RESUMO
Post-translational histone modifications have a critical role in regulating transcription, the cell cycle, DNA replication and DNA damage repair. The identification of new histone modifications critical for transcriptional regulation at initiation, elongation or termination is of particular interest. Here we report a new layer of regulation in transcriptional elongation that is conserved from yeast to mammals. This regulation is based on the phosphorylation of a highly conserved tyrosine residue, Tyr 57, in histone H2A and is mediated by the unsuspected tyrosine kinase activity of casein kinase 2 (CK2). Mutation of Tyr 57 in H2A in yeast or inhibition of CK2 activity impairs transcriptional elongation in yeast as well as in mammalian cells. Genome-wide binding analysis reveals that CK2α, the catalytic subunit of CK2, binds across RNA-polymerase-II-transcribed coding genes and active enhancers. Mutation of Tyr 57 causes a loss of H2B mono-ubiquitination as well as H3K4me3 and H3K79me3, histone marks associated with active transcription. Mechanistically, both CK2 inhibition and the H2A(Y57F) mutation enhance H2B deubiquitination activity of the Spt-Ada-Gcn5 acetyltransferase (SAGA) complex, suggesting a critical role of this phosphorylation in coordinating the activity of the SAGA complex during transcription. Together, these results identify a new component of regulation in transcriptional elongation based on CK2-dependent tyrosine phosphorylation of the globular domain of H2A.
Assuntos
Caseína Quinase II/metabolismo , Histonas/química , Histonas/metabolismo , Elongação da Transcrição Genética , Tirosina/metabolismo , Sequência de Aminoácidos , Linhagem Celular , Sequência Conservada , Histonas/genética , Humanos , Dados de Sequência Molecular , Fosforilação , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Tirosina/química , Ubiquitinação/genéticaRESUMO
XopN is a type III effector protein from Xanthomonas campestris pathovar vesicatoria that suppresses PAMP-triggered immunity (PTI) in tomato. Previous work reported that XopN interacts with the tomato 14-3-3 isoform TFT1; however, TFT1's role in PTI and/or XopN virulence was not determined. Here we show that TFT1 functions in PTI and is a XopN virulence target. Virus-induced gene silencing of TFT1 mRNA in tomato leaves resulted in increased growth of Xcv ΔxopN and Xcv ΔhrpF demonstrating that TFT1 is required to inhibit Xcv multiplication. TFT1 expression was required for Xcv-induced accumulation of PTI5, GRAS4, WRKY28, and LRR22 mRNAs, four PTI marker genes in tomato. Deletion analysis revealed that the XopN C-terminal domain (amino acids 344-733) is sufficient to bind TFT1. Removal of amino acids 605-733 disrupts XopN binding to TFT1 in plant extracts and inhibits XopN-dependent virulence in tomato, demonstrating that these residues are necessary for the XopN/TFT1 interaction. Phos-tag gel analysis and mass spectrometry showed that XopN is phosphorylated in plant extracts at serine 688 in a putative 14-3-3 recognition motif. Mutation of S688 reduced XopN's phosphorylation state but was not sufficient to inhibit binding to TFT1 or reduce XopN virulence. Mutation of S688 and two leucines (L64,L65) in XopN, however, eliminated XopN binding to TFT1 in plant extracts and XopN virulence. L64 and L65 are required for XopN to bind TARK1, a tomato atypical receptor kinase required for PTI. This suggested that TFT1 binding to XopN's C-terminal domain might be stabilized via TARK1/XopN interaction. Pull-down and BiFC analyses show that XopN promotes TARK1/TFT1 complex formation in vitro and in planta by functioning as a molecular scaffold. This is the first report showing that a type III effector targets a host 14-3-3 involved in PTI to promote bacterial pathogenesis.
Assuntos
Proteínas 14-3-3/metabolismo , Doenças das Plantas/imunologia , Proteínas de Plantas/metabolismo , Receptores de Reconhecimento de Padrão/metabolismo , Solanum lycopersicum/microbiologia , Transposases/metabolismo , Xanthomonas campestris/patogenicidade , Proteínas 14-3-3/genética , Proteínas 14-3-3/imunologia , Sistemas de Secreção Bacterianos/genética , Sistemas de Secreção Bacterianos/imunologia , Inativação Gênica , Solanum lycopersicum/genética , Solanum lycopersicum/imunologia , Solanum lycopersicum/metabolismo , Mutação , Doenças das Plantas/genética , Proteínas de Plantas/genética , Proteínas de Plantas/imunologia , RNA Mensageiro/análise , Reação em Cadeia da Polimerase em Tempo Real , Receptores de Reconhecimento de Padrão/genética , Receptores de Reconhecimento de Padrão/imunologia , Transposases/genética , Transposases/imunologia , Virulência/genética , Xanthomonas campestris/enzimologia , Xanthomonas campestris/genéticaRESUMO
Cellular synthesis of phytic acid sequesters phosphates in the sugar inositol. Phytic acid in soil represents the most abundant form of organic phosphates. The supplementation of phytase or phytase-producing organisms has been considered as a strategy to improve usable soil phosphates. However, the impacts on the environmental flow of inositol, which is generated along with phosphate by phytase, have not been examined. In this review, we discuss the origin and nature of inositol produced in soil and the several possible destinations of inositol released by phytase activities. We emphasise how an improved understanding of soil inositol flow could help to provide new solutions to the phosphate shortage problem in agriculture.