Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

TOP1α fine-tunes TOR-PLT2 to maintain root tip homeostasis in response to sugars

A Publisher Correction to this article was published on 08 September 2022

This article has been updated

Abstract

Plant development is highly dependent on energy levels. TARGET OF RAPAMYCIN (TOR) activates the proximal root meristem to promote root development in response to photosynthesis-derived sugars during photomorphogenesis in Arabidopsis thaliana. However, the mechanisms of how root tip homeostasis is maintained to ensure proper root cap structure and gravitropism are unknown. PLETHORA (PLT) transcription factors are pivotal for the root apical meristem (RAM) identity by forming gradients, but how PLT gradients are established and maintained, and their roles in COL development are not well known. We demonstrate that endogenous sucrose induces TOPOISOMERASE1α (TOP1α) expression during the skotomorphogenesis-to-photomorphogenesis transition. TOP1α fine-tunes TOR expression in the root tip columella. TOR maintains columella stem cell identity correlating with reduced quiescent centre cell division in a WUSCHEL RELATED HOMEOBOX5-independent manner. Meanwhile, TOR promotes PLT2 expression and phosphorylates and stabilizes PLT2 to maintain its gradient consistent with TOR expression pattern. PLT2 controls cell division and amyloplast formation to regulate columella development and gravitropism. This elaborate mechanism helps maintain root tip homeostasis and gravitropism in response to energy changes during root development.

This is a preview of subscription content, access via your institution

Access options

Buy article

Get time limited or full article access on ReadCube.

$32.00

All prices are NET prices.

Fig. 1: TOP1α represses TOR expression.
Fig. 2: TOR controls cell division and PLT2 accumulation.
Fig. 3: TOR phosphorylates and stabilizes PLT2.

Data availability

The RNA-seq raw data have been deposited in the NCBI SRA (https://www.ncbi.nlm.nih.gov/sra/) under the accession number: PRJNA700788. Sequence data for the genes in this article can be found in the Arabidopsis Genome Initiative or GenBank/EMBL databases under the following accession numbers: CDC6, AT2G29680; CDF4, AT2G34140; CDKB1, AT3G54180; CLE40, AT5G12990; CYCB1;1, AT4G37490; CYCD3;3, AT3G50070; CYCD5;1, AT4G37630; E2Fa, AT2G36010; ETG1, AT2G40550; GIN2, AT4G29130; KIN10, AT3G01090; KIN11, AT3G29160; MCM5, AT2G07690; MCM7, AT4G02060; PLT1, AT3G20840; PLT2, AT1G51190; SCR, AT3G54220; SHR, AT4G37650; SS1, AT5G24300; SS2, AT3G01180; SS3, AT1G11720; SS4, AT4G18240; TOP1α, AT5G55300; TOR, AT1G50030; UBQ5, AT3G62250; WOX5, AT3G11260. Source data are provided with this paper.

Change history

References

  1. Moubayidin, L. et al. The rate of cell differentiation controls the Arabidopsis root meristem growth phase. Curr. Biol. 20, 1138–1143 (2010).

    CAS  PubMed  Google Scholar 

  2. Perilli, S. & Sabatini, S. Analysis of root meristem size development. Methods Mol. Biol. 655, 177–187 (2010).

    CAS  PubMed  Google Scholar 

  3. Xiong, Y. et al. Glucose-TOR signalling reprograms the transcriptome and activates meristems. Nature 496, 181–186 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  4. Li, X. et al. Differential TOR activation and cell proliferation in Arabidopsis root and shoot apexes. Proc. Natl Acad. Sci. USA 114, 2765–2770 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  5. Kircher, S. & Schopfer, P. Photosynthetic sucrose acts as cotyledon-derived long-distance signal to control root growth during early seedling development in Arabidopsis. Proc. Natl Acad. Sci. USA 109, 11217–11221 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  6. Pfeiffer, A. et al. Integration of light and metabolic signals for stem cell activation at the shoot apical meristem. eLife https://doi.org/10.7554/eLife.17023 (2016).

