Structures of a human papillomavirus (HPV) E6 polypeptide bound to MAGUK proteins: mechanisms of targeting tumor suppressors by a high-risk HPV oncoprotein

Human papillomavirus (HPV) E6 oncoprotein targets certain tumor suppressors such as MAGI-1 and SAP97/hDlg for degradation. A short peptide at the C terminus of E6 interacts specifically with the PDZ domains of these tumor suppressors, which is a property unique to high-risk HPVs that are associated with cervical cancer. The detailed recognition mechanisms between HPV E6 and PDZ proteins are unclear. To understand the specific binding of cellular PDZ substrates by HPV E6, we have solved the crystal structures of the complexes containing a peptide from HPV18 E6 bound to three PDZ domains from MAGI-1 and SAP97/Dlg. The complex crystal structures reveal novel features of PDZ peptide recognition that explain why high-risk HPV E6 can specifically target these cellular tumor suppressors for destruction. Moreover, a new peptide-binding loop on these PDZs is identified as interacting with the E6 peptide. Furthermore, we have identified an arginine residue, unique to high-risk HPV E6 but outside the canonical core PDZ recognition motif, that plays an important role in the binding of the PDZs of both MAGI-I and SAP97/Dlg, the mutation of which abolishes E6's ability to degrade the two proteins. Finally, we have identified a dimer form of MAGI-1 PDZ domain 1 in the cocrystal structure with E6 peptide, which may have functional relevance for MAGI-1 activity. In addition to its novel insights into the biochemistry of PDZ interactions, this study is important for understanding HPV-induced oncogenesis; this could provide a basis for developing antiviral and anticancer compounds.     

Role of the PDZ domain-binding motif of the oncoprotein E6 in the pathogenesis of human papillomavirus type 31

A number of PDZ domain-containing proteins have been identified as binding partners for the oncoprotein E6 of the high-risk type human papillomaviruses (HPVs). These include hDlg, hScrib, MAGI-1, MAGI-2, MAGI-3, and MUPP1. The PDZ domain-binding motif (-X-T-X-V) at the carboxy terminus of E6 is essential for targeting PDZ proteins for proteasomal degradation. The presence of this motif only in the high-risk HPVs suggests its possible role in HPV-induced oncogenesis. To investigate the role of the PDZ domain-binding motif of E6 in the HPV life cycle, two mutant HPV31 genomes were constructed: E6ValDelta, with a deletion of the last amino acid residue of E6 (valine), and E6ETQVDelta, with a deletion of the entire PDZ domain-binding motif of E6 (ETQV). Three human foreskin keratinocyte (HFK) cell lines were established which maintained transfected wild-type HPV31 or either of two mutant genomes. Cells containing either of two mutant genomes were significantly retarded in their growth rates and reduced in their viral copy numbers compared to those transfected with wild-type genomes. Western analysis did not reveal any significant changes in the levels of PDZ proteins following stable transfection of any HPV31 genomes into HFKs. Although the E6ETQVDelta-transfected HFKs exhibited a pattern of morphological differentiation that appeared different from the HPV31 wild-type-transfected HFKs in organotypic raft cultures, immunohistochemical analysis failed to identify substantial changes in the differentiation-dependent membrane localization of hDlg proteins. These results suggest that binding of E6 to PDZ proteins modulates the early viral functions such as proliferation and maintenance of the viral copy number in undifferentiated cells.     

Association of cottontail rabbit papillomavirus E6 oncoproteins with the hDlg/SAP97 tumor suppressor

Papillomaviruses are small DNA viruses that infect epithelial tissues and cause warts. Human papillomavirus (HPV) infection is the primary risk factor for the development of cervical cancer. The E6 and E7 oncogenes are the only genes consistently expressed in HPV-positive cervical cancer cells. Cottontail rabbit papillomavirus (CRPV) induces papillomas and carcinomas on cottontail and domestic rabbits and provides an excellent animal model of HPV infection and vaccine development. CRPV encodes three transforming proteins; LE6, SE6, and E7. Each of these proteins is required for papilloma formation. Like HPV E7, the CRPV E7 protein binds to the tumor suppressor pRB. In contrast, unlike HPV E6, the CRPV E6 proteins do not bind the tumor suppressor p53. Although more than a dozen cellular proteins have been identified as HPV E6 interacting proteins, nothing is known about the cellular interacting proteins of CRPV E6s. Here we describe the association of CRPV E6s with hDlg/SAP97, the mammalian homolog of the Drosophila discs large tumor suppressor protein. HPV E6 has previously shown to bind and target hDlg/SAP97 for degradation. Our results demonstrate that both LE6 and SE6 interact with hDlg/SAP97, although their association does not lead to the degradation of hDlg/SAP97. The PDZ domains of hDlg were shown to be sufficient for interaction with CRPV E6 proteins while the C-terminus of CRPV E6 is essential for the interaction with hDlg. The association of hDlg with SE6 may be important but not sufficient for the transformation of NIH 3T3 cells by SE6. Importantly, a CRPV SE6 mutant defective for papilloma formation did not interact with hDlg. These results suggest that interaction with hDlg/SAP97 plays a role in the biological function of CRPV E6s.     

