scholarly journals Intramolecular Dimerization Is Required for the Chlamydia-Secreted Protease CPAF To Degrade Host Transcriptional Factors

2004 ◽  
Vol 72 (7) ◽  
pp. 3869-3875 ◽  
Author(s):  
Feng Dong ◽  
Jyotika Sharma ◽  
Yanming Xiao ◽  
Youmin Zhong ◽  
Guangming Zhong
Keyword(s):  
Transcription Factors ◽  
Host Cell ◽  
Proteolytic Activity ◽  
Bacterial Species ◽  
Antigen Expression ◽  
Obligate Intracellular ◽  
Histocompatibility Complex ◽  
Activity Factor ◽  
Cell Cytosol ◽  
Host Cell Cytosol

ABSTRACT We previously identified a chlamydial protein designated CPAF (chlamydia protease/proteasome-like activity factor) that is secreted into host cell cytosol for degrading host transcription factors required for major histocompatibility complex antigen expression. Here we report that CPAF, synthesized as a 70-kDa proprotein, is processed into two fragments (designated CPAFn and CPAFc) to form intramolecular dimers that are much more stable than the naïve CPAF. Precipitation with antibodies that recognized CPAF dimers removed the proteolytic activity responsible for degrading host transcription factor RFX5 from chlamydia-infected host cell cytosol, while precipitation with antibodies that recognized free CPAF fragments alone did not remove this activity. Separation of CPAFn from CPAFc resulted in a loss of proteolytic activity. Furthermore, neither expressed full-length CPAF that was not processed nor coexpressed CPAFn and CPAFc fragments that failed to form dimers degraded RFX5. These observations demonstrate that intramolecular dimerization is required for CPAF to degrade host transcription factors, a strategy that is utilized by an obligate intracellular bacterial species to evade host defenses.

scholarly journals Chlamydia pneumoniae Secretion of a Protease-Like Activity Factor for Degrading Host Cell Transcription Factors Is Required for Major Histocompatibility Complex Antigen Expression

2002 ◽  
Vol 70 (1) ◽  
pp. 345-349 ◽  
Author(s):  
Peiyi Fan ◽  
Feng Dong ◽  
Yanqing Huang ◽  
Guangming Zhong
Keyword(s):  
Transcription Factors ◽  
Major Histocompatibility Complex ◽  
Chlamydia Pneumoniae ◽  
Respiratory Infections ◽  
Antigen Expression ◽  
Major Histocompatibility ◽  
Histocompatibility Complex ◽  
Activity Factor ◽  
Infected Cells ◽  
Mhc Antigen

ABSTRACT Chlamydia pneumoniae is a causative agent for many respiratory infections and has been associated with cardiovascular diseases in humans. The pathogenicity of C. pneumoniae is thought to depend on its ability to cause persistent infection and to evade host defense. Genome sequence analysis indicates that C. pneumoniae encodes a homologue of a chlamydial protease-like activity factor from C. trachomatis (CPAFct). We designated the C. pneumoniae homologue as CPAFcp. Recombinant CPAFcp was produced and found to degrade RFX5, a host transcription factor required for major histocompatibility complex (MHC) antigen expression. The degradation was inhibitable by lactacystin, an irreversible proteasome inhibitor. Furthermore, CPAFcp was secreted into host cytosol by C. pneumoniae organisms. Depletion of the C. pneumoniae-secreted CPAFcp with specific antibodies completely ablated the RFX5 degradation activity in the infected cells, suggesting that CPAFcp is necessary for the degradation of host transcription factors required for MHC antigen expression during C. pneumoniae infection. These observations have revealed a unique molecular mechanism for C. pneumoniae to evade host adaptive immunity that may aid in its persistence.

scholarly journals Identification of a Chlamydial Protease–Like Activity Factor Responsible for the Degradation of Host Transcription Factors

2001 ◽  
Vol 193 (8) ◽  
pp. 935-942 ◽  
Author(s):  
Guangming Zhong ◽  
Peiyi Fan ◽  
Hezhao Ji ◽  
Feng Dong ◽  
Yanqing Huang
Keyword(s):  
Transcription Factors ◽  
Bacterial Pathogen ◽  
Antigen Expression ◽  
Microbial Pathogens ◽  
Host Responses ◽  
Immune Recognition ◽  
Obligate Intracellular ◽  
Activity Factor ◽  
Obligate Intracellular Pathogen ◽  
Necessary And Sufficient