    Article  PubMed  PubMed Central  Google Scholar 

  7. Brunkard, J. O. Exaptive evolution of Target of Rapamycin signaling in multicellular eukaryotes. Dev. Cell 54, 142–155 (2020).

    CAS  PubMed  PubMed Central  Google Scholar 

  8. Liu, G. Y. & Sabatini, D. M. mTOR at the nexus of nutrition, growth, ageing and disease. Nat. Rev. Mol. Cell Biol. 21, 183–203 (2020).

    CAS  PubMed  PubMed Central  Google Scholar 

  9. Ryabova, L. A., Robaglia, C. & Meyer, C. Target of Rapamycin kinase: central regulatory hub for plant growth and metabolism. J. Exp. Bot. 70, 2211–2216 (2019).

    CAS  PubMed  PubMed Central  Google Scholar 

  10. Wu, Y. et al. Integration of nutrient, energy, light, and hormone signalling via TOR in plants. J. Exp. Bot. 70, 2227–2238 (2019).

    CAS  PubMed  PubMed Central  Google Scholar 

  11. Fu, L. et al. The TOR-EIN2 axis mediates nuclear signalling to modulate plant growth. Nature 591, 288–292 (2021).

    CAS  PubMed  Google Scholar 

  12. Shi, L., Wu, Y. & Sheen, J. TOR signaling in plants: conservation and innovation. Development https://doi.org/10.1242/dev.160887 (2018)

  13. Menand, B. et al. Expression and disruption of the Arabidopsis TOR (target of rapamycin) gene. Proc. Natl Acad. Sci. USA 99, 6422–6427 (2002).

    CAS  PubMed  PubMed Central  Google Scholar 

  14. Van Leene, J. et al. Capturing the phosphorylation and protein interaction landscape of the plant TOR kinase. Nat. Plants 5, 316–327 (2019).

    PubMed  Google Scholar 

  15. Lee, Y., Lee, W. S. & Kim, S. H. Hormonal regulation of stem cell maintenance in roots. J. Exp. Bot. 64, 1153–1165 (2013).

    CAS  PubMed  Google Scholar 

  16. Liu, X. et al. DNA topoisomerase I affects polycomb group protein-mediated epigenetic regulation and plant development by altering nucleosome distribution in Arabidopsis. Plant Cell 26, 2803–2817 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  17. Liu, X. G. et al. AGAMOUS terminates floral stem cell maintenance in Arabidopsis by directly repressing WUSCHEL through recruitment of polycomb group proteins. Plant Cell 23, 3654–3670 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  18. Capranico, G., Marinello, J. & Chillemi, G. Type I DNA topoisomerases. J. Med. Chem. 60, 2169–2192 (2017).

    CAS  PubMed  Google Scholar 

  19. Wang, J. C. Cellular roles of DNA topoisomerases: a molecular perspective. Nat. Rev. Mol. Cell Biol. 3, 430–440 (2002).

    CAS  PubMed  Google Scholar 

  20. Dinh, T. T. et al. DNA topoisomerase 1alpha promotes transcriptional silencing of transposable elements through DNA methylation and histone lysine 9 dimethylation in Arabidopsis. PLoS Genet. 10, e1004446 (2014).

    PubMed  PubMed Central  Google Scholar 

  21. Takahashi, T., Matsuhara, S., Abe, M. & Komeda, Y. Disruption of a DNA topoisomerase I gene affects morphogenesis in Arabidopsis. Plant Cell 14, 2085–2093 (2002).

    CAS  PubMed  PubMed Central  Google Scholar 

  22. Graf, P. et al. MGOUN1 encodes an Arabidopsis type IB DNA topoisomerase required in stem cell regulation and to maintain developmentally regulated gene silencing. Plant Cell Online 22, 716–728 (2010).

