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The Sulfated PSY Peptide Negatively Regulates Receptor Kinase Activity to Promote Growth

Devin V. TulioiD, Alexandra M. ShigenagaiD, Dawn Lim, Artur Teixeira de AraújoiD, Shu‐Zon WuiD, Pamela C. RonaldiD, Magdalena BezanillaiD

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Abstract

Complex signaling pathways organize cell expansion and proliferation across cells to pattern tissues and organs in plants. The sulfotyrosine peptide hormone family, PLANT PEPTIDE CONTAINING SULFATED TYROSINE (PSY), contributes to these processes. We identified two plasma membrane-localized receptors, PSYR1 and PSYR2, that are necessary for PSY signaling and regulate growth in Physcomitrium patens. Membrane-associated PSYRs accumulate to high levels in a mutant lacking TYROSYL PROTEIN SULFOTRANSFERASE (TPST). Given that a tpst null mutant ({triangleup}tpst) is impaired in sulfation, this suggests that in the absence of sulfated peptides, PSYRs accumulate on the membrane. A null mutant of the PSY receptors,{triangleup} psyr1/2, showed increased growth and was epistatic to{triangleup} tpst, suppressing defects in gametophore formation and early senescence. The transcriptional profiles comparing wild type to{triangleup} psyr1/2 and{triangleup} psyr1/2/{triangleup}tpst showed 25 to 30 differentially expressed genes between the receptor null mutants and wild type, with a common signature of cell wall remodeling and stress responses. Similarly, a PSYR1 kinase-inactive mutation rescued{triangleup} tpst and relieved the accumulation of membrane-associated PSYRs. In contrast, overexpression of PSYRs inhibited plant growth, with phenotypic severity correlating with the amount of overexpression. These data are consistent with a constitutive activation model in which membrane-associated PSYRs unbound to PSY serve to inhibit growth through an active kinase. In the presence of the PSY peptide, the kinase is inactivated, promoting growth and driving PSY expression. The relationship between growth-repressive PSYR kinase activity and growth-promoting PSYR kinase inactivation in P. patens serves as a model for optimizing plant growth and development.

In-depth summary

Claude Opus 5.5 · xhigh effort · 8 Nov 2026 · from the full text of version 2 (2 Sept 2026) · CC BY 4.0

In Physcomitrium patens, genetic evidence suggests sulfated PSY peptide binding inactivates its receptor kinases rather than activating them; kinase-dead receptors rescue tpst mutants, while receptor overexpression phenocopies loss of sulfation.

Leucine-rich repeat receptor kinases are normally activated when a peptide binds, and Arabidopsis PSY receptors are described as active in the absence of ligand, but the underlying mechanism had not been tested in other species. Using endogenous tags and kinase point mutations in moss, the paper tests whether ligand binding turns the receptor kinase off.

Key results

  1. ShownTwo moss PSY receptors (PSYR1, PSYR2) are needed to respond to PSY peptide, and both accumulate at the plasma membrane when sulfation is lost, without higher transcript levels.

    Wild type formed on average 2.08 times more buds and gametophores on 1 µM PpPSY; Δpsyr1/2 showed no change. In Δtpst, PSYR1 and PSYR2 transcripts were 0.42 and 0.50 of wild type, yet mScarlet signal was higher. · Identification and characterization of PSY…

  2. ShownRemoving the receptors rescues Δtpst growth defects, and most of the Δtpst transcriptional response depends on the receptors.

    Plant area vs wild type: Δpsyr1/2 18% larger, Δpsyr1/2/Δtpst 16% larger, Δtpst 69.5% smaller. DEGs vs wild type: 271 (Δtpst), 25 (Δpsyr1/2), 30 (triple). 253 of 271 were receptor-dependent; r = -0.984 across 262 shared genes. · Loss of TPST in the psyr null mutant rescues…; Transcriptional landscape of tpst and psy…

  3. ShownA kinase-dead PSYR1 (K843E, with PSYR2 knocked out) fully rescues Δtpst, so Δtpst phenotypes need receptor kinase activity, not merely receptor presence. The kinase-dead receptor also does not accumulate on the membrane in Δtpst.