Microbial pathogens have been selected for the capacity to evade or manipulate host responses in order to survive after infection. Chlamydia, an obligate intracellular pathogen and the causative agent for many human diseases, can escape T lymphocyte immune recognition by degrading host transcription factors required for major histocompatibility complex (MHC) antigen expression. We have now identified a chlamydial protease– or proteasome–like activity factor (CPAF) that is secreted into the host cell cytosol and that is both necessary and sufficient for the degradation of host transcription factors RFX5 and upstream stimulation factor 1 (USF-1). The CPAF gene is highly conserved among chlamydial strains, but has no significant overall homology with other known genes. Thus, CPAF represents a unique secreted protein produced by an obligate intracellular bacterial pathogen to interfere with effective host adaptive immunity.

scholarly journals The Chlamydial Plasmid-Encoded Protein pgp3 Is Secreted into the Cytosol of Chlamydia-Infected Cells

2008 ◽  
Vol 76 (8) ◽  
pp. 3415-3428 ◽  
Author(s):  
Zhongyu Li ◽  
Ding Chen ◽  
Youmin Zhong ◽  
Shiping Wang ◽  
Guangming Zhong
Keyword(s):  
Host Cell ◽  
Chlamydial Infection ◽  
Open Reading Frames ◽  
Cryptic Plasmid ◽  
Activity Factor ◽  
Infected Cells ◽  
Cell Cytosol ◽  
Endogenous Proteins ◽  
Chlamydial Genome ◽  
Host Cell Cytosol

ABSTRACT The chlamydial cryptic plasmid encodes eight putative open reading frames (ORFs), designated pORF1 to -8. Antibodies raised against these ORF proteins were used to localize the endogenous proteins during chlamydial infection. We found that the pORF5 protein (also known as pgp3) was detected mainly in the cytosol of Chlamydia-infected cells, while the remaining seven proteins were found inside the chlamydial inclusions only. The pgp3 distribution pattern in the host cell cytosol is similar to but not overlapping with that of chlamydial protease/proteasome-like activity factor (CPAF), a chlamydial genome-encoded protein known to be secreted from chlamydial inclusions into the host cell cytosol. The anti-pgp3 labeling was removed by preabsorption with pgp3 but not CPAF fusion proteins and vice versa, demonstrating that pgp3 is a unique secretion protein. This conclusion is further supported by the observation that pgp3 was highly enriched in cytosolic fractions and had a minimal presence in the inclusion-containing nuclear fractions prepared from Chlamydia-infected cells. The pgp3 protein was detected as early as 12 h after infection and was secreted by all chlamydial species that carry the cryptic plasmid, suggesting that there is a selection pressure for maintaining pgp3 secretion during chlamydial infection. Although expression of pgp3 in the host cell cytosol via a transgene did not alter the susceptibility of the transfected cells to the subsequent chlamydial infection, purified pgp3 protein stimulated macrophages to release inflammatory cytokines, suggesting that pgp3 may contribute to chlamydial pathogenesis.

Parasitophorous vacuoles of Leishmania mexicana acquire macromolecules from the host cell cytosol via two independent routes

1999 ◽  
Vol 112 (5) ◽  
pp. 681-693
Author(s):  
U.E. Schaible ◽  
P.H. Schlesinger ◽  
T.H. Steinberg ◽  
W.F. Mangel ◽  
T. Kobayashi ◽  
...  
Keyword(s):  
Host Cell ◽  
Lucifer Yellow ◽  
Cysteine Proteinase ◽  
Organic Anion ◽  
Organic Anion Transporter ◽  
Anion Transporter ◽  
Infected Cells ◽  
Lysobisphosphatidic Acid ◽  
Cell Cytosol ◽  
Host Cell Cytosol

The intracellular parasite Leishmania survives and proliferates in host macrophages. In this study we show that parasitophorous vacuoles of L. mexicana gain access to cytosolic material via two different routes. (1) Small anionic molecules such as Lucifer Yellow are rapidly transported into the vacuoles by an active transport mechanism that is sensitive to inhibitors of the host cell's organic anion transporter. (2) Larger molecules such as fluorescent dextrans introduced into the host cell cytosol are also delivered to parasitophorous vacuoles. This transport is slower and sensitive to modulators of autophagy. Infected macrophages were examined by two novel assays to visualize and quantify this process. Immunoelectron microscopy of cells loaded with digoxigenin-dextran revealed label in multivesicular endosomes, which appeared to fuse with parasitophorous vacuoles. The inner membranes of the multivesicular vesicles label strongly with antibodies against lysobisphosphatidic acid, suggesting that they represent a point of confluence between the endosomal and autophagosomal pathways. Although the rate of autophagous transfer was comparable in infected and uninfected cells, infected cells retained hydrolyzed cysteine proteinase substrate to a greater degree. These data suggest that L. mexicana-containing vacuoles have access to potential nutrients in the host cell cytosol via at least two independent mechanisms.

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