    CAS  Google Scholar 

  23. Shafiq, S. et al. DNA topoisomerase 1 prevents R-loop accumulation to modulate auxin-regulated root development in rice. Mol. Plant 10, 821–833 (2017).

    CAS  PubMed  Google Scholar 

  24. Zhang, Y. et al. TOPOISOMERASE1alpha acts through two distinct mechanisms to regulate stele and columella stem cell maintenance. Plant Physiol. 171, 483–493 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  25. Fisher, A. P. & Sozzani, R. Uncovering the networks involved in stem cell maintenance and asymmetric cell division in the Arabidopsis root. Curr. Opin. Plant Biol. 29, 38–43 (2016).

    CAS  PubMed  Google Scholar 

  26. Forzani, C. et al. WOX5 suppresses CYCLIN D activity to establish quiescence at the center of the root stem cell niche. Curr. Biol. 24, 1939–1944 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  27. Zhang, W., Swarup, R., Bennett, M., Schaller, G. E. & Kieber, J. J. Cytokinin induces cell division in the quiescent center of the Arabidopsis root apical meristem. Curr. Biol. 23, 1979–1989 (2013).

    CAS  PubMed  Google Scholar 

  28. Sheen, J. Master regulators in plant glucose signaling networks. J. Plant Biol. 57, 67–79 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  29. Montane, M. H. & Menand, B. ATP-competitive mTOR kinase inhibitors delay plant growth by triggering early differentiation of meristematic cells but no developmental patterning change. J. Exp. Bot. 64, 4361–4374 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  30. Kunz, S., Pesquet, E. & Kleczkowski, L. A. Functional dissection of sugar signals affecting gene expression in Arabidopsis thaliana. PLoS One 9, e100312 (2014).

    PubMed  PubMed Central  Google Scholar 

  31. De Veylder, L., Joubes, J. & Inze, D. Plant cell cycle transitions. Curr. Opin. Plant Biol. 6, 536–543 (2003).

    PubMed  Google Scholar 

  32. Delgado, J. L., Hsieh, C. M., Chan, N. L. & Hiasa, H. Topoisomerases as anticancer targets. Biochem J. 475, 373–398 (2018).

    CAS  PubMed  Google Scholar 

  33. Drisch, R. C. & Stahl, Y. Function and regulation of transcription factors involved in root apical meristem and stem cell maintenance. Front. Plant Sci. 6, 505 (2015).

    PubMed  PubMed Central  Google Scholar 

  34. Aida, M. et al. The PLETHORA genes mediate patterning of the Arabidopsis root stem cell niche. Cell 119, 109–120 (2004).

    CAS  PubMed  Google Scholar 

  35. Galinha, C. et al. PLETHORA proteins as dose-dependent master regulators of Arabidopsis root development. Nature 449, 1053–1057 (2007).

    CAS  PubMed  Google Scholar 

  36. Smith, Z. R. & Long, J. A. Control of Arabidopsis apical-basal embryo polarity by antagonistic transcription factors. Nature 464, 423–426 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  37. Blilou, I. et al. The PIN auxin efflux facilitator network controls growth and patterning in Arabidopsis roots. Nature 433, 39–44 (2005).

    CAS  PubMed  Google Scholar 

  38. Xu, J. et al. A molecular framework for plant regeneration. Science 311, 385–388 (2006).

    CAS  PubMed  Google Scholar 

  39. Mähönen, A. P. et al. PLETHORA gradient formation mechanism separates auxin responses. Nature 515, 125–129 (2014).

    PubMed  PubMed Central  Google Scholar 

  40. Di Laurenzio, L. et al. The SCARECROW gene regulates an asymmetric cell division that is essential for generating the radial organization of the Arabidopsis root. Cell 86, 423–433 (1996).

    CAS  PubMed  Google Scholar 

  41. Helariutta, Y. et al. The SHORT-ROOT gene controls radial patterning of the Arabidopsis root through radial signaling. Cell 101, 555–567 (2000).

