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The TAL effector PthA4 interacts with nuclear factors involved in RNA-dependent processes including a HMG protein that selectively binds poly(U) RNA.

de Souza TA, Soprano AS, de Lira NP, Quaresma AJ, Pauletti BA, Paes Leme AF, Benedetti CE - PLoS ONE (2012)

Bottom Line: Plant pathogenic bacteria utilize an array of effector proteins to cause disease.Surprisingly, both CsHMG and PthA4 interacted with PABP1 and 2 and showed selective binding to poly(U) RNA, a property that is novel among HMGs and TAL effectors.Given that homologs of CsHMG, CsPABP1, CsPABP2, CsSMC and CsTRAX in other organisms assemble into protein complexes to regulate mRNA stability and translation, we suggest a novel role of TAL effectors in mRNA processing and translational control.

View Article: PubMed Central - PubMed

Affiliation: Laboratório Nacional de Biociências, Centro Nacional de Pesquisa em Energia e Materiais, Campinas, SP, Brazil.

ABSTRACT
Plant pathogenic bacteria utilize an array of effector proteins to cause disease. Among them, transcriptional activator-like (TAL) effectors are unusual in the sense that they modulate transcription in the host. Although target genes and DNA specificity of TAL effectors have been elucidated, how TAL proteins control host transcription is poorly understood. Previously, we showed that the Xanthomonas citri TAL effectors, PthAs 2 and 3, preferentially targeted a citrus protein complex associated with transcription control and DNA repair. To extend our knowledge on the mode of action of PthAs, we have identified new protein targets of the PthA4 variant, required to elicit canker on citrus. Here we show that all the PthA4-interacting proteins are DNA and/or RNA-binding factors implicated in chromatin remodeling and repair, gene regulation and mRNA stabilization/modification. The majority of these proteins, including a structural maintenance of chromosomes protein (CsSMC), a translin-associated factor X (CsTRAX), a VirE2-interacting protein (CsVIP2), a high mobility group (CsHMG) and two poly(A)-binding proteins (CsPABP1 and 2), interacted with each other, suggesting that they assemble into a multiprotein complex. CsHMG was shown to bind DNA and to interact with the invariable leucine-rich repeat region of PthAs. Surprisingly, both CsHMG and PthA4 interacted with PABP1 and 2 and showed selective binding to poly(U) RNA, a property that is novel among HMGs and TAL effectors. Given that homologs of CsHMG, CsPABP1, CsPABP2, CsSMC and CsTRAX in other organisms assemble into protein complexes to regulate mRNA stability and translation, we suggest a novel role of TAL effectors in mRNA processing and translational control.

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Schematic model of the citrus multiprotein complex comprising the PthA4-interacting partners.Protein-protein and protein-RNA contacts involving the PthA4 interactors based on the yeast two-hybrid, GST-pulldown and gel-shift assays described here, and literature data. The citrus multiprotein complex is reminiscent of that of mammalian miRISC involved in miRNA-mediated deadenylation [51]. Importin-α, which interacts with all PthA variants [16] is also a component of the cap-binding complex (CBC) which inhibits mRNA deadenylation when in the presence of a poly(A)-specific ribonucleases [55]–[58]. It is suggested that by interacting with such proteins and with poly(U) RNA (not necessarily simultaneously), PthA proteins may displace some of the components of this complex thought to promote deadenylation and mRNA decay and thus increase mRNA stabilization and translation initiation. U-rich sequences found in both 5′ and 3′ ends of mRNAs could represent binding sites of CsHMG and PthA4.
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pone-0032305-g008: Schematic model of the citrus multiprotein complex comprising the PthA4-interacting partners.Protein-protein and protein-RNA contacts involving the PthA4 interactors based on the yeast two-hybrid, GST-pulldown and gel-shift assays described here, and literature data. The citrus multiprotein complex is reminiscent of that of mammalian miRISC involved in miRNA-mediated deadenylation [51]. Importin-α, which interacts with all PthA variants [16] is also a component of the cap-binding complex (CBC) which inhibits mRNA deadenylation when in the presence of a poly(A)-specific ribonucleases [55]–[58]. It is suggested that by interacting with such proteins and with poly(U) RNA (not necessarily simultaneously), PthA proteins may displace some of the components of this complex thought to promote deadenylation and mRNA decay and thus increase mRNA stabilization and translation initiation. U-rich sequences found in both 5′ and 3′ ends of mRNAs could represent binding sites of CsHMG and PthA4.