The Invasive Capacity of HPV Transformed Cells Requires the hDlg-Dependent Enhancement of SGEF/RhoG Activity

A major target of the HPV E6 oncoprotein is the human Discs Large (hDlg) tumour suppressor, although how this interaction contributes to HPV-induced malignancy is still unclear. Using a proteomic approach we show that a strong interacting partner of hDlg is the RhoG-specific guanine nucleotide exchange factor SGEF. The interaction between hDlg1 and SGEF involves both PDZ and SH3 domain recognition, and directly contributes to the regulation of SGEF''s cellular localization and activity. Consistent with this, hDlg is a strong enhancer of RhoG activity, which occurs in an SGEF-dependent manner. We also show that HPV-18 E6 can interact indirectly with SGEF in a manner that is dependent upon the presence of hDlg and PDZ binding capacity. In HPV transformed cells, E6 maintains a high level of RhoG activity, and this is dependent upon the presence of hDlg and SGEF, which are found in complex with E6. Furthermore, we show that E6, hDlg and SGEF each directly contributes to the invasive capacity of HPV-16 and HPV-18 transformed tumour cells. These studies demonstrate that hDlg has a distinct oncogenic function in the context of HPV induced malignancy, one of the outcomes of which is increased RhoG activity and increased invasive capacity.     

Structure of the Human Discs Large 1 PDZ2– Adenomatous Polyposis Coli Cytoskeletal Polarity Complex: Insight into Peptide Engagement and PDZ Clustering

The membrane associated guanylate kinase (MAGUK) family member, human Discs Large 1 (hDlg1) uses a PDZ domain array to interact with the polarity determinant, the Adenomatous Polyposis Coli (APC) microtubule plus end binding protein. The hDLG1-APC complex mediates a dynamic attachment between microtubule plus ends and polarized cortical determinants in epithelial cells, stem cells, and neuronal synapses. Using its multi-domain architecture, hDlg1 both scaffolds and regulates the polarity factors it engages. Molecular details underlying the hDlg1-APC interaction and insight into how the hDlg1 PDZ array may cluster and regulate its binding factors remain to be determined. Here, I present the crystal structure of the hDlg1 PDZ2-APC complex and the molecular determinants that mediate APC binding. The hDlg1 PDZ2-APC complex also provides insight into potential modes of ligand-dependent PDZ domain clustering that may parallel Dlg scaffold regulatory mechanisms. The hDlg1 PDZ2-APC complex presented here represents a core biological complex that bridges polarized cortical determinants with the dynamic microtubule cytoskeleton.     

Human scribble (Vartul) is targeted for ubiquitin-mediated degradation by the high-risk papillomavirus E6 proteins and the E6AP ubiquitin-protein ligase

The high-risk human papillomavirus (HPV) E6 proteins stimulate the ubiquitination and degradation of p53, dependent on the E6AP ubiquitin-protein ligase. Other proteins have also been shown to be targeted for degradation by E6, including hDlg, the human homolog of the Drosophila melanogaster Discs large (Dlg) tumor suppressor. We show here that the human homolog of the Drosophila Scribble (Vartul) (hScrib) tumor suppressor protein is also targeted for ubiquitination by the E6-E6AP complex in vitro and that expression of E6 induces degradation of hScrib in vivo. Characterization of the E6AP-E6-hScrib complex indicated that hScrib binds directly to E6 and that the binding is mediated by the PDZ domains of hScrib and a carboxyl-terminal epitope conserved among the high-risk HPV E6 proteins. Green fluorescent protein-hScrib was localized to the periphery of MDCK cells, where it colocalized with ZO-1, a component of tight junctions. E6 expression resulted in loss of integrity of tight junctions, as measured by ZO-1 localization, and this effect was dependent on the PDZ binding epitope of E6. Thus, the high-risk HPV E6 proteins induce the degradation of the human homologs of two Drosophila PDZ domain-containing tumor suppressor proteins, hDlg and hScrib, both of which are associated with cell junction complexes. The fact that Scrib/Vart and Dlg appear to cooperate in a pathway that controls Drosophila epithelial cell growth suggests that the combined targeting of hScrib and hDlg is an important component of the biologic activity of high-risk HPV E6 proteins.     