    CAS  PubMed  Google Scholar 

  42. Santuari, L. et al. The PLETHORA gene regulatory network guides growth and cell differentiation in Arabidopsis roots. Plant Cell 28, 2937–2951 (2016).

    PubMed  PubMed Central  Google Scholar 

  43. Yamada, M., Han, X. & Benfey, P. N. RGF1 controls root meristem size through ROS signalling. Nature 577, 85–88 (2020).

    CAS  PubMed  Google Scholar 

  44. Forzani, C. et al. Mutations of the AtYAK1 kinase suppress TOR deficiency in Arabidopsis. Cell Rep. 27, 3696–3708.e3695 (2019).

    CAS  PubMed  Google Scholar 

  45. Barrada, A. et al. A TOR-YAK1 signaling axis controls cell cycle, meristem activity and plant growth in Arabidopsis. Development 146, dev171298 (2019).

  46. Watson, M. J. et al. Metabolic support of tumour-infiltrating regulatory T cells by lactic acid. Nature 591, 645–651 (2021).

    CAS  PubMed  PubMed Central  Google Scholar 

  47. Grieneisen, V. A., Xu, J., Marée, A. F., Hogeweg, P. & Scheres, B. Auxin transport is sufficient to generate a maximum and gradient guiding root growth. Nature 449, 1008–1013 (2007).

    CAS  PubMed  Google Scholar 

  48. Yuan, X., Xu, P., Yu, Y. & Xiong, Y. Glucose-TOR signaling regulates PIN2 stability to orchestrate auxin gradient and cell expansion in Arabidopsis root. Proc. Natl Acad. Sci. USA 117, 32223–32225 (2020).

    CAS  PubMed  PubMed Central  Google Scholar 

  49. Di Mambro, R. et al. The lateral root cap acts as an auxin sink that controls meristem size. Curr. Biol. 29, 1199–1205 e1194 (2019).

    PubMed  Google Scholar 

  50. Matsuzaki, Y., Ogawa-Ohnishi, M., Mori, A. & Matsubayashi, Y. Secreted peptide signals required for maintenance of root stem cell niche in Arabidopsis. Science 329, 1065–1067 (2010).

    CAS  PubMed  Google Scholar 

  51. Pi, L. et al. Organizer-derived WOX5 signal maintains root columella stem cells through chromatin-mediated repression of CDF4 expression. Dev. Cell 33, 576–588 (2015).

    CAS  PubMed  Google Scholar 

  52. Moore, B. et al. Role of the Arabidopsis glucose sensor HXK1 in nutrient, light, and hormonal signaling. Science 300, 332–336 (2003).

    CAS  PubMed  Google Scholar 

  53. Tu, B. et al. Distinct and cooperative activities of HESO1 and URT1 nucleotidyl transferases in microRNA turnover in Arabidopsis. PLoS Genet. 11, e1005119 (2015).

    PubMed  PubMed Central  Google Scholar 

  54. Earley, K. W. et al. Gateway-compatible vectors for plant functional genomics and proteomics. Plant J. 45, 616–629 (2006).

    CAS  PubMed  Google Scholar 

  55. Wang, Z. P. et al. Egg cell-specific promoter-controlled CRISPR/Cas9 efficiently generates homozygous mutants for multiple target genes in Arabidopsis in a single generation. Genome Biol. 16, 144 (2015).

    PubMed  PubMed Central  Google Scholar 

  56. Zhou, W. et al. A jasmonate signaling network activates root stem cells and promotes regeneration. Cell 177, 942–956.e14 (2019).

  57. Grabov, A. et al. Morphometric analysis of root shape. New Phytol. 165, 641–651 (2005).

    CAS  PubMed  Google Scholar 

  58. Kim, D. et al. TopHat2: accurate alignment of transcriptomes in the presence of insertions, deletions and gene fusions. Genome Biol. 14, R36 (2013).

