ETHYLENE-INSENSITIVE3 Is a Senescence-Associated Gene That Accelerates Age-Dependent Leaf Senescence by Directly Repressing miR164 Transcription in Arabidopsis    

Abstract

Numerous endogenous and environmental signals regulate the intricate and highly orchestrated process of plant senescence. Ethylene is a well-known inducer of senescence, including fruit ripening and flower and leaf senescence. However, the underlying molecular mechanism of ethylene-induced leaf senescence remains to be elucidated. Here, we examine ETHYLENE-INSENSITIVE3 (EIN3), a key transcription factor in ethylene signaling, and find that EIN3 is a functional senescence-associated gene. Constitutive overexpression or temporary activation of EIN3 is sufficient to accelerate leaf senescence symptoms. Conversely, loss of EIN3 and EIN3-Like1 (its close homolog) function leads to a delay in agedependent and ethylene-, jasmonic acid-, or dark-induced leaf senescence. We further found that EIN3 acts downstream of ORESARA2 (ORE2)/ORE3/EIN2 to repress miR164 transcription and upregulate the transcript levels of ORE1/NAC2, a target gene of miR164. EIN3 directly binds to the promoters of microRNA164 (miR164), and this binding activity progressively increases during leaf ageing. Genetic analysis revealed that overexpression of miR164 or knockout of ORE1/NAC2 represses EIN3-induced early-senescence phenotypes. Collectively, our study defines a continuation of the signaling pathway involving EIN2-EIN3-miR164-NAC2 in regulating leaf senescence and provides a mechanistic insight into how ethylene promotes the progression of leaf senescence in Arabidopsis thaliana.

A Genome-Wide Chronological Study of Gene Expression and Two Histone Modifications, H3K4me3 and H3K9ac, during Developmental Leaf Senescence

“…WRKY53 expression is down-regulated by Whirly1 (Miao et al, 2013), which may explain the coincidence of low transcript levels and high levels of H3K4me3 marks. Examples of posttranscriptional regulation mediated by small RNAs have been identified during leaf senescence (Kim et al, 2009a(Kim et al, , 2009bLi et al, 2013b;Thatcher et al, 2015) and may also explain some of the inconsistencies between H3K4me3 marks and gene expression. Senescence associated gene12 (SAG12), a molecular marker for senescence, was up-regulated 3,300-fold between 29 and 57 d (0.022-72.5 RPKM) and showed increased levels of H3K4me3 marks 250 to 650 bp upstream of the TSS but no clear H3K9ac marks (Supplemental Fig.…”

“…Toward understanding the mechanisms that regulate gene expression during senescence, numerous studies have shown that transcription factors, such as WRKY53 (Hinderhofer and Zentgraf, 2001;Miao et al, 2004;Miao and Zentgraf, 2010), ORESARA1/NAC2 (Oh et al, 1997;Matallana-Ramirez et al, 2013), and ETHYLENE-INSENSITIVE3 (Li et al, 2013b), promote leaf senescence as well as many others listed in the Leaf Senescence Database (Liu et al, 2011). In addition, mutants that affect chromatin structure have been shown to affect senescence; however, phenotypes are often pleiotropic (Lim et al, 2007b;Wu et al, 2008;Ay et al, 2009;Humbeck, 2013).…”

Ethylene Exerts Species-Specific and Age-Dependent Control of Photosynthesis

“…It 226 has been shown that external ethylene stimulates leaf senescence (Bleecker et al, 1988;Zacarias & 227 Reid, 1990), while ethylene-insensitive mutants (etr1-1, ers1, ein2-1, ein2-5, ein3-1 and ein3-1 eil1-1 228 in Arabidopsis and Nr in tomato) show a reduced rate of senescence (Bleecker, 1988;Hua et al, 229 1995;Grbic and Bleecker, 1995;Oh et al, 1997;Lanahan et al, 1994, Li et al, 2013Kim et al, 2014). 230…”

“…It can activate the expression of NAP (NAM/ATAF1,2/CUC2) and NAC2 (ORE1/NAC092), two 252 important transcription factors controlling leaf senescence Kim et al, 2009;Kim et 253 al., 2014). Furthermore, EIN3 binds to the promotor of the microRNA (miRNA) miR164 and 254 progressively inhibits its expression during leaf development (Li et al, 2013). In turn, miR164 inhibits 255 the expression of the transcription factor NAC2 (Kim et al, 2009;Kim et al, 2014;Li et al, 2013), 256 unraveling a dual feed-forward and feedback regulation of senescence by ethylene through the 257 action of EIN3 ( Figure 1).…”

Interaction between MYC2 and ETHYLENE INSENSITIVE3 Modulates Antagonism between Jasmonate and Ethylene Signaling inArabidopsis

“…Both ethylene (ET) and jasmonate (JA) are essential plant hormones that regulate various plant developmental processes and diverse defense responses (Kieber, 1997;Bleecker and Kende, 2000;Guo and Ecker, 2004;Broekaert et al, 2006;Howe and Jander, 2008;Browse, 2009;Shan et al, 2012;Wasternack and Hause, 2013). ET signal is perceived by its receptors ETHYLENE RESPONSE1 (ETR1), ETR2, ETHYLENE RESPONSE SENSOR1 (ERS1), ERS2, and ETHYLENE INSENSITIVE4 (EIN4) (Hua and Meyerowitz, 1998) to repress CONSTITUTIVE TRIPLE RESPONSE1 (CTR1) (Kieber et al, 1993), which activates EIN2 (Alonso et al, 1999;Ju et al, 2012;Qiao et al, 2012;Wen et al, 2012) and subsequently stabilizes EIN3 and EIN3-LIKE1 (EIL1) (Chao et al, 1997;Guo and Ecker, 2003;Potuschak et al, 2003;Gagne et al, 2004) to mediate various ET responses, including hypocotyl growth (Zhong et al, 2012), apical hook formation (Knight et al, 1910;An et al, 2012), root growth (Ortega-Martínez et al, 2007;Růzicka et al, 2007), flowering (Ogawara et al, 2003;Achard et al, 2007), fruit ripening (Burg and Burg, 1962;Theologis et al, 1992), leaf senescence (Gepstein and Thimann, 1981;Li et al, 2013), freezing tolerance , and resistance against pathogen infection (Alonso et al, 2003;Chen et al, 2009).…”