Tyrosine Phosphorylation of the Chlamydial Effector Protein Tarp Is Species Specific and Not Required for Recruitment of Actin

Clifton, Dawn R.; Dooley, Cheryl A.; Grieshaber, Scott S.; Carabeo, Rey A.; Fields, Kenneth A.; Hackstadt, Ted

doi:10.1128/iai.73.7.3860-3868.2005

Cited by 102 publications

(159 citation statements)

References 42 publications

Supporting

Mentioning

152

Contrasting

Order By: Relevance

“…The TARP orthologs demonstrate striking sequence diversity among C. trachomatis strains and species of Chlamydophila (7,28). Interestingly, the tyrosine-phosphorylated tandem repeat domain of TARP is not conserved between species, yet both the actin-binding helix and proline-rich oligomerization domains are conserved among the sequenced strains, suggesting that these domains are integral to the function of TARP (7,28).…”

Section: Discussionmentioning

confidence: 96%

“…To define the residues within TARP responsible for actin polymerization, a series of truncated GST fusions (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11) with the TARP C domain were generated (see Fig. 8, which is published as supporting information on the PNAS web site).…”

Section: Tarp Binds To Actin But Not the Arp2͞3 Complex From Hela Extmentioning

confidence: 99%

“…TARP is present in all pathogenic Chlamydia species examined to date. Analysis of TARP orthologs from C. trachomatis, Chlamydia muridarium, Chlamydia caviae, and Chlamydia pneumoniae indicates that only C. trachomatis serovars are phosphorylated, despite all Chlamydia strains demonstrating the recruitment of actin to the site of entry (6,7).…”

mentioning

confidence: 99%

See 2 more Smart Citations

Chlamydial TARP is a bacterial nucleator of actin

Jewett¹,

Fischer

Mead³

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

166

223

View full text Add to dashboard Cite

Chlamydia trachomatis entry into host cells results from a parasitedirected remodeling of the actin cytoskeleton. A type III secreted effector, TARP (translocated actin recruiting phosphoprotein), has been implicated in the recruitment of actin to the site of internalization. To elucidate the role of TARP in actin recruitment, we identified host cell proteins that associated with recombinant GST-TARP fusions. TARP directly associated with actin, and this interaction promoted actin nucleation as determined by in vitro polymerization assays. Domain analysis of TARP identified an actin-binding domain that bears structural and primary amino acid sequence similarity to WH2 domain family proteins. In addition, a proline-rich domain was found to promote TARP oligomerization and was required for TARP-dependent nucleation of new actin filaments. Our findings reveal a mechanism by which chlamydiae induce localized cytoskeletal changes by the translocated effector TARP during entry into host cells.Chlamydia ͉ type III secretion ͉ cytoskeleton ͉ endocytosis A Gram-negative obligate intracellular bacterium, Chlamydia trachomatis, is the leading cause of preventable blindness worldwide and the most prevalent bacterial pathogen causing sexually transmitted disease in the western world (1). Chlamydiae initiate their intracellular developmental cycle by actively gaining entry into host cells. The extracellular infectious form of the developmental cycle is referred to as an elementary body (EB). Once engulfed by the host cell, the EB differentiates into the replicative reticulate body within the protective confines of a membrane-bound parasitophorous vacuole called an inclusion (2).The ligands on the EB surface and cognate host cell receptor have not been definitively identified; however, it appears that chlamydiae use an entry mechanism that involves several distinct levels of interaction (3, 4). EB invasion of nonphagocytic cells is thus the product of coordinated cytoskeletal remodeling characterized by the formation of pedestal-like structures and hypertrophic microvilli that are directly triggered by the invading chlamydiae (5).A recently identified C. trachomatis type III secretion system secreted protein called TARP (for translocated actin recruiting phosphoprotein) is tyrosine-phosphorylated by a host cell kinase and is spatially and temporally associated with the recruitment of actin at the site of EB invasion (6). TARP is present in all pathogenic Chlamydia species examined to date. Analysis of TARP orthologs from C. trachomatis, Chlamydia muridarium, Chlamydia caviae, and Chlamydia pneumoniae indicates that only C. trachomatis serovars are phosphorylated, despite all Chlamydia strains demonstrating the recruitment of actin to the site of entry (6, 7).We demonstrate here that TARP associates directly with actin by a small domain contained within the C-terminal region of the protein. Furthermore, TARP independently nucleates new actin filaments by forming a large homogenous multimeric protein complex mediated by a prolin...

