interest to declare.

scholarly journals MAVS: A Two-Sided CARD Mediating Antiviral Innate Immune Signaling and Regulating Immune Homeostasis

2021 ◽  
Vol 12 ◽  
Author(s):  
Yunqiang Chen ◽  
Yuheng Shi ◽  
Jing Wu ◽  
Nan Qi
Keyword(s):  
Signal Transduction ◽  
Innate Immune ◽  
Immune Homeostasis ◽  
Type I ◽  
Interferon Signaling ◽  
Immune Signaling ◽  
Post Translational Modifications ◽  
Innate Immune Signaling ◽  
Mitochondrial Antiviral Signaling ◽  
Mitochondrial Antiviral Signaling Protein

Mitochondrial antiviral signaling protein (MAVS) functions as a “switch” in the immune signal transduction against most RNA viruses. Upon viral infection, MAVS forms prion-like aggregates by receiving the cytosolic RNA sensor retinoic acid-inducible gene I-activated signaling and further activates/switches on the type I interferon signaling. While under resting state, MAVS is prevented from spontaneously aggregating to switch off the signal transduction and maintain immune homeostasis. Due to the dual role in antiviral signal transduction and immune homeostasis, MAVS has emerged as the central regulation target by both viruses and hosts. Recently, researchers show increasing interest in viral evasion strategies and immune homeostasis regulations targeting MAVS, especially focusing on the post-translational modifications of MAVS, such as ubiquitination and phosphorylation. This review summarizes the regulations of MAVS in antiviral innate immune signaling transduction and immune homeostasis maintenance.

scholarly journals IMMU-34. ATRX MUTATIONS PREDICT RESPONSE TO INNATE BASED THERAPY IN GLIOMA

2019 ◽  
Vol 21 (Supplement_6) ◽  
pp. vi126-vi126
Author(s):  
Michelle Bowie ◽  
Seethalakshmi Hariharan ◽  
Janell Hostettler ◽  
Kristen Roso ◽  
Yiping He ◽  
...  
Keyword(s):  
Cell Lines ◽  
Glioma Cell ◽  
Innate Immune ◽  
Type I ◽  
Loss Of Function ◽  
Who Grade ◽  
Immune Signaling ◽  
Innate Immune Signaling ◽  
Recurrent Mutations ◽  
Glioma Cell Lines

Abstract BACKGROUND Innate based immunotherapies are becoming increasingly important for treating brain tumor patients. Gliomas carry recurrent mutations in regulatory genes that control innate immune signaling responses. About 71% of adult WHO grade II and III gliomas and 57% of secondary glioblastomas also carry a loss-of-function mutation in the ATRX gene. ATRX is a SWI-SNF chromatin remodeling protein that has major roles in processes such as cell cycle regulation and maintenance of genomic stability. Recent studies have implicated ATRX in dysfunctional innate immune signaling in cancer cells. However, the role of ATRX in mediating innate immune responses has not been investigated in gliomas. METHODS AND RESULTS Human and mouse glioma cell lines from a variety of genetic contexts have been examined including models which carry IDH/ATRX mutations, IDH 1p-/19q- and ATRX -/- status. Additionally, using Crispr-Cas9 technology and cloning cell lines with ATRX deletions, we have derived a series of immune competent and nude mice models. Treating these cell lines with double-stranded RNA based innate stimuli led to an enhanced early induction in phospho-interferon regulatory factor 3 (IRF3) and late induction in phospho-STAT1 in the ATRX knockout (KO) cell lines. A differential increase in interferon-stimulated gene 15 (ISG15) release was also noted in the ATRX KO cell lines, further suggesting that ATRX deletion may enable a potent activation of type I interferon production. A combination of patient-derived glioma cell lines in xenograft models and syngeneic murine glioma models derived from ATRX KO cell lines and controls confirm a survival advantage in both immuno-competent mice and xenografts. Our models are under evaluation with PVSRIPO and other innate based RNA therapies. CONCLUSION Our data suggests that ATRX mutations may confer sensitivity to RNA-based innate immune signaling agonists in gliomas. This potential vulnerability can be targeted in future therapies.

