Publications by authors named "Nimi Marcel"

13 Publications

  • Page 1 of 1

CopyCatchers are versatile active genetic elements that detect and quantify inter-homolog somatic gene conversion.

Nat Commun 2021 05 11;12(1):2625. Epub 2021 May 11.

Section of Cell and Developmental Biology, University of California San Diego, La Jolla, CA, USA.

CRISPR-based active genetic elements, or gene-drives, copied via homology-directed repair (HDR) in the germline, are transmitted to progeny at super-Mendelian frequencies. Active genetic elements also can generate widespread somatic mutations, but the genetic basis for such phenotypes remains uncertain. It is generally assumed that such somatic mutations are generated by non-homologous end-joining (NHEJ), the predominant double stranded break repair pathway active in somatic cells. Here, we develop CopyCatcher systems in Drosophila to detect and quantify somatic gene conversion (SGC) events. CopyCatchers inserted into two independent genetic loci reveal unexpectedly high rates of SGC in the Drosophila eye and thoracic epidermis. Focused RNAi-based genetic screens identify several unanticipated loci altering SGC efficiency, one of which (c-MYC), when downregulated, promotes SGC mediated by both plasmid and homologous chromosome-templates in human HEK293T cells. Collectively, these studies suggest that CopyCatchers can serve as effective discovery platforms to inform potential gene therapy strategies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-021-22927-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8113449PMC
May 2021

FOXO1 constrains activation and regulates senescence in CD8 T cells.

Cell Rep 2021 Jan;34(4):108674

Division of Biological Sciences, Molecular Biology Section, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0377, USA; Department of Cellular and Molecular Medicine, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0377, USA. Electronic address:

Naive and memory T cells are maintained in a quiescent state, yet capable of rapid response and differentiation to antigen challenge via molecular mechanisms that are not fully understood. In naive cells, the deletion of Foxo1 following thymic development results in the increased expression of multiple AP-1 family members, rendering T cells less able to respond to antigenic challenge. Similarly, in the absence of FOXO1, post-infection memory T cells exhibit the characteristics of extended activation and senescence. Age-based analysis of human peripheral T cells reveals that levels of FOXO1 and its downstream target, TCF7, are inversely related to host age, whereas the opposite is found for AP-1 factors. These characteristics of aging also correlate with the formation of T cells manifesting features of cellular senescence. Our work illustrates a role for FOXO1 in the active maintenance of stem-like properties in T cells at the timescales of acute infection and organismal life span.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.celrep.2020.108674DOI Listing
January 2021

A key control point in the T cell response to chronic infection and neoplasia: FOXO1.

Curr Opin Immunol 2020 04 2;63:51-60. Epub 2020 Mar 2.

Molecular Biology Section, Division of Biological Sciences, Department of Cellular and Molecular Medicine, TATA Institute for Genetics and Society, University of California, San Diego, La Jolla, CA 92093-0377, United States. Electronic address:

T cells able to control neoplasia or chronic infections display a signature gene expression profile similar or identical to that of central memory T cells. These cells have qualities of self-renewal and a plasticity that allow them to repeatedly undergo activation (growth, proliferation, and differentiation), followed by quiescence. It is these qualities that define the ability of T cells to establish an equilibrium with chronic infectious agents, and also preserve the ability of T cells to be re-activated (by checkpoint therapy) in response to malignant cancers. Here we describe distinctions between the forms of inhibition mediated by tumors and persistent viruses, we review the properties of T cells associated with long-term immunity, and we identify the transcription factor, FOXO1, as the control point for a program of gene expression that allows CD8 T cells to undergo serial reactivation and self-renewal.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.coi.2020.02.001DOI Listing
April 2020

The lysine deacetylase Sirtuin 1 modulates the localization and function of the Notch1 receptor in regulatory T cells.

Sci Signal 2017 Apr 4;10(473). Epub 2017 Apr 4.

National Centre for Biological Sciences, Bengaluru, Karnataka 560065, India.

The ability to tune cellular functions in response to nutrient availability has important consequences for immune homeostasis. The activity of the receptor Notch in regulatory T (T) cells, which suppress the functions of effector T cells, is indispensable for T cell survival under conditions of diminished nutrient supply. Anti-apoptotic signaling induced by the Notch1 intracellular domain (NIC) originates from the cytoplasm and is spatially decoupled from the nuclear, largely transcriptional functions of NIC. We showed that Sirtuin 1 (Sirt1), which is an NAD (nicotinamide adenine dinucleotide)-dependent lysine deacetylase that inhibits NIC-dependent gene transcription, stabilized NIC proximal to the plasma membrane to promote the survival and function of activated T cells. Sirt1 was required for NIC-dependent protection from apoptosis in cell lines but not for the activity of the anti-apoptotic protein Bcl-xL. In addition, a variant NIC protein in which four lysines were mutated to arginines (NIC4KR) retained anti-apoptotic activity, but was not regulated by Sirt1, and reconstituted the functions of nonnuclear NIC in Notch1-deficient T cells. Loss of Sirt1 compromised T cell survival, resulting in antigen-induced T cell proliferation and inflammation in two mouse models. Thus, the Sirt1-Notch interaction may constitute an important checkpoint that tunes noncanonical Notch1 signaling.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1126/scisignal.aah4679DOI Listing
April 2017

The NOTCH1-autophagy interaction: Regulating self-eating for survival.

