**3.2 The negative effect of abiotic and biotic stress on photosynthetic gene expression**

The down-regulation of photosynthetic gene transcription is frequently observed in plants subjected to stress. Thereby, environmental stresses, including drought, salinity and low temperatures can negatively affect photosynthetic gene expression in addition to an induction of compensating physiological and biochemical alterations (Saibo et al., 2009; Chaves et al., 2009). Similarly, a whole gamut of biotic insults caused by arthropods, fungi, bacteria and viral pathogens triggers a uniform and apparently regulated reduction in transcription of nuclear genes coding for the major components of photosynthesis, regardless of the plant host. The widespread negative effect on photosynthesis caused by biotic stressors was recently evidenced by a meta-genomic analysis in which the transcriptome data from microarray experiments representing twenty two different forms of

Friend or Foe? Exploring the Factors that Determine the Difference

**3.4 Main photosynthetic genes targeted by biotic stress** 

of the protein domains involved in ATP binding (Thivierge et al., 2010).

Curiously, the gene coding for the Rubisco enzyme, an absolutely vital component of the carbon assimilation machinery in plants, was found to be one of the primary photosynthetic genes targeted by herbivore attack, in addition to genes coding for the components of the antenna complexes in both photosystems (Logemann et al., 1995; Ehness et al., 1997; Hermsmeier et al., 2001; Hahlbrock et al., 2003; Hui et al., 2003; Montesano et al., 2004; Zou et al., 2005). Proteomic investigations provided further evidence of the vulnerability of the CO2 fixing process to insect attack by showing that herbivory, or even the application of the insect´s saliva, also reduced the abundance of Rubisco activase (RCA) in *Nicotiana attenuata* and Arabidopsis (Giri et al., 2006; Thivierge et al., 2010). RCA is a key regulatory enzyme of photosynthetic carbon assimilation that modulates the activity of Rubisco by facilitating the removal of inhibiting sugar phosphates from its active site (Portis, 1995). Additional findings derived from the proteomic experimental approaches indicated that Rubisco large subunit and RCA, in addition to PS I P700 apoprotein A1 suffered several caterpillar-specific modifications, including the conversion of Cys192 in Rubisco to the the thiolate anion, which may lead to decreased enzyme activity and protein degradation, and adverse modifications, in RCA,

et al., 2009).

Between Positive and Negative Effects on Photosynthesis in Response to Insect Herbivory 171

1997; Wasternack & Parthier, 1997; Hui et al., 2003; Reymond et al., 2004; Giri et al., 2006; Zavala & Baldwin, 2006; Yan et al., 2007). Ample evidence demonstrating the direct and indirect negative effect on plant growth and/or photosynthesis exerted by JA is available. Early reports described that JA treatment of barley leaves inhibits activity of PSII electron transport (Maslenkova et al., 1999), whereas barley plants treated with methyl jasmonate (MeJA) suffered a repressed translation of transcripts coding for Rubisco small subunit, chlorophyll a/b binding protein and photosystem II proteins (Roloff et al., 1994). Moreover, plants treated with MeJA or genetically manipulated to accumulate higher JA concentrations were found to develop shorter petioles or have a reduced total seed production (Cipollini, 2007; Bonaventure et al., 2007), or suffer reduced root growth (Henkes et al., 2008). Also, induction by diverse types of herbivores of the lipoxygenase pathway, which represents the initial step of JA biosynthesis and jasmonate signaling, was found to be associated with reduced photosynthesis and vegetative growth (Heidel & Baldwin, 2004; De Vos et al., 2005; Kempema et al., 2007; Bilgin et al., 2010), similarly to experiments showing that herbivore-induced JA signaling suppressed re-growth and contributed to apical dominance (Zavala & Baldwin, 2006). It has been proposed that the above effects on plant growth are modulated by the gene *JASMONATE-ASSOCIATED1*  (*JAS1*) (Yan et al., 2007). Moreover, a cross-talk between ABA- and JA-responsive gene expression in response to insect herbivory, mediated by the action of MYC and MYB TFs, has been proposed as a mechanism to coordinate the expression of defensive and dehydration-responsive genes (Yamaguchi-Shinozaki & Shinozaki, 2006). Such interaction is deemed to be needed by the plant in order to deal with the increased leaf dehydration and accompanying senescence produced by defoliating herbivores (Aldea et al., 2005; Lim et al., 2007; see below). The observed influence of light quality and perception by phytochromes on JA induced defense responses and resource allocation was also indicative of an indirect connection between JA and photosynthesis (Ballare, 2009; Moreno