scholarly journals STING signaling and host defense against microbial infection

2019 ◽  
Vol 51 (12) ◽  
pp. 1-10 ◽  
Author(s):  
Jeonghyun Ahn ◽  
Glen N. Barber
Keyword(s):  
Host Defense ◽  
Innate Immune ◽  
Type I Interferons ◽  
Type I ◽  
Microbial Infection ◽  
Immune Signaling ◽  
Pathogen Invasion ◽  
Innate Immune Signaling ◽  
Specific Pathogen ◽  
Microbial Dna

AbstractThe first line of host defense against infectious agents involves activation of innate immune signaling pathways that recognize specific pathogen-associated molecular patterns (PAMPs). Key triggers of innate immune signaling are now known to include microbial-specific nucleic acid, which is rapidly detected in the cytosol of the cell. For example, RIG-I-like receptors (RLRs) have evolved to detect viral RNA species and to activate the production of host defense molecules and cytokines that stimulate adaptive immune responses. In addition, host defense countermeasures, including the production of type I interferons (IFNs), can also be triggered by microbial DNA from bacteria, viruses and perhaps parasites and are regulated by the cytosolic sensor, stimulator of interferon genes (STING). STING-dependent signaling is initiated by cyclic dinucleotides (CDNs) generated by intracellular bacteria following infection. CDNs can also be synthesized by a cellular synthase, cGAS, following interaction with invasive cytosolic self-DNA or microbial DNA species. The importance of STING signaling in host defense is evident since numerous pathogens have developed strategies to prevent STING function. Here, we review the relevance of STING-controlled innate immune signaling in host defense against pathogen invasion, including microbial endeavors to subvert this critical process.

scholarly journals STAT5: a Target of Antagonism by Neurotropic Flaviviruses

2019 ◽  
Vol 93 (23) ◽  
Author(s):  
Matthew G. Zimmerman ◽  
James R. Bowen ◽  
Circe E. McDonald ◽  
Ellen Young ◽  
Ralph S. Baric ◽  
...  
Keyword(s):  
Immune Responses ◽  
Innate Immune ◽  
Type I ◽  
Molecular Signatures ◽  
Zikv Infection ◽  
Immune Signaling ◽  
Gm Csf ◽  
Innate Immune Signaling ◽  
Significant Public Health ◽  
Dengue Virus Serotypes

ABSTRACT Flaviviruses are a diverse group of arthropod-borne viruses responsible for numerous significant public health threats; therefore, understanding the interactions between these viruses and the human immune response remains vital. West Nile virus (WNV) and Zika virus (ZIKV) infect human dendritic cells (DCs) and can block antiviral immune responses in DCs. Previously, we used mRNA sequencing and weighted gene coexpression network analysis (WGCNA) to define molecular signatures of antiviral DC responses following activation of innate immune signaling (RIG-I, MDA5, or type I interferon [IFN] signaling) or infection with WNV. Using this approach, we found that several genes involved in T cell cosignaling and antigen processing were not enriched in DCs during WNV infection. Using cis-regulatory sequence analysis, STAT5 was identified as a regulator of DC activation and immune responses downstream of innate immune signaling that was not activated during either WNV or ZIKV infection. Mechanistically, WNV and ZIKV actively blocked STAT5 phosphorylation downstream of RIG-I, IFN-β, and interleukin-4 (IL-4), but not granulocyte-macrophage colony-stimulating factor (GM-CSF), signaling. Unexpectedly, dengue virus serotypes 1 to 4 (DENV1 to DENV4) and the yellow fever 17D vaccine strain (YFV-17D) did not antagonize STAT5 phosphorylation. In contrast to WNV, ZIKV inhibited JAK1 and TYK2 phosphorylation following type I IFN treatment, suggesting divergent mechanisms used by these viruses to inhibit STAT5 activation. Combined, these findings identify STAT5 as a target of antagonism by specific pathogenic flaviviruses to subvert the immune response in infected DCs. IMPORTANCE Flaviviruses are a diverse group of insect-borne viruses responsible for numerous significant public health threats. Previously, we used a computational biology approach to define molecular signatures of antiviral DC responses following activation of innate immune signaling or infection with West Nile virus (WNV). In this work, we identify STAT5 as a regulator of DC activation and antiviral immune responses downstream of innate immune signaling that was not activated during either WNV or Zika virus (ZIKV) infection. WNV and ZIKV actively blocked STAT5 phosphorylation downstream of RIG-I, IFN-β, and IL-4, but not GM-CSF, signaling. However, other related flaviviruses, dengue virus serotypes 1 to 4 and the yellow fever 17D vaccine strain, did not antagonize STAT5 phosphorylation. Mechanistically, WNV and ZIKV showed differential inhibition of Jak kinases upstream of STAT5, suggesting divergent countermeasures to inhibit STAT5 activation. Combined, these findings identify STAT5 as a target of antagonism by specific pathogenic flaviviruses to subvert antiviral immune responses in human DCs.

