Ribosomal protein S3: A multifunctional target of attaching/effacing bacterial pathogens

MINI REVIEW ARTICLE published: 27 June 2011 doi: 10.3389/fmicb.2011.00137 Ribosomal protein S3: a multifunctional target of attaching/effacing bacterial pathogens Xiaofei Gao and Philip R. Hardwidge* Department of Microbiology, Molecular Genetics and Immunology, University of Kansas Medical Center, Kansas City, KS, USA Edited by: Elizabeth L. Hartland, University of Melbourne, Australia Reviewed by: Elizabeth L. Hartland, University of Melbourne, Australia Toru Tobe, Osaka University, Japan *Correspondence: Philip R. Hardwidge, Department of Microbiology, Molecular Genetics and Immunology, University of Kansas Medical Center, 3901 Rainbow Boulevard, Kansas City, KS 66160, USA. e-mail: The extraribosomal functions of ribosomal proteins have drawn significant recent attention. Ribosomal protein S3 (RPS3), a component of the eukaryotic 40S ribosomal subunit, is a multifunctional protein that regulates DNA repair, apoptosis, and the innate immune response to bacterial infection. Here we the review the latest findings about RPS3 extraribosomal functions, with special emphasis on their relation to microbial pathogenesis and enteropathogenic Escherichia coli. Keywords: apoptosis, DNA repair, EPEC, extraribosomal function, NF-κB, NleH1, ribosomal protein S3 INTRODUCTION Ribosomal proteins function not only in protein translation, but also in multiple extraribosomal activities (Blumenthal and Carmichael, 1979). These functions include, but are not limited to, DNA repair, cell death, inflammation, tumorigenesis, and transcriptional regulation (Warner and McIntosh, 2009). Here we focus on a eukaryotic 40S ribosome component, the ribosomal protein S3 (RPS3), and its emerging regulatory roles in DNA repair, apoptosis, and pro-inflammatory signaling during bacterial infection. We propose that RPS3 may play a central role in regulating numerous aspects of host–pathogen interactions. RPS3 AND MICROBIAL PATHOGENESIS Ribosomal protein S3 has been directly and indirectly implicated in host–pathogen interactions. A clone of human RPS3 was obtained in a yeast three-hybrid screen designed to identify proteins that bind the 3 untranslated region (UTR) of hepatitis C virus (Wood et al., 2001). Suppression subtractive hybridization studies of mast cell gene expression modulated by Pseudomonas aeruginosa suggested that RPS3 might be involved in P. aeruginosa pathogenesis (Sun et al., 2005). RPS3 expression levels may also be important to mouse resistance to the H5N1 influenza virus (Boon et al., 2009). The NF-κB family of transcription factors regulates the expression of genes involved in a variety of cellular functions such as immune responses and cellular proliferation (Lenardo and Baltimore, 1989). NF-κB is normally sequestered in the cytoplasm by inhibitory IκB proteins that mask NF-κB nuclear localization signals (Hacker and Karin, 2006). After a cell recognizes a pathogen-associated molecular pattern (PAMP), the IκB kinase (IKK) complex is activated and subsequently phosphorylates the IκBs, leading to their ubiquitination and degradation by the 26S proteasome, permitting NF-κB subunits to translocate into the nucleus to function in transcription. It was recently discovered that RPS3 is also inducibly associated with and phosphorylated by IKKβ on serine 209 (S209) in response www.frontiersin.org to NF-κB pathway activation (Wan et al., 2011). This phosphorylation event is essential to the nuclear translocation of RPS3, after it associates with importin-α (Wan et al., 2011). Affinity purification experiments had also revealed that RPS3 interacts with the p65 NF-κB subunit through its K homology (KH) domain (Wan et al., 2007; Figure 1). After entering the nucleus, NF-κB binds to κB sites within target gene promoters and regulate transcription by recruiting co-activators/repressors (Wan et al., 2007). This newly discovered NF-κB subunit, RPS3, guides NF-κB to specific κB sites by increasing the affinity of the p65 NF-κB subunit for a subset of target gene promoters (Wan et al., 2007). Likewise, silencing RPS3 expression alters a subset of NF-κB signal transduction pathways. RPS3 thus provides for selective NF-κB recruitment to specific promoters and tailors cellular transcriptional responses to specific stimuli. Interestingly, RPS3 also forms a complex with NF-κB in human islet cells after stimulation with IL-1β (Mokhtari et al., 2009). The function of type III secretion system (T3SS) effector proteins has been a subject of intense research in recent years (Dean and Kenny, 2009). Some effectors (e.g., NleB, NleC, NleD, NleE, NleH) are key modulators of the innate immune system of intestinal epithelial cells, especially pathways regulated by NF-κB. For example, NleC is a protease that cleaves the NF-κB p65 subunit (Marches et al., 2005; Yen et al., 2010; Baruch et al., 2011; Muhlen et al., 2011; Pearson et al., 2011). NleD cleaves the c-Jun N-terminal kinase (JNK) thus blocking activator protein-1 (AP-1) activation (Baruch et al., 2011). NleE inhibits both p65 nuclear translocation and IκBα degradation (Newton et al., 2010) to block NF-κB activation, in conjunction with NleB (Nadler et al., 2010; Newton et al., 2010). During attaching/effacing (A/E) pathogen infection, the T3SS effectors NleH1 and NleH2 bind to the N-terminus of RPS3 after their translocation into host cells (Gao et al., 2009). NleH1, but not NleH2, inhibits the nuclear translocation of RPS3, consequently inhibiting the transcription of genes encoding pro-inflammatory June 2011 | Volume 2 | Article 137 | 1 Gao and Hardwidge Emerging functions of RPS3 Rb E2F1 PRMT1 KKRK NLS? importin-α RPS3 NleH1 NleH2 p65 ERK FIGURE 1 | Known and postulated interactions between RPS3 and mammalian or bacterial proteins. Specific RPS3 phosphorylation sites and the protein kinases responsible (PKCδ, ERK, Akt) are indicated. NleH1 and NleH2 are E. coli virulence proteins. NleH1 cytokines, such as IL-8 and TNF-α, indicating that pathogens target RPS3 to inhibit host immune defenses (Gao et al., 2009). NleH1 functions by inhibiting the IKKβ-mediated phosphorylation of RPS3 S209 (Wan et al., 2011). NleH1 is an autophosphorylated Ser/Thr protein kinase with an active site at lysine 159 (K159; Gao et al., 2009). While the kinase substrate for NleH1 is not yet known, it does not appear to phosphorylate either IKKβ or RPS3. However, NleH1 kinase activity is required to inhibit IKKβ from phosphorylating RPS3, as mutating the NleH1 K159 residue to alanine (K159A) prevented NleH1 from inhibiting RPS3 S209 phosphorylation, both in vitro and in cell culture models (Wan et al., 2011). Studies of gnotobiotic piglets infected with Escherichia coli O157:H7 also demonstrated that RPS3 S209 phosphorylation is inhibited by NleH1 in vivo, possibly to benefit bacterial colonization and transmission (Wan et al., 2011). It is interesting that IKKβ activation and IκBα degradation appear to be unaffected by NleH1 (Wan et al., 2011) suggesting that it may be benefic (...truncated)