biotic damage on eight different plant species (predominantly Arabidopsis, five other herbaceous species and two tree species) was performed. In this study, transcript levels of photosynthesis light reaction, carbon reduction cycle and pigment synthesis genes were decreased regardless of the type of biotic attack. Interestingly, down-regulation of photosynthesis-related genes contrasted with the corresponding up-regulation of genes coding for the synthesis of JA and those involved in the responses to SA and ethylene. This clear difference in expression patterns suggested that the up-regulation of defense-related biosynthetic genes could be part of the overall defense response responsible for re-allocating resources from growth to defense (see below; Bilgin et al., 2010).

Apparently, insect herbivory caused by chewing insects, had the weakest negative effect on photosynthetic gene expression as compared to other biotic stressors, whereas a general down-regulation of photosynthesis genes was observed in plants infested by aphids and the whitefly *Bemisia tabaci* (Bt) (Heidel & Baldwin, 2004; Zhu-Salzman et al., 2004; Voelckel et al., 2004; Qubbaj et al., 2005; Yuan et al., 2005; Kempema et al., 2007; Bilgin et al., 2010). Additional data pertaining modifications in gene expression produced by Bt larval feeding in tomato plants at different stages of development was generated using a suppressionsubtractive-hybridization (SSH) approach (Estrada-Hernández et al., 2009). In addition to the identification of several genes whose expression was differentially modified at different larval phases during the infestation process, the study showed a down-regulation of photosynthetic gene expression which was in accordance to the general negative trend associated with biotic-stress. However, upon closer examination, it became apparent that a more or less defined phase-dependent change in photosynthetic gene expression occurred in response to Bt infestation, which favored an up-regulation of photosystem II genes in the late two phases of Bt development in detriment of genes coding for components of other photosynthetic complexes, and also of the oxygen evolving complex and the Calvin cycle. A similar behavior was observed in Bt-infested tomatillo (*Physalis* spp.) plants (C. Sánchez-Hernández, personal communication). This led to the proposal that the contrasting pattern of gene expression, which occurred concomitantly with an up-regulation of oxidative stress genes leading to tissue senescence, could represent an additional strategy used by Bt, besides their reported ability avoid plant defenses, to favor infestation, (Walling, 2008; Estrada-Hernández et al., 2009; Délano-Frier et al., 2009). Support for this proposal was recently provided by a study showing that the application of the bacterial toxin coronatine to tomato seedlings, led to a reduced expression of photosynthesis related genes and a 1.5 to 2-fold reduction in maximum quantum efficiency of PS II, which occurred concomitantly with ROS generation and necrotic cell death (Ishiga et al., 2009)

Mention should be made, however, that given the long functional lifetime of most photosynthesis-related proteins (the highly labile D1 protein is a notable exception, see above), reduced gene photosynthetic gene expression does not necessarily translate into an immediate loss of function. Such behavior is believed to permit reallocation of nitrogen to the defense response, due to repressed transcription, without significantly affecting the rate of carbon assimilation (Bilgin et al., 2010).

#### **3.3 The role of JA in on photosynthetic gene expression and growth regulation**

Jasmonates play a central role in regulating plant defense responses to herbivores (Howe & Jander, 2008; Spoel & Dong, 2008) and also inhibit growth and photosynthesis by participating in the down-regulation of photosynthesis-related genes (Creelman & Mullet,

biotic damage on eight different plant species (predominantly Arabidopsis, five other herbaceous species and two tree species) was performed. In this study, transcript levels of photosynthesis light reaction, carbon reduction cycle and pigment synthesis genes were decreased regardless of the type of biotic attack. Interestingly, down-regulation of photosynthesis-related genes contrasted with the corresponding up-regulation of genes coding for the synthesis of JA and those involved in the responses to SA and ethylene. This clear difference in expression patterns suggested that the up-regulation of defense-related biosynthetic genes could be part of the overall defense response responsible for re-allocating