Autophagy 2017 Feb 27;13(2):446-447. Epub 2016 Dec 27.

a Institute for Stem Cell Biology & Regenerative Medicine , Bengaluru , Karnataka , India.

T-cell subsets in the mammalian immune system use varied mechanisms for survival, a demand imposed by the diverse and dynamic niches that they function in. In a recent study, we showed that survival of natural T-regulatory cells (Tregs) was determined by spatially regulated NOTCH1 activity signaling leading to the activation of macroautophagy/autophagy. While this interaction was revealed in experimental conditions of limited nutrient availability in vitro, the consequences of this interaction were confirmed in the context of immune physiology. Consistently, disrupting NOTCH signaling or the autophagy cascade was deleterious to Tregs. At the molecular level, ligand-activated NOTCH1, which is enriched outside the nucleus in Tregs, was detected in complexes that included specific molecular intermediates controlling the progression of autophagy. Mitochondria were a prominent cellular target, with organelle remodeling and function dependent on NOTCH1 signaling to autophagy. It is tempting to speculate that the link between autophagy and the developmental regulator NOTCH1 identified in this work may be conserved in other biological contexts.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1080/15548627.2016.1268303DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5324846PMC
February 2017

Notch1 regulated autophagy controls survival and suppressor activity of activated murine T-regulatory cells.

Elife 2016 06 6;5. Epub 2016 Jun 6.

National Centre for Biological Sciences, Bengaluru, India.

Cell survival is one of several processes regulated by the Notch pathway in mammalian cells. Here we report functional outcomes of non-nuclear Notch signaling to activate autophagy, a conserved cellular response to nutrient stress, regulating survival in murine natural T-regulatory cells (Tregs), an immune subset controlling tolerance and inflammation. Induction of autophagy required ligand-dependent, Notch intracellular domain (NIC) activity, which controlled mitochondrial organization and survival of activated Tregs. Consistently, NIC immune-precipitated Beclin and Atg14, constituents of the autophagy initiation complex. Further, ectopic expression of an effector of autophagy (Atg3) or recombinant NIC tagged to a nuclear export signal (NIC-NES), restored autophagy and suppressor function in Notch1(-/-) Tregs. Furthermore, Notch1 deficiency in the Treg lineage resulted in immune hyperactivity, implicating Notch activity in Treg homeostasis. Notch1 integration with autophagy, revealed in these experiments, holds implications for Notch regulated cell-fate decisions governing differentiation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.7554/eLife.14023DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4894756PMC
June 2016

Role of actin dependent nuclear deformation in regulating early gene expression.

PLoS One 2012 28;7(12):e53031. Epub 2012 Dec 28.

National Centre for Biological Sciences, Tata Institute of Fundamental Research, Bangalore, India.

The nucleus of a living cell is constantly undergoing changes in shape and size as a result of various mechanical forces in physiology. These changes correlate with alterations in gene expression, however it is unclear whether nuclear deformation alone is sufficient to elicit these alterations. We used T-cell activation as a model system to test the coupling between nuclear deformation (elongation) and gene expression. Naïve T-cell activation with surrogate antigens resulted in actin dependent nuclear elongation. This was accompanied with Erk and NF-κB signaling to the nucleus to induce CD69 expression. Importantly, inhibiting actin polymerization abolished both nuclear elongation and CD69 expression, while inhibiting Erk, NF-κB or microtubule depolymerization only abolished expression but not elongation. Immobilization of antigen-coated beads, under conditions where actin polymerization was inhibited, rescued both nuclear elongation and CD69 expression. In addition, fibroblast cells plated on fibronectin micropatterns of different sizes showed correlation between nuclear shape index and tenascin C expression. Upon inhibiting the signaling intermediate Erk, tenascin C expression was down regulated although the nuclear shape index remained unaltered. Our results highlight the importance of specific signaling intermediates accompanied with nuclear deformation in the modulation of cellular genomic programs.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0053031PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3532443PMC
July 2013

Apoptotic programs are determined during lineage commitment of CD4+ T effectors: selective regulation of T effector-memory apoptosis by inducible nitric oxide synthase.

J Immunol 2013 Jan 5;190(1):97-105. Epub 2012 Dec 5.