Stimulator of Interferon Genes Signaling Pathway and its Role in Anti-tumor Immune Therapy

2020 ◽  
Vol 26 (26) ◽  
pp. 3085-3095 ◽  
Author(s):  
Yuanjin Gong ◽  
Chang Chang ◽  
Xi Liu ◽  
Yan He ◽  
Yiqi Wu ◽  
...  
Keyword(s):  
Signaling Pathway ◽  
Immune Therapy ◽  
Signal Transduction Pathway ◽  
The Body ◽  
Innate Immune ◽  
Type I ◽  
Innate Immune Signaling ◽  
Critical Problems ◽  
Molecular Patterns ◽  
Damage Associated Molecular Patterns

Stimulator of interferon genes is an important innate immune signaling molecule in the body and is involved in the innate immune signal transduction pathway induced by pathogen-associated molecular patterns or damage-associated molecular patterns. Stimulator of interferon genes promotes the production of type I interferon and thus plays an important role in the innate immune response to infection. In addition, according to a recent study, the stimulator of interferon genes pathway also contributes to anti-inflammatory and anti-tumor reactions. In this paper, current researches on the Stimulator of interferon genes signaling pathway and its relationship with tumor immunity are reviewed. Meanwhile, a series of critical problems to be addressed in subsequent studies are discussed as well.

CS17-5. Arteri- and nairovirus OTU domain-containing proteases target ubiquitin-regulated factors in the RLR-mediated innate immune signaling pathway

Cytokine ◽  
2011 ◽  
Vol 56 (1) ◽  
pp. 108
Author(s):  
Marjolein Kikkert ◽  
Puck B. van Kasteren ◽  
Corrine Beugeling ◽  
Dennis Ninaber ◽  
Sander van Boheemen ◽  
...  
Keyword(s):  
Signaling Pathway ◽  
Innate Immune ◽  
Immune Signaling ◽  
Innate Immune Signaling ◽  
Immune Signaling Pathway ◽  
Innate Immune Signaling Pathway

scholarly journals Dependency of Spliceosomal Mutant MDS on Innate Immune Signaling

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 937-937
Author(s):  
Stanley C Lee ◽  
Khrystyna North ◽  
Chi-Chao Chen ◽  
Eunhee Kim ◽  
Sydney X. Lu ◽  
...  
Keyword(s):  
Cell Survival ◽  
Myeloid Cells ◽  
K562 Cells ◽  
Genetic Alterations ◽  
Innate Immune ◽  
Type I ◽  
Immune Signaling ◽  
Innate Immune Signaling ◽  
Genes Encoding ◽  
Mutant Cells