A systematic analysis of human papillomavirus (HPV) E6 PDZ substrates identifies MAGI-1 as a major target of HPV type 16 (HPV-16) and HPV-18 whose loss accompanies disruption of tight junctions

The E6 proteins from high-risk, cancer-causing types of human papillomavirus (HPV) are characterized by the presence of a PDZ (PSD95/Dlg/ZO-1) binding motif in their extreme carboxy termini, through which they interact with a number of cellular PDZ domain-containing substrates. In order to ascertain how many of these are degraded by E6 in vivo, we performed an extensive analysis of the effects of E6 ablation on the expression levels of a number of previously reported E6 PDZ substrates. Using HPV type 16 (HPV-16)-positive CaSKi cells and HPV-18-positive HeLa cells, we have found that MAGI-1 is a major degradation target of both HPV-16 and HPV-18 E6. In contrast, hDlg, hScrib, PTPN3, TIP2, FAP1, and PSD95 all exhibit various degrees of susceptibility to E6-induced degradation, and a high degree of HPV type specificity is observed for certain substrates. We also show that E6 preferentially targets MAGI-1 within the nucleus and at membrane sites. One of the direct consequences of MAGI-1 degradation is a loss of tight-junction integrity, as determined by mislocalization of the tight-junction protein ZO-1. Ablation of E6 expression restores tight junctions, and this restoration is dependent on the presence of MAGI-1. These results demonstrate that oncogenic HPV E6 proteins disrupt cellular tight junctions through the degradation of MAGI-1, and they provide further evidence of how the PDZ binding potential of E6 can contribute to HPV-induced malignancy.     

The Role of Protein Kinase A Regulation of the E6 PDZ-Binding Domain during the Differentiation-Dependent Life Cycle of Human Papillomavirus Type 18

Human papillomavirus (HPV) E6 proteins of high-risk alpha types target a select group of PSD95/DLG1/ZO1 (PDZ) domain-containing proteins by using a C-terminal PDZ-binding motif (PBM), an interaction that can be negatively regulated by phosphorylation of the E6 PBM by protein kinase A (PKA). Here, we have mutated the canonical PKA recognition motif that partially overlaps with the E6 PBM in the HPV18 genome (E6153PKA) and compared the effect of this mutation on the HPVl8 life cycle in primary keratinocytes with the wild-type genome and with a second mutant genome that lacks the E6 PBM (E6ΔPDZ). Loss of PKA recognition of E6 was associated with increased growth of the genome-containing cells relative to cells carrying the wild-type genome, and upon stratification, a more hyperplastic phenotype, with an increase in the number of S-phase competent cells in the upper suprabasal layers, while the opposite was seen with the E6ΔPDZ genome. Moreover, the growth of wild-type genome-containing cells was sensitive to changes in PKA activity, and these changes were associated with increased phosphorylation of the E6 PBM. In marked contrast to E6ΔPDZ genomes, the E6153PKA mutation exhibited no deleterious effects on viral genome amplification or expression of late proteins. Our data suggest that the E6 PBM function is differentially regulated by phosphorylation in the HPV18 life cycle. We speculate that perturbation of protein kinase signaling pathways could lead to changes in E6 PBM function, which in turn could have a bearing on tumor promotion and progression.     