Mentions: Several lines of evidence support this idea. Firstly, some of the protein-protein interactions identified by two-hybrid, relating for instance CsSMC with CsTRAX and CsPABP2, were confirmed by mass spectrometry analysis (Table 2). CsPABP1 and CsPABP2 are, respectively, homologous to human PABPN and PABPC, which function as mRNA stabilizing and translation initiation factors [46], [47]. Notably, translation initiation and elongation factors, as well as a homolog of tobacco AGO1 [48], were identified as binding partners of CsSMC and CsTRAX (Table 2). Furthermore, human PABPN binds to PABPC in the presence of RNA and to RRM proteins similar to CsRRMP1 in a protein complex involved in mRNA turnover [49], [50]. Most importantly however, mammalian PABPC was recently shown to be recruited by RISC and to associate with CAF1/CCR4-NOT deadenylases in a multiprotein complex that regulates gene silencing through a miRNA-mediated mRNA deadenylation [51], [52]. Thus, considering that TRAX is a component of RISC [40], [41], AGO1 was found associated with CsSMC and CsTRAX (Table 2), and CsVIP2 is a CCR4-NOT domain protein, we propose a model (Fig. 8) in which many the PthA4 interactors identified here are components of a multiprotein complex similar to the mammalian miRISC involved in miRNA-mediated deadenylation [51]. In this model, CsHMG and CsPCBP would bind to UC-rich sequences in the 3′ end of the mRNA [45], whereas PABP1 and PABP2 would attach to the adjacent poly(A) tail creating a scaffold for the assembly of CsSMC, CsTRAX, CsVIP2, CsRRMP1 and AGO1 (Fig. 8). The fact that PthA4 selectively binds poly(U) RNA, apparently forming a ternary complex with CsHMG, is also interesting. Even though not compatible with the TAL code [7], [8], it is worth noting that the DNA-binding domain of PthA, and related TAL effectors, shows a superhelical structure though to be similar to that of PPR domains [28], [53], [54] involved in the recognition of U-rich sequences in 5′ and 3′mRNA termini [29]–[31].


The TAL effector PthA4 interacts with nuclear factors involved in RNA-dependent processes including a HMG protein that selectively binds poly(U) RNA.

de Souza TA, Soprano AS, de Lira NP, Quaresma AJ, Pauletti BA, Paes Leme AF, Benedetti CE - PLoS ONE (2012)

Schematic model of the citrus multiprotein complex comprising the PthA4-interacting partners.Protein-protein and protein-RNA contacts involving the PthA4 interactors based on the yeast two-hybrid, GST-pulldown and gel-shift assays described here, and literature data. The citrus multiprotein complex is reminiscent of that of mammalian miRISC involved in miRNA-mediated deadenylation [51]. Importin-α, which interacts with all PthA variants [16] is also a component of the cap-binding complex (CBC) which inhibits mRNA deadenylation when in the presence of a poly(A)-specific ribonucleases [55]–[58]. It is suggested that by interacting with such proteins and with poly(U) RNA (not necessarily simultaneously), PthA proteins may displace some of the components of this complex thought to promote deadenylation and mRNA decay and thus increase mRNA stabilization and translation initiation. U-rich sequences found in both 5′ and 3′ ends of mRNAs could represent binding sites of CsHMG and PthA4.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3285215&req=5