    PubMed  PubMed Central  Google Scholar 

  59. Anders, S., Pyl, P. T. & Huber, W. HTSeq–A Python framework to work with high-throughput sequencing data. Bioinformatics 31, 166–169 (2015).

    CAS  PubMed  Google Scholar 

  60. Robinson, M. D., McCarthy, D. J. & Smyth, G. K. edgeR: a Bioconductor package for differential expression analysis of digital gene expression data. Bioinformatics 26, 139–140 (2010).

    CAS  PubMed  Google Scholar 

  61. Yu, G., Wang, L. G., Han, Y. & He, Q. Y. clusterProfiler: an R package for comparing biological themes among gene clusters. OMICS 16, 284–287 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

We thank X. Chen and Y. Xiong for helpful discussion, Y. Xiong for sharing the e2fa and tor-es mutants and the TOR constructs for Y2H, L. Pi for the CSC marker J2341, B. Menand for the AtTOR::GUS line, C. Li for the pPLT2::CFP line and X. Huang and Y. Wang for the technical support on IP–MS. This research was supported by the project 'Full-time introduction of high-end talent research project' (2020HBQZYC004; A202105008 to X.L.) from Hebei province, funding from Hebei Natural Science Foundation (grant nos C2021205013 to X.L. and C2021205043 to L.G.) and grants from NSFC (grant nos 31970824 to X.L. and 31401039 to L.G.).

Author information

Authors and Affiliations

Authors

Contributions

J.D., L.G. and X.L. designed the experiments. H.Z., Y.L., D.Z., L.L., W.C., Y.N., Y.M., Y. Zheng, Y. Zhang and Ke Z. carried out most of the experiments with technical support from F.W., J.X., Kang Z. and S.C. H.Z., B.Z., S.Z. and W.T. performed the IP–MS experiment. J.H., K.T. and C.F. performed the CPT treatment NSCLC cell assay. X.L., S.Z. and W.T. analysed data. H.Z., Y.L., L.G. and X.L. wrote the manuscript with input from J.D.

Corresponding authors

Correspondence to Lin Guo, Jingao Dong or Xigang Liu.

Ethics declarations

Competing interests

The authors declare no competing interests.

Peer review

Peer review information

Nature Plants thanks José Manuel Pérez-Pérez, Benoit Menand and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Extended data

Extended Data Fig. 1 Maintenance of root tip homeostasis.

a, Cartoon depicting a longitudinal section of a wild type (WT) Arabidopsis root tip. QC: quiescent center; CSC: columella stem cell; PM: proximal root meristem; DM: distal root meristem; COL: columella; S1-S4: columella layers 1–4. b, Morphological observation of seedlings grown under the indicated treatments. Seedlings were grown on sugar-free medium for 5 days in the dark, transferred to different conditions, and grown for 3 more days. Scale bar: 5 mm. c, Examination of cell division in the RAMs of pCYCB1;1::GUS plants under the indicated treatments. Scale bar: 100 µm. d, f, Quantification of PM and COL size in WT plants grown under the indicated conditions. Data were analyzed from 15 seedlings for each treatment from 3 experiments. Statistical significance was determined by one-way ANOVA and Tukey test (P < 0.05) (see Supplementary Data 1 for specific P values) (d) or by Student’s t-test (two-tailed) (f) and are expressed as mean ± s.d. e, DM and CC homeostasis is maintained during the skotomorphogenesis-to-photomorphogenesis transition. The seedlings described in (b) harboring the QC marker pWOX5::GFP were selected for structural observation of the root tip. Arrows mark the CSC layer; lines mark the columella story (S). Scale bar: 50 µm.