Apparently, insect herbivory caused by chewing insects, had the weakest negative effect on photosynthetic gene expression as compared to other biotic stressors, whereas a general down-regulation of photosynthesis genes was observed in plants infested by aphids and the whitefly *Bemisia tabaci* (Bt) (Heidel & Baldwin, 2004; Zhu-Salzman et al., 2004; Voelckel et al., 2004; Qubbaj et al., 2005; Yuan et al., 2005; Kempema et al., 2007; Bilgin et al., 2010). Additional data pertaining modifications in gene expression produced by Bt larval feeding in tomato plants at different stages of development was generated using a suppressionsubtractive-hybridization (SSH) approach (Estrada-Hernández et al., 2009). In addition to the identification of several genes whose expression was differentially modified at different larval phases during the infestation process, the study showed a down-regulation of photosynthetic gene expression which was in accordance to the general negative trend associated with biotic-stress. However, upon closer examination, it became apparent that a more or less defined phase-dependent change in photosynthetic gene expression occurred in response to Bt infestation, which favored an up-regulation of photosystem II genes in the late two phases of Bt development in detriment of genes coding for components of other photosynthetic complexes, and also of the oxygen evolving complex and the Calvin cycle. A similar behavior was observed in Bt-infested tomatillo (*Physalis* spp.) plants (C. Sánchez-Hernández, personal communication). This led to the proposal that the contrasting pattern of gene expression, which occurred concomitantly with an up-regulation of oxidative stress genes leading to tissue senescence, could represent an additional strategy used by Bt, besides their reported ability avoid plant defenses, to favor infestation, (Walling, 2008; Estrada-Hernández et al., 2009; Délano-Frier et al., 2009). Support for this proposal was recently provided by a study showing that the application of the bacterial toxin coronatine to tomato seedlings, led to a reduced expression of photosynthesis related genes and a 1.5 to 2-fold reduction in maximum quantum efficiency of PS II, which occurred concomitantly

Mention should be made, however, that given the long functional lifetime of most photosynthesis-related proteins (the highly labile D1 protein is a notable exception, see above), reduced gene photosynthetic gene expression does not necessarily translate into an immediate loss of function. Such behavior is believed to permit reallocation of nitrogen to the defense response, due to repressed transcription, without significantly affecting the rate

**3.3 The role of JA in on photosynthetic gene expression and growth regulation** 

Jasmonates play a central role in regulating plant defense responses to herbivores (Howe & Jander, 2008; Spoel & Dong, 2008) and also inhibit growth and photosynthesis by participating in the down-regulation of photosynthesis-related genes (Creelman & Mullet,

resources from growth to defense (see below; Bilgin et al., 2010).

with ROS generation and necrotic cell death (Ishiga et al., 2009)

of carbon assimilation (Bilgin et al., 2010).

1997; Wasternack & Parthier, 1997; Hui et al., 2003; Reymond et al., 2004; Giri et al., 2006; Zavala & Baldwin, 2006; Yan et al., 2007). Ample evidence demonstrating the direct and indirect negative effect on plant growth and/or photosynthesis exerted by JA is available. Early reports described that JA treatment of barley leaves inhibits activity of PSII electron transport (Maslenkova et al., 1999), whereas barley plants treated with methyl jasmonate (MeJA) suffered a repressed translation of transcripts coding for Rubisco small subunit, chlorophyll a/b binding protein and photosystem II proteins (Roloff et al., 1994). Moreover, plants treated with MeJA or genetically manipulated to accumulate higher JA concentrations were found to develop shorter petioles or have a reduced total seed production (Cipollini, 2007; Bonaventure et al., 2007), or suffer reduced root growth (Henkes et al., 2008). Also, induction by diverse types of herbivores of the lipoxygenase pathway, which represents the initial step of JA biosynthesis and jasmonate signaling, was found to be associated with reduced photosynthesis and vegetative growth (Heidel & Baldwin, 2004; De Vos et al., 2005; Kempema et al., 2007; Bilgin et al., 2010), similarly to experiments showing that herbivore-induced JA signaling suppressed re-growth and contributed to apical dominance (Zavala & Baldwin, 2006). It has been proposed that the above effects on plant growth are modulated by the gene *JASMONATE-ASSOCIATED1*  (*JAS1*) (Yan et al., 2007). Moreover, a cross-talk between ABA- and JA-responsive gene expression in response to insect herbivory, mediated by the action of MYC and MYB TFs, has been proposed as a mechanism to coordinate the expression of defensive and dehydration-responsive genes (Yamaguchi-Shinozaki & Shinozaki, 2006). Such interaction is deemed to be needed by the plant in order to deal with the increased leaf dehydration and accompanying senescence produced by defoliating herbivores (Aldea et al., 2005; Lim et al., 2007; see below). The observed influence of light quality and perception by phytochromes on JA induced defense responses and resource allocation was also indicative of an indirect connection between JA and photosynthesis (Ballare, 2009; Moreno et al., 2009).