Abstract Mutations in genes encoding RNA splicing factors constitute the most common class of genetic alterations in patients with myelodysplastic syndromes (MDS). These occur as heterozygous point mutations at specific amino acid residues in SF3B1, SRSF2, and U2AF1, and are almost always mutually exclusive with one another. Recent studies have identified that mutations in each of these genes results in activation of the innate immune signaling through altered splicing of mRNAs encoding key enzymes in this pathway. Now, through an unbiased genetic screen as well as focused genetic studies, we have identified that SF3B1-mutant MDS depends on aberrant immune signaling for cell survival. Recent work has identified that aberrant splicing of MAP3K7 (also known as TAK1; TGF-b Activating Kinase 1) is pervasive across SF3B1-mutant human and mouse cells and results in reduced MAP3K7 protein expression and increased NF-κB signaling. Consistent with this, Map3k7 haploinsufficiency in myeloid cells is known to cause myeloproliferation, while at the same time, complete loss of Map3k7 is intolerable for hematopoietic cells. We therefore hypothesized that partial inhibition of MAP3K7 might preferentially impact SF3B1-mutant cells. To test this hypothesis, we generated mice with inducible deletion of 1 or 2 copies of Map3k7 (Mx1-cre Map3k7fl/+,Mx1-cre Map3k7fl/fl) alone or in the presence of mutant Sf3b1K700E (Mx1-cre Map3k7fl/+Sf3b1K700E/+,Mx1-cre Map3k7fl/flSf3b1K700E/+), along with all controls (Mx1-cre Sf3b1+/+ Map3k7+/+ (Wildtype; WT) and Mx1-cre Sf3b1K700E/+ mice). We then performed bone marrow transplantation (BMT) to assess the effect of Map3k7 deletion on aberrant hematopoiesis driven by mutant SF3B1. Consistent with prior reports, heterozygous deletion of Map3k7did not affect repopulating potential in BMT assays compared to controls while homozygous deletion of Map3k7 resulted in complete failure of hematopoiesis (Figure A). Interestingly, however, in the presence of Sf3b1K700E mutation, deletion of a single copy of Map3k7 completely rescued the hematopoietic defects characteristic of mutant SF3B1 in both mature and immature cells (Figure B-C). These data suggest that inhibition of residual MAP3K7 function may preferentially target SF3B1-mutant MDS cells. In parallel to the above studies, we also performed a negative selection RNAi screen to uncover novel genetic dependencies in SF3B1-mutant myeloid neoplasms. We performed pooled lentiviral infection of shRNAs targeting ~2,200 genes encoding proteins which are drug targets ("The Druggable Genome") under the control of a doxycycline-inducible vector in isogenic K562 cells expressing the two most commonly occurring SF3B1 mutations, SF3B1K666N and SF3B1K700E, from the endogenous SF3B1 locus. Two individual clones per SF3B1-mutant line were used to improve the robustness of the screen. On Day 21 following shRNA activation, genes with ≥3 shRNAs depleted in SF3B1-mutant cells while remaining unchanged in parental K562 cells were selected. This identified 101 candidates that are potentially synthetic-lethal with SF3B1 mutation (Figure D). Interestingly, pathway analysis of these potential candidates revealed of genes involved in immune and inflammatory signaling as well as in metabolic processes (Figure E). Further target validation was performed using in vitro competitive growth assay in K562 cells, and another set of SF3B1 isogenic lymphoid leukemia cell lines (NALM-6) expressing the same mutations. This revealed consistent dependency of SF3B1-mutant cells on STAT1, an essential component of the interferon (IFN) signaling pathway (Figure F). Upon exposure to Type-I IFNs, SF3B1-mutant K562 cells showed increased transcriptional response in IFN-responsive genes containing interferon stimulated response elements (ISREs) compared with SF3B1 WT cells (Figure G). These data highlight that SF3B1-mutant cells are hyper-responsive to IFN signaling and require intact IFN-signaling responses for cell survival. Taken together, the above studies indicate that sustained IFN signaling as well as activated innate immune signaling downstream of TAK1 are required for the survival of SF3B1-mutant myeloid cells. These results therefore have important therapeutic implications as they suggest that pharmacologic inhibition of STAT1/Type I IFN activation and/or TAK1 may serve as important therapeutic agents for SF3B1-mutant MDS. Figure. Figure. Disclosures No relevant conflicts of interest to declare.

scholarly journals Alternative Splice Variants of IRAK4 That Activate Innate Immune Signaling Are Associated with U2AF1 Mutations in Myelodysplastic Syndrome and Acute Myeloid Leukemia

