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Abstract Many root parasitic plants in the Orobanchaceae use host-derived strigolactones (SLs) as germination cues. This adaptation facilitates attachment to a host and is particularly important for the success of obligate parasitic weeds that cause substantial crop losses globally. Parasite seeds sense SLs through ‘divergent’ KARRIKIN INSENSITIVE2 (KAI2d)/HYPOSENSITIVE TO LIGHT α/β-hydrolases that have undergone substantial duplication and diversification in Orobanchaceae genomes. After germination, chemotropic growth of parasite roots toward a SL source also occurs in some species. We investigated which of the seven KAI2d genes found in a facultative hemiparasite, Phtheirospermum japonicum, may enable chemotropic responses to SLs. To do so, we developed a triple mutant Nbd14a,b kai2i line of Nicotiana benthamiana in which SL-induced degradation of SUPPRESSOR OF MORE AXILLARY GROWTH2 (MAX2) 1 (SMAX1), an immediate downstream target of KAI2 signaling, is disrupted. In combination with a transiently expressed, ratiometric reporter of SMAX1 protein abundance, this mutant forms a system for the functional analysis of parasite KAI2d proteins in a plant cellular context. Using this system, we unexpectedly found three PjKAI2d proteins that do not trigger SMAX1 degradation in the presence of SLs. Instead, these PjKAI2d proteins inhibit the perception of low SL concentrations by SL-responsive PjKAI2d in a dominant-negative manner that depends upon an active catalytic triad. Similar dominant-negative KAI2d paralogs were identified in an obligate hemiparasitic weed, Striga hermonthica. These proteins suggest a mechanism for attenuating SL signaling in parasites, which might be used to enhance the perception of shallow SL gradients during root growth toward a host or to restrict germination responses to specific SLs.
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Rapid analysis of strigolactone receptor activity in a Nicotiana benthamiana dwarf14 mutant
Abstract DWARF14 (D14) is an ɑ/β‐hydrolase and receptor for the plant hormone strigolactone (SL) in angiosperms. Upon SL perception, D14 works with MORE AXILLARY GROWTH2 (MAX2) to trigger polyubiquitination and degradation of DWARF53(D53)‐type proteins in the SUPPRESSOR OF MAX2 1‐LIKE (SMXL) family. We used CRISPR‐Cas9 to generate knockout alleles of the two homoeologousD14genes in theNicotiana benthamianagenome. TheNbd14a,bdouble mutant had several phenotypes that are consistent with the loss of SL perception in other plants, including increased axillary bud outgrowth, reduced height, shortened petioles, and smaller leaves. A ratiometric fluorescent reporter system was used to monitor degradation of SMXL7 fromArabidopsis thaliana(AtSMXL7) after transient expression inN. benthamianaand treatment with the strigolactone analog GR24. AtSMXL7 was degraded after treatment with GR245DS, which has the stereochemical configuration of natural SLs, as well as its enantiomer GR24ent‐5DS. InNbd14a,bleaves, AtSMXL7 abundance was unaffected byrac‐GR24 or either GR24 stereoisomer. Transient coexpression of AtD14 with the AtSMXL7 reporter inNbd14a,brestored the degradation response torac‐GR24, but required an active catalytic triad. We used this platform to evaluate the ability of several AtD14 mutants that had not been characterized in plants to target AtSMXL7 for degradation.
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- PAR ID:
- 10364578
- Publisher / Repository:
- Wiley Blackwell (John Wiley & Sons)
- Date Published:
- Journal Name:
- Plant Direct
- Volume:
- 6
- Issue:
- 3
- ISSN:
- 2475-4455
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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