pone-0032305-g008: Schematic model of the citrus multiprotein complex comprising the PthA4-interacting partners.Protein-protein and protein-RNA contacts involving the PthA4 interactors based on the yeast two-hybrid, GST-pulldown and gel-shift assays described here, and literature data. The citrus multiprotein complex is reminiscent of that of mammalian miRISC involved in miRNA-mediated deadenylation [51]. Importin-α, which interacts with all PthA variants [16] is also a component of the cap-binding complex (CBC) which inhibits mRNA deadenylation when in the presence of a poly(A)-specific ribonucleases [55]–[58]. It is suggested that by interacting with such proteins and with poly(U) RNA (not necessarily simultaneously), PthA proteins may displace some of the components of this complex thought to promote deadenylation and mRNA decay and thus increase mRNA stabilization and translation initiation. U-rich sequences found in both 5′ and 3′ ends of mRNAs could represent binding sites of CsHMG and PthA4.
Mentions: Several lines of evidence support this idea. Firstly, some of the protein-protein interactions identified by two-hybrid, relating for instance CsSMC with CsTRAX and CsPABP2, were confirmed by mass spectrometry analysis (Table 2). CsPABP1 and CsPABP2 are, respectively, homologous to human PABPN and PABPC, which function as mRNA stabilizing and translation initiation factors [46], [47]. Notably, translation initiation and elongation factors, as well as a homolog of tobacco AGO1 [48], were identified as binding partners of CsSMC and CsTRAX (Table 2). Furthermore, human PABPN binds to PABPC in the presence of RNA and to RRM proteins similar to CsRRMP1 in a protein complex involved in mRNA turnover [49], [50]. Most importantly however, mammalian PABPC was recently shown to be recruited by RISC and to associate with CAF1/CCR4-NOT deadenylases in a multiprotein complex that regulates gene silencing through a miRNA-mediated mRNA deadenylation [51], [52]. Thus, considering that TRAX is a component of RISC [40], [41], AGO1 was found associated with CsSMC and CsTRAX (Table 2), and CsVIP2 is a CCR4-NOT domain protein, we propose a model (Fig. 8) in which many the PthA4 interactors identified here are components of a multiprotein complex similar to the mammalian miRISC involved in miRNA-mediated deadenylation [51]. In this model, CsHMG and CsPCBP would bind to UC-rich sequences in the 3′ end of the mRNA [45], whereas PABP1 and PABP2 would attach to the adjacent poly(A) tail creating a scaffold for the assembly of CsSMC, CsTRAX, CsVIP2, CsRRMP1 and AGO1 (Fig. 8). The fact that PthA4 selectively binds poly(U) RNA, apparently forming a ternary complex with CsHMG, is also interesting. Even though not compatible with the TAL code [7], [8], it is worth noting that the DNA-binding domain of PthA, and related TAL effectors, shows a superhelical structure though to be similar to that of PPR domains [28], [53], [54] involved in the recognition of U-rich sequences in 5′ and 3′mRNA termini [29]–[31].

Bottom Line: Plant pathogenic bacteria utilize an array of effector proteins to cause disease.Surprisingly, both CsHMG and PthA4 interacted with PABP1 and 2 and showed selective binding to poly(U) RNA, a property that is novel among HMGs and TAL effectors.Given that homologs of CsHMG, CsPABP1, CsPABP2, CsSMC and CsTRAX in other organisms assemble into protein complexes to regulate mRNA stability and translation, we suggest a novel role of TAL effectors in mRNA processing and translational control.

View Article: PubMed Central - PubMed

Affiliation: Laboratório Nacional de Biociências, Centro Nacional de Pesquisa em Energia e Materiais, Campinas, SP, Brazil.

ABSTRACT
Plant pathogenic bacteria utilize an array of effector proteins to cause disease. Among them, transcriptional activator-like (TAL) effectors are unusual in the sense that they modulate transcription in the host. Although target genes and DNA specificity of TAL effectors have been elucidated, how TAL proteins control host transcription is poorly understood. Previously, we showed that the Xanthomonas citri TAL effectors, PthAs 2 and 3, preferentially targeted a citrus protein complex associated with transcription control and DNA repair. To extend our knowledge on the mode of action of PthAs, we have identified new protein targets of the PthA4 variant, required to elicit canker on citrus. Here we show that all the PthA4-interacting proteins are DNA and/or RNA-binding factors implicated in chromatin remodeling and repair, gene regulation and mRNA stabilization/modification. The majority of these proteins, including a structural maintenance of chromosomes protein (CsSMC), a translin-associated factor X (CsTRAX), a VirE2-interacting protein (CsVIP2), a high mobility group (CsHMG) and two poly(A)-binding proteins (CsPABP1 and 2), interacted with each other, suggesting that they assemble into a multiprotein complex. CsHMG was shown to bind DNA and to interact with the invariable leucine-rich repeat region of PthAs. Surprisingly, both CsHMG and PthA4 interacted with PABP1 and 2 and showed selective binding to poly(U) RNA, a property that is novel among HMGs and TAL effectors. Given that homologs of CsHMG, CsPABP1, CsPABP2, CsSMC and CsTRAX in other organisms assemble into protein complexes to regulate mRNA stability and translation, we suggest a novel role of TAL effectors in mRNA processing and translational control.

Show MeSH
Related in: MedlinePlus