Extended Data Fig. 2 TOP1α controls root gravitropism in response to sucrose.

a, b, Root gravitropism (a) and statistical analysis of vertical growth index (VGI) (b) in roots of the indicated genotypes under ±Suc treatment. Data were analyzed from 15 seedlings for each treatment from 3 experiments. Statistical significance was determined by Student’s t-test (two-tailed) (b) and are expressed as mean ± s.d. Scale bar: 1 cm. c, Graphical representation of VGI to define root gravitropism. d, Time course of the gravity response and amyloplast formation in WT and top1α ± Suc treatment. Seedlings were turned 90° from the vertical axis and fixed at the indicated time points. Amyloplast formation and levels in layers S1 and S2 were examined. Yellow lines represent amyloplast levels. Green arrows mark amyloplasts in premature CSCs. (m/n) indicates m in n of biological repeats showing the displayed features. Scale bar: 25 µm. e, f, Root gravity response (e) and statistical analysis (f) of WT and top1α in response to Suc. Data represent mean ± s.d. of 3 biological repeats. Statistical significance was determined by Student’s t-test (two-tailed) (f) and are expressed as mean ± s.d. Scale bar: 0.5 mm. (m/n) in (d and e) indicates m in n of biological repeats showing the displayed features.

Extended Data Fig. 3 TOP1α controls root tip homeostasis in response to sucrose.

a-c, Quantification of QC cell number (a), CSC layer (b), and CC number (c) of WT and top1α ± Suc treatment. Data represent mean ± s.d. of 3 biological repeats (a, b). Data were analyzed from 15 seedlings for each treatment from 3 experiments (c). Statistical significance was determined by one-way ANOVA with Tukey’s post hoc test (P < 0.05). d, RT-qPCR of CLE40 and CDF4 expression in WT and top1α under the indicated treatments. Gene expression in WT –Suc was set to 1. Data represent mean ± s.d. of 3 biological repeats. Student’s t test (two-tailed). e, Glucose contents in Ler and top1a. Statistical significance was determined by one-way ANOVA and Tukey test (P < 0.05). Data represent mean ±s.d. of 3 biological repeats (n = 3). Each replicate (n) is from 500–550 root tips. f. TOP1α expression patterns in pTOP1a::GFP root tips. Scale bar: 50 µm. g. TOP1α expression in pTOP1a::GFP root under indicated treatments. (m/n) indicates m in n of biological repeats showing the displayed features. Scale bar: 100 μm.

Extended Data Fig. 4 The role of TOP1α is independent of Glc and energy sensors.

a, Genetic analysis of the roles of GIN2 and TOP1α in root gravitropism. Scale bars: 1 cm. b, RT-qPCR of GIN2 expression in WT and top1α under the indicated treatments. Gene expression in WT –Suc was set to 1. Data represent mean ± s.d. of 3 biological repeats. Student’s t test (two-tailed).

Extended Data Fig. 5 Transcriptome analysis of TOP1α and TOR regulated genes.

a, Venn diagram showing the number of overlapping TOR-activated and TOR-repressed as well as Glc-TOR-activated and Glc-TOR repressed DEGs identified by Xiong et al., (2013). b, Venn diagram showing the number of overlapping DEGs in top1α vs. Ler and AZD-8055-treated Ler vs. Ler (left); and overlapping DEGs in top1α vs. Ler and AZD-8055-treated top1α vs. top1α (right). c, GO analysis of TOR-activated genes (upper panel) and TOR-repressed genes (lower panel). d, GO analysis of TOP1α-activated genes (left) and TOP1α-repressed genes (right). e, f, Hierarchical clustering analysis of TOR-activated (e) and TOR-repressed genes (f). TOR-activated and TOP1α-repressed DEGs (e) and TOR-repressed and TOP1α-activated DEGs (f) were grouped and subjected to GO analysis, as shown in the graphs. g, h, Hierarchical clustering analysis of TOP1α-activated (g) genes and TOP1α-repressed genes (h). TOP1α-activated and TOR-repressed DEGs (g) and TOP1α-repressed and TOR-activated DEGs (h) were grouped and subjected to GO analysis, as shown in the graphs. The GO enrichment analysis was performed by R package ClusterProfiler3.0, in which the pvalue was calculated by Fisher’ exact test, and was adjusted by Benjamini-Hochbery method in c-h.