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 1531-1531
Author(s):  
Molly A Smith ◽  
Kwangmin Choi ◽  
Nathan Salomonis ◽  
Matthew J. Walter ◽  
Kakajan Komurov ◽  
...  
Keyword(s):  
Rna Splicing ◽  
Myeloid Leukemia ◽  
Innate Immune ◽  
Tlr Signaling ◽  
Immune Signaling ◽  
Normal Tissues ◽  
Innate Immune Signaling ◽  
Alternatively Spliced ◽  
Exon 4 ◽  
Isoform Expression

Abstract Alternative RNA splicing and mutations in spliceosome genes are common features of human cancer. For example, recurring mutations of spliceosome genes in Myelodysplastic Syndrome (MDS) and Acute Myeloid Leukemia (AML) most frequently involve U2AF1, SF3B1, and SRSF2. Although aberrant RNA splicing is implicated in the pathogenesis of human cancers, it is less understood which misspliced genes contribute the malignant state. A global analysis of alternatively spliced genes and RNA isoforms in breast cancer, lung cancer, and AML revealed enrichment of alternatively spliced genes associated with inflammatory and immune pathways in the cancer cells as compared to the respective normal tissues. One such example is cancer-specific isoform expression of Interleukin Receptor Associated Kinase 4 (IRAK4), a serine/threonine kinase downstream of toll-like receptor (TLR) signaling and implicated in the pathogenesis of MDS/AML (Rhyasen et al., Cancer Cell 2013). Examination of the spliced isoforms by RNA sequencing showed that normal tissues preferentially express an alternatively spliced isoform of IRAK4 resulting from exclusion of the exon 4 (IRAK4-Short). In contrast, the majority of MDS and AML samples exclusively express an IRAK4 isoform that includes exon 4 (IRAK4-Long). Skipping of IRAK4 exon 4 results in an in-frame deletion of the N-terminal death domain, which is required for IRAK4 oligomerization and efficient TLR signaling, yet retains its C-terminal kinase domain. Immunoblotting confirmed that MDS/AML samples predominantly express the IRAK4-Long protein, while normal hematopoietic BM cells express the IRAK4-Short protein lacking the N-terminal domain. IRAK4-Long expression is significantly associated with increased NF-kB and innate immune signaling and correlates with poor AML patient outcome. Functional characterization of the IRAK4 isoforms in human AML cell lines revealed that IRAK4-Long induces NF-kB activation. In contrast, IRAK4-Short is less efficient at activating NF-kB (via phosphorylation of IKKbeta), yet it activates p38/MAPK signaling. To gain insight into the alternative splicing regulation of IRAK4 exon 4, we examined IRAK4 isoform expression and associated spliceosome gene mutations in MDS/AML patients. Of all examined genetic associations, mutation of U2AF1 (S34F) significantly correlated with inclusion of exon 4 and expression of IRAK4-Long, suggesting that mutations in U2AF1 instruct expression of IRAK4 RNA isoforms with maximal functional potential. To explore the direct regulation of IRAK4 by U2AF1, wildtype or mutant (S34F) U2AF1 were expressed in CD34+ cord blood cells, and IRAK4 isoform expression and exon usage was determined by RNA-sequencing. Expression of U2AF1-S34F resulted in significant retention of IRAK4 exon 4 (i.e. IRAK4-Long), while expression of wildtype U2AF1 correlated with exclusion of IRAK4 exon 4 (i.e., IRAK4-Short). Moreover, exon 4 and flanking intron sequences were cloned into a splicing reporter. Overexpression of U2AF1-S34F induced retention of the cassette exon 4, while wild-type U2AF1 mediated exclusion of exon 4. Taken together, these findings illustrate the importance of cancer-associated RNA splicing alterations and their consequences on downstream molecular networks required for cancer pathogenesis. In addition, we find that U2AF1 mutations result in expression of IRAK4 isoforms providing a genetic link to chronic innate immune signaling and IRAK1/4 activation in MDS and AML. Disclosures No relevant conflicts of interest to declare.

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