show abstract

Section: Discussionmentioning

confidence: 96%

Section: Tarp Binds To Actin But Not the Arp2͞3 Complex From Hela Extmentioning

confidence: 99%

See 1 more Smart Citation

Chlamydial TARP is a bacterial nucleator of actin

Jewett¹,

Fischer

Mead³

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

166

223

View full text Add to dashboard Cite

show abstract

“…The biological significance of tyrosine phosphorylation by PknD of C. pneumoniae is currently unknown, but it is noteworthy that infection of host cells with Chlamydia results in tyrosine phosphorylation of host proteins (3,8,33) despite the absence of predicted tyrosine kinases in Chlamydia. Additionally, Chlamydia secretes the actin-recruiting protein Tarp into the host cytosol, and Tarp is believed to be phosphorylated by a host cell kinase (5,6). Tarp, PknD, and Cpn0712 are the only three chlamydial proteins that have been shown to be phosphorylated on tyrosine.…”

Section: Vol 189 2007 Topology and Dual Specificity Of C Pneumoniamentioning

confidence: 99%

Chlamydophila pneumoniae PknD Exhibits Dual Amino Acid Specificity and Phosphorylates Cpn0712, a Putative Type III Secretion YscD Homolog

Johnson

Mahony

2007

J Bacteriol

View full text Add to dashboard Cite

Chlamydophila pneumoniae is an obligate intracellular bacterium that causes bronchitis, pharyngitis, and pneumonia and may be involved in atherogenesis and Alzheimer's disease. Genome sequencing has identified three eukaryote-type serine/threonine protein kinases, Pkn1, Pkn5, and PknD, that may be important signaling molecules in Chlamydia. Full-length PknD was cloned and expressed as a histidine-tagged protein in Escherichia coli. Differential centrifugation followed by sodium carbonate treatment of E. coli membranes demonstrated that His-PknD is an integral membrane protein. Fusions of overlapping PknD fragments to alkaline phosphatase revealed that PknD contains a single transmembrane domain and that the kinase domain is in the cytoplasm. To facilitate solubility, the kinase domain was cloned and expressed as a glutathione Stransferase (GST) fusion protein in E. coli. Purified GST-PknD kinase domain autophosphorylated, and catalytic mutants (K33G, D156G, and K33G-D156G mutants) and activation loop mutants (T185A and T193A) were inactive. PknD phosphorylated recombinant Cpn0712, a type III secretion YscD homolog that has two forkhead-associated domains. Thin-layer chromatography revealed that the PknD kinase domain autophosphorylated on threonine and tyrosine and phosphorylated the FHA-2 domain of Cpn0712 on serine and tyrosine. To our knowledge, this is the first demonstration of a bacterial protein kinase with amino acid specificity for both serine/threonine and tyrosine residues and this is the first study to show phosphorylation of a predicted type III secretion structural protein.

show abstract

“…At least one early secreted effector, TARP, contributes to bacterial internalization by its ability to directly nucleate actin polymerization through a WH2 actin-binding domain mimic (Jewett et al 2006) and by recruiting the guanine nucleotide exchange factors (GEFs) Sos1 and Vav2, which activate Rac1 and signal to the actin machinery (Carabeo et al 2007;Lane et al 2008). This latter mechanism is only relevant in chlamydial species inwhich the polymorphic TARP gene (Clifton et al 2005) can be targeted for tyrosine phosphorylation by Abl, Src, and Syk kinases (Elwell et al 2008;Jewett et al 2008;Lane et al 2008;Mehlitz et al 2008Mehlitz et al , 2010. Microinjection of cells with antibodies directed to the actin-binding domain of TARP before infection significantly reduces bacterial invasion, providing the most conclusive support for a direct role of TARP in mediating bacterial entry (Jewett et al 2010).…”

Section: Mechanisms Of Chlamydia Invasion Of Epithelial Cellsmentioning

confidence: 99%

Chlamydial Intracellular Survival Strategies

Bastidas

Elwell

Engel

et al. 2013

Cold Spring Harbor Perspectives in Medicine

195

193

View full text Add to dashboard Cite

Chlamydia trachomatis is the most common sexually transmitted bacterial pathogen and the causative agent of blinding trachoma. Although Chlamydia is protected from humoral immune responses by residing within remodeled intracellular vacuoles, it still must contend with multilayered intracellular innate immune defenses deployed by its host while scavenging for nutrients. Here we provide an overview of Chlamydia biology and highlight recent findings detailing how this vacuole-bound pathogen manipulates host -cellular functions to invade host cells and maintain a replicative niche.

show abstract

Tyrosine Phosphorylation of the Chlamydial Effector Protein Tarp Is Species Specific and Not Required for Recruitment of Actin

Cited by 102 publications

References 42 publications

Chlamydial TARP is a bacterial nucleator of actin

Chlamydial TARP is a bacterial nucleator of actin

Chlamydophila pneumoniae PknD Exhibits Dual Amino Acid Specificity and Phosphorylates Cpn0712, a Putative Type III Secretion YscD Homolog

Chlamydial Intracellular Survival Strategies

Contact Info

Product

Resources

About