Extended Data Fig. 6 TOP1a regulates cell cycle gene expression.

a, Heatmap showing the expression of cell-cycle-related genes in WT and top1α ± AZD-8055 treatment. b, RT-qPCR of CYCD3;3 and E2Fa as well as E2Fa target gene expression in WT and top1α under the indicated treatments. Gene expression in WT –Suc was set to 1. Data represent mean ± s.d. of 3 biological repeats. Statistical significance was determined by one-way ANOVA and Tukey test (P < 0.05) (see Supplementary Data 1 for specific P values). c, Diagram of the CYCD3;3 locus. Gray and black rectangles and black lines represent untranslated regions, coding regions, and introns, respectively. Two nucleotides (GA) were deleted, resulting in a premature stop codon (red), in the cycd3;3 mutant. d, e, Quantification of CSC layer (d) and CC number (e) of the indicated genotypes. Data represent mean ± s.d. of 3 biological repeats (d). Data were analyzed from 15 seedlings for each genotype from 3 experiments (e). Statistical significance was determined by one-way ANOVA and Tukey test (P < 0.05) (see Supplementary Data 1 for specific P values). f, Root gravity responses and amyloplast formation in the indicated genotypes. Seedlings were turned 90° from the vertical axis and fixed at the indicated time points. Amyloplast formation and levels in layers S1 and S2 were examined. Yellow lines represent amyloplast levels, white stars mark the QC, green arrows mark the CSC layer, white arrows mark amyloplast in CSC. (m/n) indicates m in n of biological repeats showing the displayed features. Scale bar: 25 µm.

Extended Data Fig. 7 TOP1 regulates mTOR and cell cycle gene expression in response to Glc.

a, The cell viability of human NSCLC cell line HCC827 under the indicated treatments. Data represent mean ± s.d. of 3 biological repeats. Student’s t test (two-tailed). b, RT-qPCR of mTOR and cell cycle gene expression in HCC827 under the indicated treatments. Gene expression in untreated cells in 5 mM Glc was set to 1. Data represent mean ± s.d. of 3 biological repeats. Student’s t test (two-tailed). GAPDH served as negative control.

Extended Data Fig. 8 TOP1α and TOR regulate PLT2 expression.

a, b, Gene expression (a) and protein distribution of SHR, SCR, and PLT1 in plants (b) of the indicated genotypes. Gene expression in WT –Suc was set to 1. Data represent mean ± s.d. of 3 biological repeats. Student’s t test (two-tailed). (m/n) indicates m in n of biological repeats showing the displayed features. Scale bars: 50 µm. c, Diagram of the PLT2 locus. Gray and black rectangles and black lines represent untranslated regions, coding regions, and introns, respectively. A DNA fragment (shown in red) was deleted in the plt2 mutant. d, Quantification of CC number in plants of the indicated genotypes. Data were analyzed from 15 seedlings for each genotype from 3 experiments. Statistical significance was determined by one-way ANOVA and Tukey test (P < 0.05) (see Supplementary Data 1 for specific P values). e, Root gravity responses and amyloplast formation in top1α and top1α plt2 plants of the indicated genotypes. Seedlings were turned 90° from the vertical axis and fixed at the indicated time points. Amyloplast formation and levels in layers S1 and S2 were examined. Yellow lines represent amyloplast levels, white arrow marks amyloplasts and green arrows mark the premature CSC indicating that plt2 mutation failed to rescue the QC quiescence. (m/n) indicates m in n of biological repeats showing the displayed features. Scale bars: 25 µm. f, Diagram of the SS1/2/3/4 and CYCD5;1 loci. Gray and black rectangles and black lines represent untranslated regions, coding regions, and introns, respectively. The regions tested by ChIP are marked by black lines. g, RT-qPCR of SS1-4 in Ler and top1a plants under the indicated treatment. Gene expression in WT was set to 1. Data represent mean ± s.d. of 3 biological repeats. Statistical significance was determined by one-way ANOVA and Tukey test (P < 0.05) (see Supplementary Data 1 for specific P values).

