Dynamics and origin of cytokinins involved in plant manipulation by a leaf‐mining insect

Abstract

Several herbivorous insects and plant-associated microorganisms control the phytohormonal balance, thus enabling them to successfully exploit the plant by inhibiting plant defenses and withdrawing plant resources for their own benefit. The leaf-mining moth Phyllonorycter blancardella modifies the cytokinin (CK) profile of mined leaf-tissues, and the insect symbiotic bacteria Wolbachia is involved in the plant manipulation to the benefit of the insect host. To gain a deeper understanding into the possible origin and dynamics of CKs, we conducted an extensive characterization of CKs in larvae and in infected apple leaves. Our results show the enhanced CK levels in mines, both on green and yellow leaves, allowing insects to control their nutritional supply under fluctuating environmental conditions. The spatial distribution of CKs within the mined leaves shows that hormone manipulation is strictly limited to the mine suggesting the absence of CK translocation from distant leaf areas toward the insect feeding site. Mass spectrometry analyses reveal that major CK types accumulating in mines and larvae are similar to what is observed for most gall-inducers, suggesting that strategies underlying the plant manipulation may be shared between herbivorous insects with distinct life histories. Results further show that CKs are detected in the highest levels in larvae, reinforcing our hypothesis that CKs accumulating in the mines originate from the insect itself. Presence of bacteriaspecific methylthio-CKs is consistent with previous results suggesting that insect bacterial symbionts contribute to the observed phenotype. Our study provides key findings toward the understanding of molecular mechanisms underlying this intricate plant-insect-microbe interaction.

Modulation of plant cytokinin levels in the Wolbachia‐free leaf‐mining species Phyllonorycter mespilella

“…However, significant amounts of phytohormones have been found in the saliva, body and accessory glands of some insect species suggesting that insects may also directly produce relevant levels of phytohormones or other effectors with mimicking functions Bartlett & Connor, 2014;Tooker & Helms, 2014;Zhang et al, 2017). Whether or not insects can be the source of CKs found at high levels at the insects feeding sitesrather than simply manipulating plant phytohormonal biosynthesis/signalingremains a challenging question to be answered in most systems.…”

“…However, significant amounts of phytohormones have been found in the saliva, body and accessory glands of some insect species suggesting that insects may also directly produce relevant levels of phytohormones or other effectors with mimicking functions Bartlett & Connor, 2014;Tooker & Helms, 2014;Zhang et al, 2017). CKs have been detected in the body, saliva or accessory glands of several other gallinducing (Ohkawa, 1974;Mapes & Davies, 2001b;Dorchin et al, 2009;Straka et al, 2010;Tooker & De Moraes, 2011a,b;Giron et al, 2013;Tanaka et al, 2013) and leaf-mining insect species (Engelbrecht et al, 1969;Engelbrecht, 1971;Body et al, 2013;Zhang et al, 2017), suggesting their ability to produce and deliver these effectors to the plant Bartlett & Connor, 2014;Tooker & Helms, 2014;Zhang et al, 2017). that induce galls on Salix japonica Thunb., high CK levels have been detected in larvae (Yamaguchi et al, 2012).…”

“…There is also growing evidence that insect-associated microbes are active players in the ability of insects to modulate the plant hormonal balance to the benefit of their insect host (Kaiser et al, 2010;Body et al, 2013;Sugio et al, 2014;Zhang et al, 2016Zhang et al, , 2017Giron et al, 2017). Metabolic profiling targeted to the bacterial chemical signatures (Zhang et al, 2017), and comparisons between insects deprived of their endosymbiotic bacteria through antibiotic treatments (Kaiser et al, 2010) Phytohormonal modulation in a Wolbachia-free leaf-miner 429 or between distinct populations that do or do not harbor endosymbionts (Barr et al, 2010) have provided key indirect evidence of the role of symbionts in the modulation of plant phytohormones. However, without labeling experiments, artificial rearing media, or knockout mutants it often remains difficult to distinguish conclusively between plant-, insect-, and symbiont-derived hormones.…”

“…These leaf areas are enriched in CKs, and CKs accumulated in the mined area do not originate from the plant CK-biosynthetic pathway (Zhang et al, 2016). Insect bacterial symbionts (Wolbachia) contribute to the observed phenotype (Giron et al, 2007;Kaiser et al, 2010;Body et al, 2013;Gutzwiller et al, 2015) likely synthesizing CKs delivered to the plant (Zhang et al, 2017), including bacteriaspecific methylthio-CKs (2-MeS-CKs). Insect bacterial symbionts (Wolbachia) contribute to the observed phenotype (Giron et al, 2007;Kaiser et al, 2010;Body et al, 2013;Gutzwiller et al, 2015) likely synthesizing CKs delivered to the plant (Zhang et al, 2017), including bacteriaspecific methylthio-CKs (2-MeS-CKs).…”

“…Whether or not insects can be the source of CKs found at high levels at the insects feeding sitesrather than simply manipulating plant phytohormonal biosynthesis/signalingremains a challenging question to be answered in most systems. However, significant amounts of phytohormones have been found in the saliva, body and accessory glands of some insect species suggesting that insects may also directly produce relevant levels of phytohormones or other effectors with mimicking functions Bartlett & Connor, 2014;Tooker & Helms, 2014;Zhang et al, 2017). In sawflies (Hymenoptera: Pontania spec.)…”