National Centre for Biological Sciences, Bangalore 560065, Karnataka, India.

Lineage-committed T effectors generated in response to Ag during the inflammatory phase are destined to die during termination of the immune response. We present evidence to suggest that molecular signatures of lineage commitment are reflected in apoptotic cascades activated in CD4(+) T effectors. Exemplifying this, ablation of inducible NO synthase (iNOS) protected effector-memory T (TEM) cells, but not T(Naive) or central-memory T cells, activated in vitro, from apoptosis triggered by cytokine deprivation. Furthermore, attrition of T effectors generated in the secondary, but not the primary, response to Ag was substantially reduced in mice, which received iNOS inhibitors. Distinct patterns of iNOS expression were revealed in wild-type TEM effectors undergoing apoptosis, and ablation of iNOS protein in primary and TEM wild-type effectors confirmed observations made in iNOS(-/-) cells. Describing molecular correlates of this dependence, mitochondrial damage, activation of the protein Bax, and release from mitochondria of the apoptosis-inducing factor were selectively abrogated in iNOS(-/-) TEM effectors. Suggesting that iNOS dependence was linked to the functional identity of T cell subsets, both iNOS induction and apoptosis were compromised in IFN-γ(-/-) TEM effectors, which mirrored the response patterns of iNOS(-)(/)(-) TEM. Collectively, these observations suggest that programs regulating deletion and differentiation are closely integrated and likely encoded during lineage commitment of T effectors.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.4049/jimmunol.1103694DOI Listing
January 2013

Developmental heterogeneity in DNA packaging patterns influences T-cell activation and transmigration.

PLoS One 2012 5;7(9):e43718. Epub 2012 Sep 5.

National Centre for Biological Sciences, Tata Institute for Fundamental Research, Bangalore, Karnataka, India.

Cellular differentiation programs are accompanied by large-scale changes in nuclear organization and gene expression. In this context, accompanying transitions in chromatin assembly that facilitates changes in gene expression and cell behavior in a developmental system are poorly understood. Here, we address this gap and map structural changes in chromatin organization during murine T-cell development, to describe an unusual heterogeneity in chromatin organization and associated functional correlates in T-cell lineage. Confocal imaging of DNA assembly in cells isolated from bone marrow, thymus and spleen reveal the emergence of heterogeneous patterns in DNA organization in mature T-cells following their exit from the thymus. The central DNA pattern dominated in immature precursor cells in the thymus whereas both central and peripheral DNA patterns were observed in naïve and memory cells in circulation. Naïve T-cells with central DNA patterns exhibited higher mechanical pliability in response to compressive loads in vitro and transmigration assays in vivo, and demonstrated accelerated expression of activation-induced marker CD69. T-cell activation was characterized by marked redistribution of DNA assembly to a central DNA pattern and increased nuclear size. Notably, heterogeneity in DNA patterns recovered in cells induced into quiescence in culture, suggesting an internal regulatory mechanism for chromatin reorganization. Taken together, our results uncover an important component of plasticity in nuclear organization, reflected in chromatin assembly, during T-cell development, differentiation and transmigration.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0043718PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3434176PMC
February 2013

Distinct spatial and molecular features of notch pathway assembly in regulatory T cells.

Sci Signal 2012 Jul 24;5(234):ra53. Epub 2012 Jul 24.

National Centre for Biological Sciences, Tata Institute of Fundamental Research, GKVK Campus, Bangalore 560065, India.

Variations in the spatial localization of signaling components and crosstalk among signaling cascades are mechanisms through which diversity in signaling networks is generated. The receptor Notch provides an example of regulation by spatial localization: In the canonical Notch signaling pathway, Notch is cleaved to produce the Notch intracellular domain (NICD, also known as NIC), which translocates to the nucleus to regulate gene expression. We describe a T cell receptor-dependent, non-nuclear distribution and function of the processed receptor Notch, which was associated with the improved survival of regulatory T cells (T(regs)) in vitro and in vivo and was compromised by T cell-specific deletion of Notch1. Unlike a nuclear-restricted mutant of NICD, mutant NICD that underwent nuclear export or was targeted to the plasma membrane protected Notch1(-/-) T(regs) from apoptosis induced by nutrient deprivation and oxidative stress. Notch signaling integrated with phosphatidylinositol 3-kinase signaling and mammalian target of rapamycin complex 2 (mTORC2) for this cell survival function. Biochemical and imaging approaches revealed a membrane-proximal complex containing NICD and the mTORC2 component Rictor, and this complex was stabilized by specific interactions with the Notch ligand Delta-like-1 and mediated the survival of T(regs). Together, our evidence for the spatial control of Notch and the crosstalk of Notch signaling with other pathways reveals coupling between the localization of Notch and diverse intracellular signaling pathways.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1126/scisignal.2002859DOI Listing
July 2012

Rv1218c, an ABC transporter of Mycobacterium tuberculosis with implications in drug discovery.