    Δtpst/psyr1-KD was similar in size to wild type. PSYR1-mScarlet signal in Δtpst/PSYR1-mScarlet was 2.4 times that of PSYR1-mScarlet and psyr1-KD, and 2.2 times that of Δtpst/psyr1-KD (p < 0.001). · Loss of PSYR kinase activity rescues gametophore…; Discussion

  4. SuggestedKinase-dead receptors strongly raise PSY expression, whereas receptor-null lines do not. Overexpressing PSYRs lowers PSY and mimics Δtpst, and this is reversed by kinase-dead mutations.

    PSY was up 14.1-fold (psyr1-KD) and 19.2-fold (Δtpst/psyr1-KD). Kinase-dead overexpression lines had PSY about 2.5 times wild type (p=0.011, 0.012) and about 37 times Δtpst (p<0.0001). Overexpression lines: two PSYR1, one PSYR2. · Loss of PSYR kinase activity rescues gametophore…; Overexpression of PSYRs phenocopy Δtpst

  5. SuggestedLoss of PSY receptors enlarges plants in moss and lengthens roots in Arabidopsis, but through different cell wall gene sets, which the authors read as a conserved growth-repressive role with diverged downstream targets.

    Moss receptor nulls down-regulated wall-stiffening genes; Arabidopsis tri-1 showed expansins and xyloglucan endotransglucosylases/hydrolases mostly up (16 up, 7 down). Only 1,104 of 2,237 moss DEGs (49%) had an assignable Arabidopsis ortholog. · Transcriptional landscape of tpst and psy…; Discussion; RNA-seq analysis

Methods, data and reproducibility

CRISPR-Cas9 and homology-directed repair in P. patens were used to make null, kinase-dead, endogenously tagged and overexpression lines of the PSY receptors and TPST. These lines were compared by morphometry, confocal imaging, RT-qPCR and RNA-seq (DESeq2).

  • P. patens lines: wild type, Δtpst, Δpsyr1/2, Δpsyr1/2/Δtpst, kinase-dead (psyr1-KD) and PSYR-overexpression lines; bulk RNA-seq (about 20 million paired-end reads per sample) · Physcomitrium patens · n = 3 biological replicates per genotype (RNA-seq)
  • Published Arabidopsis tri-1 (psyr1,2,3) vs Col-0 root DEG tables, used for cross-species comparison · Arabidopsis thaliana
Methods & tools
CRISPR-Cas9 with homology-directed repair, NCBI BlastP, Phytozome, Trimmomatic v.0.36, STAR v.2.5.2b, featureCounts v.1.5.2, DESeq2, Ensembl Plants BioMart, Fiji, R 4.4.1, RT-qPCR, Confocal microscopy (mScarlet, mStayGold)
Reagents & environment
Trimmomatic v.0.36, STAR v.2.5.2b, featureCounts v.1.5.2, DESeq2, R 4.4.1, RStudio v2023.12.1+402; reference genome Ppatens_318_v3.3_ERCC.
Reproducibility gaps
RNA-seq reads are said to be submitted to GEO, but no accession is printed. No analysis code is mentioned; oligos and line details are in supplementary tables.
Limitations (3)
  • Whether the TPST-dependent, receptor-independent transcriptional changes reflect sulfated substrates other than PSY remains to be determined. · stated by the authors
  • Only 49% of moss DEGs had an assignable Arabidopsis ortholog (mean 39% identity), which limits how far the cross-species comparison of downstream targets can go. · noted in this summary
  • The claim that ligand binding inactivates the kinase is inferred genetically (kinase-dead and null lines, expression and fluorescence); no direct kinase or phosphorylation assay with and without peptide is reported. · noted in this summary

Read this if you study receptor kinase signaling or sulfated peptide hormones and want genetic evidence that ligand binding switches an LRR receptor kinase off.

Citations
1

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