Abstract
BackgroundHistone H2B monoubiquitination pathway has been shown to play critical roles in regulating growth/development and stress response in Arabidopsis. In the present study, we explored the involvement of the tomato histone H2B monoubiquitination pathway in defense response against Botrytis cinerea by functional analysis of SlHUB1 and SlHUB2, orthologues of the Arabidopsis AtHUB1/AtHUB2.MethodsWe used the TRV-based gene silencing system to knockdown the expression levels of SlHUB1 or SlHUB2 in tomato plants and compared the phenotype between the silenced and the control plants after infection with B. cinerea and Pseudomonas syringae pv. tomato (Pst) DC3000. Biochemical and interaction properties of proteins were examined using in vitro histone monoubiquitination and yeast two-hybrid assays, respectively. The transcript levels of genes were analyzed by quantitative real time PCR (qRT-PCR).ResultsThe tomato SlHUB1 and SlHUB2 had H2B monoubiquitination E3 ligases activity in vitro and expression of SlHUB1 and SlHUB2 was induced by infection of B. cinerea and Pst DC3000 and by treatment with salicylic acid (SA) and 1-amino cyclopropane-1-carboxylic acid (ACC). Silencing of either SlHUB1 or SlHUB2 in tomato plants showed increased susceptibility to B. cinerea, whereas silencing of SlHUB1 resulted in increased resistance against Pst DC3000. SlMED21, a Mediator complex subunit, interacted with SlHUB1 but silencing of SlMED21 did not affect the disease resistance to B. cinerea and Pst DC3000. The SlHUB1- and SlHUB2-silenced plants had thinner cell wall but increased accumulation of reactive oxygen species (ROS), increased callose deposition and exhibited altered expression of the genes involved in phenylpropanoid pathway and in ROS generation and scavenging system. Expression of genes in the SA-mediated signaling pathway was significantly upregulated, whereas expression of genes in the jasmonic acid (JA)/ethylene (ET)-mediated signaling pathway were markedly decreased in SlHUB1- and SlHUB2-silenced plants after infection of B. cinerea.ConclusionVIGS-based functional analyses demonstrate that both SlHUB1 and SlHUB2 contribute to resistance against B. cinerea most likely through modulating the balance between the SA- and JA/ET-mediated signaling pathways.Electronic supplementary materialThe online version of this article (doi:10.1186/s12870-015-0614-2) contains supplementary material, which is available to authorized users.
Highlights
Histone H2B monoubiquitination pathway has been shown to play critical roles in regulating growth/ development and stress response in Arabidopsis
We found that virus-induced gene silencing (VIGS) of either SlHUB1 or SlHUB2 in tomato plants resulted in increased susceptibility to B. cinerea and led to thinner cell wall, increased accumulation of reactive oxygen species (ROS) and callose around the infection sites, demonstrating that both of the SlHUB1 and SlHUB2 are positive regulators of defense response against B. cinerea most likely through modulation of cell wall strengthen and ROS balance
Silencing of SlHUB1 but not SlHUB2 and SlMED21 affected resistance to Pst DC3000 To explore the possible involvement of SlHUB1, SlHUB2 and SlMED21 in defense response against other pathogens, we further examined whether silencing of SlHUB1, SlHUB2 or SlMED21 affects the resistance to Pst DC3000
Summary
Histone H2B monoubiquitination pathway has been shown to play critical roles in regulating growth/ development and stress response in Arabidopsis. The first layer is triggered upon the detection of pathogen- or microbial-associated molecular patterns (PAMPs/MAMPs) by pattern recognition receptors on the external face of plant cells and is called PAMPtriggered immunity (PTI) [2]. Plants have acquired additional intracellular receptors called resistance (R) proteins to recognize pathogen effectors, resulting in initiation of the second layer of defense, known as effector-triggered immunity (ETI) [1, 4,5,6]. Extensive genetic and biochemical studies have shown that ubiquitin-mediated protein modification plays critical roles in plant immune responses [8, 11,12,13,14,15]
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