The E6 oncoprotein from HPV16 enhances the canonical Wnt/β-catenin pathway in skin epidermis in vivo

The contribution of the Wnt signaling pathway to human papilloma virus (HPV)-induced carcinogenesis is poorly understood. In high-grade dysplastic lesions that are caused by high-risk HPVs (HR-HPV), β-catenin is often located in the cell nucleus, which suggests that Wnt pathway may be involved in the development of HPV-related carcinomas. Most of the oncogenic potential of HR-HPVs resides on the PDZ-binding domain of E6 protein. We hypothesized that the PDZ-binding domain of the HPV16-E6 oncoprotein induces the nuclear accumulation of β-catenin due to its capacity to degrade PDZ-containing cellular targets. To test this hypothesis, we evaluated the staining pattern of β-catenin in the skin epidermis of transgenic mice expressing the full-length E6 oncoprotein (K14E6 mice) and measured LacZ gene expression in K14E6 mice that were crossed with a strain expressing LacZ that was knocked into the Axin2 locus (Axin2(+/LacZ) mice). Here, we show that the E6 oncoprotein enhances the nuclear accumulation of β-catenin, the accumulation of cellular β-catenin-responsive genes, and the expression of LacZ. None of these effects were observed when a truncated E6 oncoprotein that lacks the PDZ-binding domain was expressed alone (K14E6ΔPDZ mice) or in combination with Axin2(+/LacZ). Conversely, cotransfection with either E6 or E6ΔPDZ similarly enhanced canonical Wnt signaling in short-term in vitro assays that used a luciferase Wnt/β-catenin/TCF-dependent promoter. We propose that the activation of canonical Wnt signaling could be induced by the HPV16-E6 oncoprotein; however, the participation of the E6 PDZ-binding domain seems to be important in in vivo models only.     

Two independent domains of hDlg are sufficient for subcellular targeting: the PDZ1-2 conformational unit and an alternatively spliced domain

hDlg, a human homologue of the Drosophila Dig tumor suppressor, contains two binding sites for protein 4.1, one within a domain containing three PSD-95/Dlg/ZO-1 (PDZ) repeats and another within the alternatively spliced I3 domain. Here, we further define the PDZ- protein 4.1 interaction in vitro and show the functional role of both 4.1 binding sites in situ. A single protease-resistant structure formed by the entirety of both PDZ repeats 1 and 2 (PDZ1-2) contains the protein 4.1-binding site. Both this PDZ1-2 site and the I3 domain associate with a 30-kD NH2-terminal domain of protein 4.1 that is conserved in ezrin/radixin/moesin (ERM) proteins. We show that both protein 4.1 and the ezrin ERM protein interact with the murine form of hDlg in a coprecipitating immune complex. In permeabilized cells and tissues, either the PDZ1-2 domain or the I3 domain alone are sufficient for proper subcellular targeting of exogenous hDlg. In situ, PDZ1-2- mediated targeting involves interactions with both 4.1/ERM proteins and proteins containing the COOH-terminal T/SXV motif. I3-mediated targeting depends exclusively on interactions with 4.1/ERM proteins. Our data elucidates the multivalent nature of membrane-associated guanylate kinase homologue (MAGUK) targeting, thus beginning to define those protein interactions that are critical in MAGUK function.     

NMR assignment of a PDZ domain in complex with a HPV51 E6 derived N-terminally pyroglutamic acid modified peptide

The resonance assignment of an amino-terminal pyroglutamic acid containing peptide derived from the E6 protein of human papillomavirus (HPV) type 51 in complex with PDZ domain 2 of hDlg/SAP-97 is reported. The assignments include ¹H, ¹³C and ¹⁵N resonances for the protein and peptide in the complex and all of the peptide’s pyroglutamic acid nuclei.     

Defining the minimal interacting regions of the tight junction protein MAGI-1 and HPV16 E6 oncoprotein for solution structure studies

The oncoprotein E6 produced by tumorigenic high-risk genital human papillomaviruses targets a number of cellular proteins containing PDZ domains for proteasome-mediated degradation. In particular, E6 targets the tight junction protein MAGI-1 by binding to its PDZ1 domain. Using light scattering and NMR, we explored different fragments of both the HPV16 E6 and the MAGI-1 PDZ1 domain to define the best-behaving complex for solution structure studies. We showed that the 70-residue HPV16 E6 C-terminal domain (E6C) can be efficiently substituted by a peptide spanning the 11 C-terminal residues of E6. The construct of MAGI-1 PDZ1 best suited for solution structure analysis presents a 14-residue N-terminal extension and a 26-residue C-terminal extension as compared to the construct used for the recently solved X-ray structure of a MAGI-1 PDZ1/HPV18 E6 complex. These data suggest a stabilizing role for the interdomain linker regions which separate the PDZ1 domain from its neighboring domains.     