Extended Data Fig. 9 TOR stabilizes PLT2.

a, CFP florescence showing PLT2 transcript levels in pPLT2::CFP and pPLT2::CFP top1α under the indicated treatments. Scale bars: 50 µm. b, e, Y2H assays to examine TOR-PLT2 and TOR-PLT1 interactions (b) and to identify particular domains of PLT2 for the TOR-PLT2 interaction (e). The indicated construct combinations were co-transformed into yeast cells and screened on plates containing SD/–Leu,–Trp or SD/–Ade,–His,–Leu,–Trp medium. c, Amino acid sequence alignment of PLT1 and PLT2. The AP2 domains are marked by red lines. d, Diagram of the constructs used in the yeast two-hybrid (Y2H) assay. F: full length; N: N-terminus; C: C-terminus; KD: kinase domain; AP2: AP2 domain. f, g, Examination of PLT2 stability in a cell-free system. MBP-PLT2 was purified and added to cell extracts from WT plants treated with DMSO or MG132 (f) or from WT and tor-es plants (g). Anti-MBP and anti-P-S/T antibodies were used to detect MBP-PLT2 and phosphorylated MBP-PLT2, respectively. h, MBP-PLT2 signal intensity of (g) quantified by ImageJ. Values represent the mean ± s.d. (n = 3). Statistical significance was determined by Student’s t-test (two-tailed). i, Examination of MBP-PLT2S12A stability in a cell-free system. MBP-PLT2 and MBP-PLT2S12A were purified and added to cell extracts from WT plants. Anti-MBP and anti-P-S/T antibodies were used to detect MBP-PLT2 and phosphorylated MBP-PLT2, respectively. j, l, CC number (j) and meristematic zone (l) of the indicted plants. Seedling was grown on -Suc under dark condition for 5 days. The CC number (j) and PLT2-GFP level (Fig. 3j) were examined. Data were analyzed from 15 seedlings for each genotype from 3 experiments. Statistical significance was determined by Student’s t test (two-tailed). k, The root phenotypes of the indicated plants. The seedlings were grown on 1/2MS plate under light condition for 5 days after germination. The PLT2-GFP level and RAM size were examined. The white arrows indicate the first elongated cortex cell and the yellow bars indicate the RAM size. Scale bars: 50 µm. For a and k, (m/n) indicates m in n of biological repeats showing the displayed features. For f, g and i, experiments were repeated three times with similar results. For gel source data, see Unmodified_Gels_Fig2.

Source data

Extended Data Fig. 10 A model of the role of TOP1α in fine-tuning TOR-PLT2 to maintain DM and CC homeostasis in the root tip.

Schematic diagram of the TOP1α- and TOR-mediated regulatory network used by plants to maintain DM and CC homeostasis in response to photosynthesis-derived Suc. Arrows represent activation; lines with bars represent repression. Dashed lines represent indirect regulation, and solid lines represent direct regulation.

Supplementary information

Supplementary Information

Supplementary Fig. 1.

Reporting Summary

Supplementary Data 1

Exact P values of one-way ANOVA and Tukey test.

Supplementary Data 2

Gene expression in RNA-seq analysis.

Supplementary Data 3

Primers used for plasmid constructs, RT–qPCR and mutagenesis.

Supplementary Data 4

All raw data for the statistical analysis that are visualized in the article.

Source data

Source Data Fig. 3

Unprocessed western blots and/or gels.

Source Data Extended Data Fig. 9

Unprocessed western blots and/or gels.

Rights and permissions

Springer Nature or its licensor holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Zhang, H., Guo, L., Li, Y. et al. TOP1α fine-tunes TOR-PLT2 to maintain root tip homeostasis in response to sugars. Nat. Plants 8, 792–801 (2022). https://doi.org/10.1038/s41477-022-01179-x

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41477-022-01179-x

This article is cited by

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing