Infection-driven activation of transglutaminase 2 boosts glucose uptake and hexosamine biosynthesis

Mapping Intimacies ◽

10.1101/634501 ◽

2019 ◽

Author(s):

Benoit Maffei ◽

Marc Laverrière ◽

Yongzheng Wu ◽

Sébastien Triboulet ◽

Stéphanie Perrinet ◽

...

Keyword(s):

Mammalian Cells ◽

Glucose Transporter ◽

Transglutaminase 2 ◽

Bacterial Development ◽

Obligate Intracellular ◽

Hexosamine Biosynthesis ◽

Infected Cells ◽

Obligate Intracellular Bacteria ◽

Dependent Pathway ◽

Transamidating Activity

ABSTRACTTransglutaminase 2 (TG2) is a ubiquitous enzyme with transamidating activity. We report here that the expression and activity of TG2 are enhanced in cells infected with the obligate intracellular bacteriaChlamydia trachomatis. Genetic or pharmacological inhibition of TG2 activity impair bacterial development. We show that TG2 increases glucose import by up-regulating the transcription of the glucose transporter genesGLUT-1andGLUT-3. Furthermore, TG2 activation drives one specific glucose-dependent pathway in the host, i.e. hexosamine biosynthesis. Mechanistically, we identify the glucosamine:fructose-6-phosphate amidotransferase (GFPT) among the substrates of TG2. GFPT modification by TG2 increases its enzymatic activity, resulting in higher levels of UDP-N-acetylglucosamine biosynthesis. As a consequence, TG2 activation results in increased proteinO-GlcNAcylation. The correlation between TG2 transamidating activity andO-GlcNAcylation is disrupted in infected cells because host hexosamine biosynthesis is being exploited by the bacteria, in particular to assist their division. In conclusion, our work establishes TG2 as a key player in controlling glucose-derived metabolic pathways in mammalian cells, themselves hijacked byC. trachomatisto sustain their own metabolic needs.

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Chlamydia preserves the mitochondrial network necessary for replication via microRNA-dependent inhibition of fission

The Journal of Cell Biology ◽

10.1083/jcb.201608063 ◽

2017 ◽

Vol 216 (4) ◽

pp. 1071-1089 ◽

Cited By ~ 43

Author(s):

Suvagata Roy Chowdhury ◽

Anastasija Reimer ◽

Malvika Sharan ◽

Vera Kozjak-Pavlovic ◽

Ana Eulalio ◽

...

Keyword(s):

Target Gene ◽

Mitochondrial Fission ◽

Host Cells ◽

Mitochondrial Network ◽

Mitochondrial Fragmentation ◽

Obligate Intracellular ◽

Mitochondrial Integrity ◽

Dependent Inhibition ◽

Infected Cells ◽

Obligate Intracellular Bacteria

Obligate intracellular bacteria such as Chlamydia trachomatis depend on metabolites of the host cell and thus protect their sole replication niche by interfering with the host cells’ stress response. Here, we investigated the involvement of host microRNAs (miRNAs) in maintaining the viability of C. trachomatis–infected primary human cells. We identified miR-30c-5p as a prominently up-regulated miRNA required for the stable down-regulation of p53, a major suppressor of metabolite supply in C. trachomatis–infected cells. Loss of miR-30c-5p led to the up-regulation of Drp1, a mitochondrial fission regulator and a target gene of p53, which, in turn, severely affected chlamydial growth and had a marked effect on the mitochondrial network. Drp1-induced mitochondrial fragmentation prevented replication of C. trachomatis even in p53-deficient cells. Additionally, Chlamydia maintain mitochondrial integrity during reactive oxygen species–induced stress that occurs naturally during infection. We show that C. trachomatis require mitochondrial ATP for normal development and hence postulate that they preserve mitochondrial integrity through a miR-30c-5p–dependent inhibition of Drp1-mediated mitochondrial fission.

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Apoptosis Functions in Defense against Infection of Mammalian Cells with Environmental Chlamydiae

Infection and Immunity ◽

10.1128/iai.00851-19 ◽

2020 ◽

Vol 88 (6) ◽

Author(s):

Dominik Brokatzky ◽

Oliver Kretz ◽

Georg Häcker

Keyword(s):

Epithelial Cells ◽

Mammalian Cells ◽

Intracellular Bacteria ◽

Mitochondrial Apoptosis ◽

Content Type ◽

Obligate Intracellular ◽

Human Epithelial Cells ◽

Antiapoptotic Activity ◽

Obligate Intracellular Bacteria ◽

Induced Apoptosis

ABSTRACT Apoptotic cell death can be an efficient defense reaction of mammalian cells infected with obligate intracellular pathogens; the host cell dies and the pathogen cannot replicate. While this is well established for viruses, there is little experimental support for such a concept in bacterial infections. All Chlamydiales are obligate intracellular bacteria, and different species infect vastly different hosts. Chlamydia trachomatis infects human epithelial cells; Parachlamydia acanthamoebae replicates in amoebae. We here report that apoptosis impedes growth of P. acanthamoebae in mammalian cells. In HeLa human epithelial cells, P. acanthamoebae infection induced apoptosis, which was inhibited when mitochondrial apoptosis was blocked by codeletion of the mediators of mitochondrial apoptosis, Bax and Bak, by overexpression of Bcl-XL or by deletion of the apoptosis initiator Noxa. Deletion of Bax and Bak in mouse macrophages also inhibited apoptosis. Blocking apoptosis permitted growth of P. acanthamoebae in HeLa cells, as measured by fluorescence in situ hybridization, assessment of genome replication and protein synthesis, and the generation of infectious progeny. Coinfection with C. trachomatis inhibited P. acanthamoebae-induced apoptosis, suggesting that the known antiapoptotic activity of C. trachomatis can also block P. acanthamoebae-induced apoptosis. C. trachomatis coinfection could not rescue P. acanthamoebae growth in HeLa; in coinfected cells, C. trachomatis even suppressed the growth of P. acanthamoebae independently of apoptosis, while P. acanthamoebae surprisingly enhanced the growth of C. trachomatis. Our results show that apoptosis can be used in the defense of mammalian cells against obligate intracellular bacteria and suggest that the known antiapoptotic activity of human pathogenic chlamydiae is indeed required to permit their growth in human cells.

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Development of a method for recovering rickettsial RNA from infected cells to analyze gene expression profiling of obligate intracellular bacteria

Journal of Microbiological Methods ◽

10.1016/j.mimet.2007.09.017 ◽

2007 ◽

Vol 71 (3) ◽

pp. 292-297 ◽

Cited By ~ 18

Author(s):

My-Van La ◽

Patrice François ◽

Clarisse Rovery ◽

Sylvianne Robineau ◽

Pascal Barbry ◽

...

Keyword(s):

Gene Expression ◽

Gene Expression Profiling ◽

Expression Profiling ◽

Intracellular Bacteria ◽

Obligate Intracellular ◽

Infected Cells ◽

Obligate Intracellular Bacteria

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Substitution of nucleotide-sugar by trehalose-dependent glycogen synthesis pathways in Chlamydiales underlines an unusual requirement for storage polysaccharides within obligate intracellular bacteria

10.1101/2020.06.02.131169 ◽

2020 ◽

Author(s):

Matthieu Colpaert ◽

Derifa Kadouche ◽

Mathieu Ducatez ◽

Trestan Pillonel ◽

Carole Kebbi-Beghdadi ◽

...

Keyword(s):

Biochemical Analysis ◽

Glycogen Synthesis ◽

Glycogen Metabolism ◽

Intracellular Bacteria ◽

Classical Pathway ◽

Nucleotide Sugar ◽

Obligate Intracellular ◽

Important Exception ◽

Obligate Intracellular Bacteria ◽

Dependent Pathway

AbstractAll obligate intracellular pathogens or symbionts of eukaryotes lack glycogen metabolism. Most members of the Chlamydiales order are exceptions to this rule as they contain the classical GlgA-GlgC-dependent pathway of glycogen metabolism that relies on the ADP-Glucose substrate. We surveyed the diversity of Chlamydiales and found glycogen metabolism to be universally present with the important exception of Criblamydiaceae and Waddliaceae families that had been previously reported to lack an active pathway. However, we now find elements of the more recently described GlgE maltose-1-P-dependent pathway in several protist-infecting Chlamydiales. In the case of Waddliaceae and Criblamydiaceae, the substitution of the classical pathway by this recently proposed GlgE pathway was essentially complete as evidenced by the loss of both GlgA and GlgC. Biochemical analysis of recombinant proteins expressed from Waddlia chondrophila and Estrella lausannensis established that both enzymes do polymerize glycogen from trehalose through the production of maltose-1-P by TreS-Mak and its incorporation into glycogen’s outer chains by GlgE. Unlike Mycobacteriaceae where GlgE-dependent polymerization is produced from both bacterial ADP-Glc and trehalose, glycogen synthesis seems to be entirely dependent on host supplied UDP-Glc and Glucose-6-P or on host supplied trehalose and maltooligosaccharides. These results are discussed in the light of a possible effector nature of these enzymes, of the chlamydial host specificity and of a possible function of glycogen in extracellular survival and infectivity of the chlamydial elementary bodies. They underline that contrarily to all other obligate intracellular bacteria, glycogen metabolism is indeed central to chlamydial replication and maintenance.

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Broad Degradation of Proapoptotic Proteins with the Conserved Bcl-2 Homology Domain 3 during Infection with Chlamydia trachomatis

Infection and Immunity ◽

10.1128/iai.73.3.1399-1403.2005 ◽

2005 ◽

Vol 73 (3) ◽

pp. 1399-1403 ◽

Cited By ~ 44

Author(s):

Songmin Ying ◽

Barbara M. Seiffert ◽

Georg Häcker ◽

Silke F. Fischer

Keyword(s):

Chlamydia Trachomatis ◽

Chlamydial Infection ◽

Death Receptor ◽

Proteolytic Degradation ◽

Intracellular Bacteria ◽

Obligate Intracellular ◽

Infected Cells ◽

Domain 3 ◽

Obligate Intracellular Bacteria ◽

Induced Apoptosis

ABSTRACT Chlamydiae are obligate intracellular bacteria that can inhibit apoptosis of their host cell. As shown recently, this inhibition is in part explained by the proteolytic degradation of the proapoptotic Bcl-2 family members (BH3-only proteins) Bim, Puma, and Bad upon chlamydial infection. In this study, we further explore this antiapoptotic mechanism. In cells infected with a Chlamydia trachomatis L2 strain, Bim, Puma, and Bad were degraded with similar kinetics, and the degradation of all three was blocked by inhibition of the proteasome. Furthermore, the BH3-only proteins Bmf, Noxa, and tBid were also targeted by chlamydial infection. The constitutively expressed Bmf disappeared during infection. When Noxa was experimentally induced, the levels were also reduced by infection with C. trachomatis. In death-receptor-induced apoptosis, cleaved and activated tBid was degraded, and this destruction was also prevented by inhibition of the proteasome. These results show that chlamydial infection leads to a broad degradation of BH3-only proteins. This loss of proapoptotic factors can explain the almost general protection of infected cells against apoptotic stimuli.

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The growing repertoire of genetic tools for dissecting chlamydial pathogenesis

Pathogens and Disease ◽

10.1093/femspd/ftab025 ◽

2021 ◽

Author(s):

Arkaprabha Banerjee ◽

David E Nelson

Keyword(s):

Genetic Engineering ◽

Intracellular Bacteria ◽

Human Pathogens ◽

Pathogenic Potential ◽

Host Preferences ◽

Genetic Tools ◽

Obligate Intracellular ◽

Obligate Intracellular Bacteria ◽

Multiple Species

Abstract Multiple species of obligate intracellular bacteria in the genus Chlamydia are important veterinary and/or human pathogens. These pathogens all share similar biphasic developmental cycles and transition between intracellular vegetative reticulate bodies and infectious elementary forms, but vary substantially in their host preferences and pathogenic potential. A lack of tools for genetic engineering of these organisms has long been an impediment to the study of their biology and pathogenesis. However, the refinement of approaches developed in C. trachomatis over the last ten years, and adaptation of some of these approaches to other Chlamydia spp. in just the last few years, has opened exciting new possibilities for studying this ubiquitous group of important pathogens.

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Dicer is involved in protection against influenza A virus infection

Journal of General Virology ◽

10.1099/vir.0.83103-0 ◽

2007 ◽

Vol 88 (10) ◽

pp. 2627-2635 ◽

Cited By ~ 80

Author(s):

Alexey A. Matskevich ◽

Karin Moelling

Keyword(s):

Virus Infection ◽

Influenza A Virus ◽

Mammalian Cells ◽

Influenza A ◽

Virus Production ◽

Vero Cells ◽

Antiviral Response ◽

Protein Levels ◽

Infected Cells ◽

Influenza A Virus Infection

In mammals the interferon (IFN) system is a central innate antiviral defence mechanism, while the involvement of RNA interference (RNAi) in antiviral response against RNA viruses is uncertain. Here, we tested whether RNAi is involved in the antiviral response in mammalian cells. To investigate the role of RNAi in influenza A virus-infected cells in the absence of IFN, we used Vero cells that lack IFN-α and IFN-β genes. Our results demonstrate that knockdown of a key RNAi component, Dicer, led to a modest increase of virus production and accelerated apoptosis of influenza A virus-infected cells. These effects were much weaker in the presence of IFN. The results also show that in both Vero cells and the IFN-producing alveolar epithelial A549 cell line influenza A virus targets Dicer at mRNA and protein levels. Thus, RNAi is involved in antiviral response, and Dicer is important for protection against influenza A virus infection.