E6 and E7 from human papillomavirus type 16 cooperate to target the PDZ protein Na/H exchange regulatory factor 1

Previous studies have shown that the PDZ-binding motif of the E6 oncoprotein from the mucosal high-risk (HR) human papillomavirus (HPV) types plays a key role in HPV-mediated cellular transformation in in vitro and in vivo experimental models. HR HPV E6 oncoproteins have the ability to efficiently degrade members of the PDZ motif-containing membrane-associated guanylate kinase (MAGUK) family; however, it is possible that other PDZ proteins are also targeted by E6. Here, we describe a novel interaction of HPV type 16 (HPV16) E6 with a PDZ protein, Na(+)/H(+) exchange regulatory factor 1 (NHERF-1), which is involved in a number of cellular processes, including signaling and transformation. HPV16 E6 associates with and promotes the degradation of NHERF-1, and this property is dependent on the C-terminal PDZ-binding motif of E6. Interestingly, HPV16 E7, via the activation of the cyclin-dependent kinase complexes, promoted the accumulation of a phosphorylated form of NHERF-1, which is preferentially targeted by E6. Thus, both oncoproteins appear to cooperate in targeting NHERF-1. Notably, HPV18 E6 is not able to induce NHERF-1 degradation, indicating that this property is not shared with E6 from all HR HPV types. Downregulation of NHERF-1 protein levels was also observed in HPV16-positive cervical cancer-derived cell lines, such as SiHa and CaSki, as well as HPV16-positive cervical intraepithelial neoplasia (CIN). Finally, our data show that HPV16-mediated NHERF-1 degradation correlates with the activation of the phosphatidylinositol-3'-OH kinase (PI3K)/AKT signaling pathway, which is known to play a key role in carcinogenesis.     

The PDZ Domain of the LIM Protein Enigma Binds to β-Tropomyosin

PDZ and LIM domains are modular protein interaction motifs present in proteins with diverse functions. Enigma is representative of a family of proteins composed of a series of conserved PDZ and LIM domains. The LIM domains of Enigma and its most related family member, Enigma homology protein, bind to protein kinases, whereas the PDZ domains of Enigma and family member actin-associated LIM protein bind to actin filaments. Enigma localizes to actin filaments in fibroblasts via its PDZ domain, and actin-associated LIM protein binds to and colocalizes with the actin-binding protein alpha-actinin-2 at Z lines in skeletal muscle. We show that Enigma is present at the Z line in skeletal muscle and that the PDZ domain of Enigma binds to a skeletal muscle target, the actin-binding protein tropomyosin (skeletal beta-TM). The interaction between Enigma and skeletal beta-TM was specific for the PDZ domain of Enigma, was abolished by mutations in the PDZ domain, and required the PDZ-binding consensus sequence (Thr-Ser-Leu) at the extreme carboxyl terminus of skeletal beta-TM. Enigma interacted with isoforms of tropomyosin expressed in C2C12 myotubes and formed an immunoprecipitable complex with skeletal beta-TM in transfected cells. The association of Enigma with skeletal beta-TM suggests a role for Enigma as an adapter protein that directs LIM-binding proteins to actin filaments of muscle cells.     

Human papillomaviruses and the specificity of PDZ domain targeting

The human papillomavirus (HPV) E6 oncoprotein is fundamental to the ability of these viruses to induce human malignancy. A defining characteristic of the HPV E6 oncoproteins found in cancer-causing HPV types is the presence of a PDZ binding motif at their extreme C-terminus. Through this motif, E6 is able to interact with a large number of cellular proteins that contain PDZ domains. Many of these cellular proteins are involved in regulation of processes associated with the control of cell attachment, cell proliferation, cell polarity and cell signaling. How E6 targets multiple proteins containing the same recognition domain is still an open question. In this review, we highlight aspects of E6 function and biology that help to answer this question, and thereby provide insight into the role of these substrates during development of HPV-induced malignancy.     

MAS C-Terminal Tail Interacting Proteins Identified by Mass Spectrometry- Based Proteomic Approach

