TAL Effectors Target the C-Terminal Domain of RNA Polymerase II (CTD) by Inhibiting the Prolyl-Isomerase Activity of a CTD-Associated Cyclophilin

Mariane Noronha Domingues,Celso Eduardo Benedetti,Bruna Medeia de Campos,Uli Quirino de Mello,Maria Luiza Peixoto de Oliveira,John W. Stiller

doi:10.1371/journal.pone.0041553

Abstract

Transcriptional activator-like (TAL) effectors of plant pathogenic bacteria function as transcription factors in plant cells. However, how TAL effectors control transcription in the host is presently unknown. Previously, we showed that TAL effectors of the citrus canker pathogen Xanthomonas citri, named PthAs, targeted the citrus protein complex comprising the thioredoxin CsTdx, ubiquitin-conjugating enzymes CsUev/Ubc13 and cyclophilin CsCyp. Here we show that CsCyp complements the function of Cpr1 and Ess1, two yeast cyclophilins that regulate transcription by the isomerization of proline residues of the regulatory C-terminal domain (CTD) of RNA polymerase II. We also demonstrate that CsCyp, CsTdx, CsUev and four PthA variants interact with the citrus CTD and that CsCyp co-immunoprecipitate with the CTD in citrus cell extracts and with PthA2 transiently expressed in sweet orange epicotyls. The interactions of CsCyp with the CTD and PthA2 were inhibited by cyclosporin A (CsA), a cyclophilin inhibitor. Moreover, we present evidence that PthA2 inhibits the peptidyl-prolyl cis-trans isomerase (PPIase) activity of CsCyp in a similar fashion as CsA, and that silencing of CsCyp, as well as treatments with CsA, enhance canker lesions in X. citri-infected leaves. Given that CsCyp appears to function as a negative regulator of cell growth and that Ess1 negatively regulates transcription elongation in yeast, we propose that PthAs activate host transcription by inhibiting the PPIase activity of CsCyp on the CTD.

Highlights

Xanthomonas citri, the bacterial pathogen responsible for citrus canker, induces hyperplasic lesions and pustule formation on the host epidermis which result from the increased division and expansion of the plant cells at the site of infection [1]
Similar to Cpr1, CsCyp suppressed the ess1H164R mutation and restored the growth of the ess1 mutant at the non-permissive temperature (Fig. 1C), indicating that its peptidyl-prolyl cis-trans isomerase (PPIase) activity complements that of Ess1
We show that all variants of the effector protein PthA of a X. citri pathogen interact with the Citrus sinensis (Cs)-terminal domain (CTD) of the citrus RNA pol II

Summary

Introduction

Xanthomonas citri, the bacterial pathogen responsible for citrus canker, induces hyperplasic lesions and pustule formation on the host epidermis which result from the increased division and expansion of the plant cells at the site of infection [1]. Many of the X. citri-induced genes, including those encoding cellulases and expansins, involved in cell-wall remodeling, were found to be regulated by auxin and gibberellin, but most importantly, both auxin and gibberellin were shown to be required for initial canker development [3] These data support the idea that X. citri promotes cell division and enlargement through changes in the auxin and gibberellin signaling pathways, how exactly the bacterium reprograms transcription in the host is not entirely clear. TAL effectors of the AvrBs3/PthA protein family are translocated into the plant cell by the type-III secretion system and targeted to the nucleus where they function as transcriptional activators [8]. The interaction of a TAL effector with its target DNA is mediated by the repeat domain, an internal region of the protein comprising variable, nearly identical tandem repeats of 34 amino acids that define the DNA specificity and determines pathogenicity and avirulence [13,14]

Methods

Results

Conclusion