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CXCR2 Blockade Influences Anaplasma phagocytophilum Propagation but Not Histopathology in the Mouse Model of Human Granulocytic Anaplasmosis

Clinical and Diagnostic Laboratory Immunology ◽

10.1128/cdli.11.5.963-968.2004 ◽

2004 ◽

Vol 11 (5) ◽

pp. 963-968 ◽

Cited By ~ 34

Author(s):

Diana G. Scorpio ◽

Mustafa Akkoyunlu ◽

Erol Fikrig ◽

J. Stephen Dumler

Keyword(s):

Anaplasma Phagocytophilum ◽

Tissue Injury ◽

Treated Group ◽

Obligate Intracellular Bacterium ◽

Human Granulocytic Anaplasmosis ◽

Obligate Intracellular ◽

Liver Histopathology ◽

Granulocytic Anaplasmosis ◽

Infected Cells ◽

And Control

ABSTRACT Anaplasma phagocytophilum is an obligate intracellular bacterium that infects neutrophils and causes human granulocytic anaplasmosis. Infection induces neutrophil secretion of interleukin-8 or murine homologs and perpetuates infection by recruiting susceptible neutrophils. We hypothesized that antibody blockade of CXCR2 would decrease A. phagocytophilum tissue load by interrupting neutrophil recruitment but would not influence murine hepatic pathology. C3H-scid mice were treated with CXCR2 antiserum or control prior to or on day 14 after infection. Quantitative PCR and immunohistochemistry for A. phagocytophilum were performed and severity of liver histopathology was ranked. Control mice had more infected cells in tissues than the anti-CXCR2-treated group. The histopathological rank was not different between treated and control animals. Infected cells of control mice clustered in tissue more than in treated mice. The results support the hypothesis of bacterial propagation through chemokine induction and confirm that tissue injury is unrelated to A. phagocytophilum tissue load.

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Initial Characterization of the Two ClpP Paralogs ofChlamydia trachomatisSuggests Unique Functionality for Each

Journal of Bacteriology ◽

10.1128/jb.00635-18 ◽

2018 ◽

Vol 201 (2) ◽

Cited By ~ 5

Author(s):

Nicholas A. Wood ◽

Krystal Y. Chung ◽

Amanda M. Blocker ◽

Nathalia Rodrigues de Almeida ◽

Martin Conda-Sheridan ◽

...

Keyword(s):

Host Cells ◽

Developmental Cycle ◽

Intracellular Bacteria ◽

Clp Protease ◽

Content Type ◽

Sexually Transmitted ◽

Obligate Intracellular ◽

Developmental Form ◽

Obligate Intracellular Bacteria

ABSTRACTMembers ofChlamydiaare obligate intracellular bacteria that differentiate between two distinct functional and morphological forms during their developmental cycle, elementary bodies (EBs) and reticulate bodies (RBs). EBs are nondividing small electron-dense forms that infect host cells. RBs are larger noninfectious replicative forms that develop within a membrane-bound vesicle, termed an inclusion. Given the unique properties of each developmental form of this bacterium, we hypothesized that the Clp protease system plays an integral role in proteomic turnover by degrading specific proteins from one developmental form or the other.Chlamydiaspp. have five uncharacterizedclpgenes,clpX,clpC, twoclpPparalogs, andclpB. In other bacteria, ClpC and ClpX are ATPases that unfold and feed proteins into the ClpP protease to be degraded, and ClpB is a deaggregase. Here, we focused on characterizing the ClpP paralogs. Transcriptional analyses and immunoblotting determined that these genes are expressed midcycle. Bioinformatic analyses of these proteins identified key residues important for activity. Overexpression of inactiveclpPmutants inChlamydiaspp. suggested independent function of each ClpP paralog. To further probe these differences, we determined interactions between the ClpP proteins using bacterial two-hybrid assays and native gel analysis of recombinant proteins. Homotypic interactions of the ClpP proteins, but not heterotypic interactions between the ClpP paralogs, were detected. Interestingly, protease activity of ClpP2, but not ClpP1, was detectedin vitro. This activity was stimulated by antibiotics known to activate ClpP, which also blocked chlamydial growth. Our data suggest the chlamydial ClpP paralogs likely serve distinct and critical roles in this important pathogen.IMPORTANCEChlamydia trachomatisis the leading cause of preventable infectious blindness and of bacterial sexually transmitted infections worldwide. Chlamydiae are developmentally regulated obligate intracellular pathogens that alternate between two functional and morphologic forms, with distinct repertoires of proteins. We hypothesize that protein degradation is a critical aspect to the developmental cycle. A key system involved in protein turnover in bacteria is the Clp protease system. Here, we characterized the two chlamydial ClpP paralogs by examining their expression inChlamydiaspp., their ability to oligomerize, and their proteolytic activity. This work will help understand the evolutionarily diverse Clp proteases in the context of intracellular organisms, which may aid in the study of other clinically relevant intracellular bacteria.

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