Propagation of signals from G protein-coupled receptors (GPCRs) in cells is primarily mediated by protein-protein interactions. MAS is a GPCR that was initially discovered as an oncogene and is now known to play an important role in cardiovascular physiology. Current literature suggests that MAS interacts with common heterotrimeric G-proteins, but MAS interaction with proteins which might mediate G protein-independent or atypical signaling is unknown. In this study we hypothesized that MAS C-terminal tail (Ct) is a major determinant of receptor-scaffold protein interactions mediating MAS signaling. Mass-spectrometry based proteomic analysis was used to comprehensively identify the proteins that interact with MAS Ct comprising the PDZ-binding motif (PDZ-BM). We identified both PDZ and non-PDZ proteins from human embryonic kidney cell line, mouse atrial cardiomyocyte cell line and human heart tissue to interact specifically with MAS Ct. For the first time our study provides a panel of PDZ and other proteins that potentially interact with MAS with high significance. A ‘cardiac-specific finger print’ of MAS interacting PDZ proteins was identified which includes DLG1, MAGI1 and SNTA. Cell based experiments with wild-type and mutant MAS lacking the PDZ-BM validated MAS interaction with PDZ proteins DLG1 and TJP2. Bioinformatics analysis suggested well-known multi-protein scaffold complexes involved in nitric oxide signaling (NOS), cell-cell signaling of neuromuscular junctions, synapses and epithelial cells. Majority of these protein hits were predicted to be part of disease categories comprising cancers and malignant tumors. We propose a ‘MAS-signalosome’ model to stimulate further research in understanding the molecular mechanism of MAS function. Identifying hierarchy of interactions of ‘signalosome’ components with MAS will be a necessary step in future to fully understand the physiological and pathological functions of this enigmatic receptor.     

The Tiam1 PDZ Domain Couples to Syndecan1 and Promotes Cell-Matrix Adhesion

The T-cell lymphoma invasion and metastasis gene 1 (Tiam1) is a guanine exchange factor (GEF) for the Rho-family GTPase Rac1 that is crucial for the integrity of adherens junctions, tight junctions, and cell-matrix interactions. This GEF contains several protein-protein interaction domains, including a PDZ domain. Earlier studies identified a consensus PDZ-binding motif and a synthetic peptide capable of binding to the Tiam1 PDZ domain, but little is known about its ligand specificity and physiological role in cells. Here, we investigated the structure, specificity, and function of the Tiam1 PDZ domain. We determined the crystal structures of the Tiam1 PDZ domain free and in complex with a “model” peptide, which revealed the structural basis for ligand specificity. Protein database searches using the consensus PDZ-binding motif identified two eukaryotic cell adhesion proteins, Syndecan1 and Caspr4, as potential Tiam1 PDZ domain binding proteins. Equilibrium binding experiments confirmed that C-terminal peptides derived from Syndecan1 and Caspr4 bound the Tiam1 PDZ domain. NMR chemical shift perturbation experiments indicated that the Tiam1 PDZ/Syndecan1 and PDZ/Caspr4 complexes were structurally distinct and identified key residues likely to be responsible for ligand selectivity. Moreover, cell biological analysis established that Syndecan1 is a physiological binding partner of Tiam1 and that the PDZ domain has a function in cell-matrix adhesion and cell migration. Collectively, our data provide insight into the structure, specificity, and function of the Tiam1 PDZ domain. Importantly, our data report on a physiological role for the Tiam1 PDZ domain and establish a novel link between two previously unrelated signal transduction pathways, both of which are implicated in cancer.     

Systematic Analysis of Bacterial Effector-Postsynaptic Density 95/Disc Large/Zonula Occludens-1 (PDZ) Domain Interactions Demonstrates Shigella OspE Protein Promotes Protein Kinase C Activation via PDLIM Proteins

Diseases caused by many Gram-negative bacterial pathogens depend on the activities of bacterial effector proteins that are delivered into eukaryotic cells via specialized secretion systems. Effector protein function largely depends on specific subcellular targeting and specific interactions with cellular ligands. PDZ domains are common domains that serve to provide specificity in protein-protein interactions in eukaryotic systems. We show that putative PDZ-binding motifs are significantly enriched among effector proteins delivered into mammalian cells by certain bacterial pathogens. We use PDZ domain microarrays to identify candidate interaction partners of the Shigella flexneri effector proteins OspE1 and OspE2, which contain putative PDZ-binding motifs. We demonstrate in vitro and in cells that OspE proteins interact with PDLIM7, a member of the PDLIM family of proteins, which contain a PDZ domain and one or more LIM domains, protein interaction domains that participate in a wide variety of functions, including activation of isoforms of protein kinase C (PKC). We demonstrate that activation of PKC during S. flexneri infection is attenuated in the absence of PDLIM7 or OspE proteins and that the OspE PDZ-binding motif is required for wild-type levels of PKC activation. These results are consistent with a model in which binding of OspE to PDLIM7 during infection regulates the activity of PKC isoforms that bind to the PDLIM7 LIM domain.