Cytokinins Induce Prehaustoria Coordinately with Quinone Signals in the Parasitic Plant Striga hermonthica

Abstract

Orobanchaceae parasitic plants are major threats to global food security, causing severe agricultural damage worldwide. Parasitic plants derive water and nutrients from their host plants through multicellular organs called haustoria. The formation of a prehaustorium, a primitive haustorial structure, is provoked by host-derived haustorium-inducing factors (HIFs). Quinones, including 2,6-dimethoxy-p-benzoquinone (DMBQ) are of the most potent HIFs for various species in Orobanchaceae, but except non-photosynthetic holoparasites, Phelipanche and Orobanche spp. Instead, cytokinin phytohormones were reported to induce prehaustoria in Phelipanche ramosa. However, little is known about whether cytokinins act as HIFs in the other parasitic species to date. Moreover, the signaling pathways for quinones and cytokinins in prehaustorium induction are not well understood. This study shows that cytokinins act as HIFs in the obligate parasite Striga hermonthica but not in the facultative parasite Phtheirospermum japonicum. Using chemical inhibitors and marker gene expression analysis, we demonstrate that cytokinins activate prehaustorium formation through cytokinin-specific signaling pathway that overlaps with the quinone HIF pathway at downstream in S. hermonthica. Moreover, host root exudates activated S. hermonthica cytokinin biosynthesis and signaling genes, and DMBQ and cytokinin inhibitors perturbed the prehaustorium-inducing activity of exudates, indicating that host root exudates include cytokinins. Our study reveals the importance of cytokinins for prehaustorium formation in obligate parasitic plants.

Phenolic signals for prehaustorium formation in Striga hermonthica

“…This was clearly supported by the inhibitory effect of ASBr and TFBQ on all of these HIFs (Figure 3) (Aoki et al, 2022). On the other hand, we found that LGR-991, a competitive inhibitor of CK that directly binds CK receptors, inhibited CKs, AS and SyA but not DMBQ (Figure 3) (Aoki et al, 2022). These results remarkably demonstrate the independence of quinone signaling transduction from CK perception and indicate that phenolic signaling is integrated into CK perception.…”

“…No detectable variation was found in the morphology of prehaustoria induced by phenolics, quinones or CKs, implying that their signaling must converge toward the same physiological output, at least in S. hermonthica. This was clearly supported by the inhibitory effect of ASBr and TFBQ on all of these HIFs (Figure 3) (Aoki et al, 2022). On the other hand, we found that LGR-991, a competitive inhibitor of CK that directly binds CK receptors, inhibited CKs, AS and SyA but not DMBQ (Figure 3) (Aoki et al, 2022).…”

“…The plant materials and growth conditions used were previously reported in Aoki et al (2022). Briefly, S. hermonthica seeds of approximately 50-100 mg were sterilized with 20% commercial bleach solution (Kao Ltd., Tokyo, Japan; final concentration of sodium hypochlorite was approximately 0.6%) for 5 min, washed at least 5 times with sterilized water, poured into petri dishes with moist glass fiber filter paper (GE Healthcare UK Ltd., Little Chalfont, UK), and preconditioned for 1 week at 25°C in the dark.…”

“…Koshihikari and A. thaliana ecotype Columbia (Col-0) were used for the exudate experiment. Root exudates of rice and Arabidopsis were collected as previously reported (Aoki et al, 2022) Inhibitor assay for prehaustorium induction Germinated S. hermonthica seedlings were transferred to 96well plates (Iwaki, Shizuoka, Japan) with each well containing HIFs with or without chemical inhibitors in a total volume of 100 µl. Chemical stocks of HIFs and inhibitors were initially prepared using DMSO as the solvent and added to each well according to the final working concentrations.…”

“…Despite differences in the haustorium initiation sites and their morphology, both facultative and obligate parasites can respond to the same HIFs, i.e., facultative parasite Phtheirospermum japonicum and the obligate parasite S. hermonthica initiate prehaustorium formation by a similar set of monolignols and quinones that bear certain structural properties (Cui et al, 2018). Diverged HIF recognition also exists among three parasitic forms: S. hermonthica but not P. japonicum reacts to cytokinins (CKs) as HIFs (Aoki et al, 2022), whereas the holoparasite Phelipanche ramosa responds primarily to CKs (Goyet et al, 2017), and forms prehaustoria against only high concentration of quinones at millimolar range (Fernandez-Aparicio et al, 2021), reflecting reduced ability of holoparasites in sensing phenolics and quinones.…”