Elucidating the signaling mechanism of strigolactones has been the key to controlling the devastating problem caused by the parasitic plant Striga hermonthica. To overcome the genetic intractability that has previously interfered with identification of the strigolactone receptor, we developed a fluorescence turn-on probe, Yoshimulactone Green (YLG), which activates strigolactone signaling and illuminates signal perception by the strigolactone receptors. Here we describe how strigolactones bind to and act via ShHTLs, the diverged family of α/β hydrolase-fold proteins in Striga. Live imaging using YLGs revealed that a dynamic wavelike propagation of strigolactone perception wakes up Striga seeds. We conclude that ShHTLs function as the strigolactone receptors mediating seed germination in Striga. Our findings enable access to strigolactone receptors and observation of the regulatory dynamics for strigolactone signal transduction in Striga.

Multiple covalent modifications on a histone tail are often recognized by linked histone reader modules. UHRF1 [ubiquitin-like, containing plant homeodomain (PHD) and really interesting new gene (RING) finger domains 1], an essential factor for maintenance of DNA methylation, contains linked two-histone reader modules, a tandem Tudor domain and a PHD finger, tethered by a 17-aa linker, and has been implicated to link histone modifications and DNA methylation. Here, we present the crystal structure of the linked histone reader modules of UHRF1 in complex with the amino-terminal tail of histone H3. Our structural and biochemical data provide the basis for combinatorial readout of unmodified Arg-2 (H3-R2) and methylated Lys-9 (H3-K9) by the tandem tudor domain and the PHD finger. The structure reveals that the intermodule linker plays an essential role in the formation of a histone H3-binding hole between the reader modules by making extended contacts with the tandem tudor domain. The histone H3 tail fits into the hole by adopting a compact fold harboring a central helix, which allows both of the reader modules to simultaneously recognize the modification states at H3-R2 and H3-K9. Our data also suggest that phosphorylation of a linker residue can modulate the relative position of the reader modules, thereby altering the histone H3-binding mode. This finding implies that the linker region plays a role as a functional switch of UHRF1 involved in multiple regulatory pathways such as maintenance of DNA methylation and transcriptional repression.epigenetics | multidomain structure | posttranslational modification | X-ray crystallography

The circadian clock provides organisms with the ability to adapt to daily and seasonal cycles. Eukaryotic clocks mostly rely on lineage-specific transcriptional-translational feedback loops (TTFLs). Posttranslational modifications are also crucial for clock functions in fungi and animals, but the posttranslational modifications that affect the plant clock are less understood. Here, using chemical biology strategies, we show that the Arabidopsis CASEIN KINASE 1 LIKE (CKL) family is involved in posttranslational modification in the plant clock. Chemical screening demonstrated that an animal CDC7/CDK9 inhibitor, PHA767491, lengthens the Arabidopsis circadian period. Affinity proteomics using a chemical probe revealed that PHA767491 binds to and inhibits multiple CKL proteins, rather than CDC7/CDK9 homologs. Simultaneous knockdown of Arabidopsis CKL-encoding genes lengthened the circadian period. CKL4 phosphorylated transcriptional repressors PSEUDO-RESPONSE REGULATOR 5 (PRR5) and TIMING OF CAB EXPRESSION 1 (TOC1) in the TTFL. PHA767491 treatment resulted in accumulation of PRR5 and TOC1, accompanied by decreasing expression of PRR5- and TOC1-target genes. A prr5 toc1 double mutant was hyposensitive to PHA767491-induced period lengthening. Together, our results reveal posttranslational modification of transcriptional repressors in plant clock TTFL by CK1 family proteins, which also modulate nonplant circadian clocks.

The parasitic plantStriga hermonthicahas been causing devastating damage to the crop production in Africa. BecauseStrigarequires host-generated strigolactones to germinate, the identification of selective and potent strigolactone agonists could help control these noxious weeds. We developed a selective agonist, sphynolactone-7, a hybrid molecule originated from chemical screening, that contains two functional modules derived from a synthetic scaffold and a core component of strigolactones. Cooperative action of these modules in the activation of a high-affinity strigolactone receptor ShHTL7 allows sphynolactone-7 to provokeStrigagermination with potency in the femtomolar range. We demonstrate that sphynolactone-7 is effective for reducingStrigaparasitism without impinging on host strigolactone-related processes.

Highlights d Keap1 H 2 O 2 sensor is distinct from that used for sensing electrophilic inducers d Keap1 uses Cys226, Cys613, and Cys622/624 residues to sense H 2 O 2 d Keap1 uses these cysteine residues to set up an elaborate fail-safe mechanism

Arginine-rich cell-penetrating peptides (CPPs) such as Tat and oligoarginine peptides have been widely used as carriers for intracellular delivery of bioactive molecules. Despite accumulating evidence for involvement of endocytosis in the cellular uptake of arginine-rich CPPs, the primary cell-surface receptors for these peptide carriers that would initiate endocytic processes leading to intracellular delivery of bioactive cargoes have remained poorly understood. Our previous attempt to identify membrane receptors for octa-arginine (R8) peptide, one of the representative arginine-rich CPPs, using the photo-cross-linking probe bearing a photoreactive diazirine was not successful due to considerable amounts of cellular proteins nonspecifically bound to the affinity beads. To address this issue, here we developed a photo-cross-linking probe in which a cleavable linker of a diazobenzene moiety was employed to allow selective elution of cross-linked proteins by reducing agent-mediated cleavage. We demonstrated that introduction of the diazobenzene moiety into the photoaffinity probe enables efficient purification of cross-linked proteins with significant reduction of nonspecific binding proteins, leading to successful identification of 17 membrane-associated proteins that would interact with R8 peptide. RNAi-mediated knockdown experiments in combination with the pharmacological inhibitors revealed that, among the proteins identified, syndecan-4, one of the heparan sulfate proteoglycans, is an endogenous membrane-associated receptor for the cellular uptake of R8 peptide via clathrin-mediated endocytosis. This syndecan-4-dependent pathway was also involved in the intracellular delivery of bioactive proteins mediated by R8 peptide. These results reveal that syndecan-4 is a primary cell-surface target for R8 peptide that allows intracellular delivery of bioactive cargo molecules via clathrin-mediated endocytosis.

No abstract

Precise directional control of pollen-tube growth by pistil tissue is critical for successful fertilization of flowering plants [1-3]. Ovular attractant peptides, which are secreted from two synergid cells on the side of the egg cell, have been identified [4-6]. Emerging evidence suggests that the ovular directional cue is not sufficient for successful guidance but that competency control by the pistil is critical for the response of pollen tubes to the attraction signal [1, 3, 7]. However, the female molecule for this competency induction has not been reported. Here we report that ovular methyl-glucuronosyl arabinogalactan (AMOR) induces competency of the pollen tube to respond to ovular attractant LURE peptides in Torenia fournieri. We developed a method for assaying the response capability of a pollen tube by micromanipulating an ovule. Using this method, we showed that pollen tubes growing through a cut style acquired a response capability in the medium by receiving a sufficient amount of a factor derived from mature ovules of Torenia. This factor, named AMOR, was identified as an arabinogalactan polysaccharide, the terminal 4-O-methyl-glucuronosyl residue of which was necessary for its activity. Moreover, a chemically synthesized disaccharide, the β isomer of methyl-glucuronosyl galactose (4-Me-GlcA-β-(1→6)-Gal), showed AMOR activity. No specific sugar-chain structure of plant extracellular matrix has been identified as a bioactive molecule involved in intercellular communication. We suggest that the AMOR sugar chain in the ovary renders the pollen tube competent to the chemotropic response prior to final guidance by LURE peptides.