Antimicrob Agents Chemother 2010 Dec 4;54(12):5167-72. Epub 2010 Oct 4.

AstraZeneca India, AVISHKAR, Bellary Road, Hebbal, Bangalore 560024, India.

Efflux systems are important in determining the efficacy of antibiotics used in the treatment of bacterial infections. In the last decade much attention has been paid to studying the efflux pumps of mycobacteria. New classes of compounds are under investigation for development into potential candidate drugs for the treatment of tuberculosis. Quite often, these have poor bactericidal activities but exhibit excellent target (biochemical) inhibition. Microarray studies conducted in our laboratories for deciphering the mode of action of experimental drugs revealed the presence of putative ABC transporters. Among these transporters, Rv1218c was chosen for studying its physiological relevance in mediating efflux in Mycobacterium tuberculosis. A ΔRv1218c mutant of M. tuberculosis displayed a 4- to 8-fold increase in the inhibitory and bactericidal potency for different classes of compounds. The MICs and MBCs were reversed to wild-type values when the full-length Rv1218c gene was reintroduced into the ΔRv1218c mutant on a multicopy plasmid. Most of the compound classes had significantly better bactericidal activity in the ΔRv1218c mutant than in the wild-type H37Rv, suggesting the involvement of Rv1218c gene product in effluxing these compounds from M. tuberculosis. The implication of these findings on tuberculosis drug discovery is discussed.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/AAC.00610-10DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2981279PMC
December 2010

Inactivation of the ilvB1 gene in Mycobacterium tuberculosis leads to branched-chain amino acid auxotrophy and attenuation of virulence in mice.

Microbiology (Reading) 2009 Sep 18;155(Pt 9):2978-2987. Epub 2009 Jun 18.

AstraZeneca R&D, Bellary Road, Hebbal, Bangalore-560024, India.

Acetohydroxyacid synthase (AHAS) is the first enzyme in the branched-chain amino acid biosynthesis pathway in bacteria. Bioinformatics analysis revealed that the Mycobacterium tuberculosis genome contains four genes (ilvB1, ilvB2, ilvG and ilvX) coding for the large catalytic subunit of AHAS, whereas only one gene (ilvN or ilvH) coding for the smaller regulatory subunit of this enzyme was found. In order to understand the physiological role of AHAS in survival of the organism in vitro and in vivo, we inactivated the ilvB1 gene of M. tuberculosis. The mutant strain was found to be auxotrophic for all of the three branched-chain amino acids (isoleucine, leucine and valine), when grown with either C(6) or C(2) carbon sources, suggesting that the ilvB1 gene product is the major AHAS in M. tuberculosis. Depletion of these branched chain amino acids in the medium led to loss of viability of the DeltailvB1 strain in vitro, resulting in a 4-log reduction in colony-forming units after 10 days. Survival kinetics of the mutant strain cultured in macrophages maintained with sub-optimal concentrations of the branched-chain amino acids did not show any loss of viability, indicating either that the intracellular environment was rich in these amino acids or that the other AHAS catalytic subunits were functional under these conditions. Furthermore, the growth kinetics of the DeltailvB1 strain in mice indicated that although this mutant strain showed defective growth in vivo, it could persist in the infected mice for a long time, and therefore could be a potential vaccine candidate.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1099/mic.0.029884-0DOI Listing
September 2009

Evaluation of killing kinetics of anti-tuberculosis drugs on Mycobacterium tuberculosis using a bacteriophage-based assay.

Chemotherapy 2008 5;54(5):404-11. Epub 2008 Sep 5.

AstraZeneca India Private Limited, Bangalore, India.

Background: Killing kinetics studies on Mycobacterium tuberculosis are labour intensive and time consuming since it takes nearly 6-7 weeks to get the data from an experiment. A modified protocol is required to increase the throughput and expedite the results.

Methods: The killing kinetics of frontline drugs used for the treatment of tuberculosis was studied using 24-well plates and 2 methods of enumeration of survivors of M. tuberculosis following drug exposure, namely conventional plating (CFU) and a phage-based assay (plaque-forming units) using mycobacteriophage D29.

Results: The use of 24-well plates enabled in reducing the volume of the compound required for the studies and the phage-based enumeration speeded up the readout and compared well with the CFU-based enumeration.

Conclusion: These results were in agreement with the earlier findings reported with respect to rifampicin, isoniazid and moxifloxacin. Also, this study shows for the first time the concentration-dependent killing of streptomycin, the time-dependent killing of ethambutol and the profiling of an experimental anti-mycobacterial compound by these 2 methods.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1159/000153314DOI Listing
February 2009