A single-nucleus and spatial transcriptomic atlas of poplar leaves reveals the regulation of leaf polarity and cuticle deposition

Yiling Li; Lingfei Kong; Xing Guo; Yang Chen; Wenwen Shao; Min Liu; Chenmeng Luo; Shaoming Liang; Ao Feng; Li Xu; Huan Liu; Tong Wei; Yuanzhong Jiang; Tao Ma

doi:10.1016/j.jgg.2026.03.015

Turn off MathJax

Article Contents

Article Navigation > Journal of Genetics and Genomics > 2026 > Accepted Manuscript

PDF( 0 KB)

A single-nucleus and spatial transcriptomic atlas of poplar leaves reveals the regulation of leaf polarity and cuticle deposition

doi: 10.1016/j.jgg.2026.03.015

a. Key Laboratory of Bio-Resource and Eco-Environment of Ministry of Education, Laboratory for Ex Situ Conservation and Resource Utilization of Montane Plants, Sichuan Zoige Alpine Wetland Ecosystem National Observation and Research Station, College of Life Sciences, Sichuan University, Chengdu, Sichuan 610065, China;
b. Chengdu Botanical Garden, Chengdu, Sichuan 610083, China;
c. Chongqing Key Laboratory of Forest Resource Innovation and Utilization, Integrative Science Center of Germplasm Creation in Western China (Chongqing) Science City, School of Life Sciences, Southwest University, Chongqing 400715, China;
d. BGI Research, Wuhan, Hubei 430047, China;
e. Guangdong Provincial Key Laboratory for Plant Epigenetics, College of Life Sciences and Oceanography, Shenzhen University, Shenzhen, Guangdong 518060, China;
f. State Key Laboratory of Genome and Multi-omics Technologies, BGI Research, Shenzhen, Guangdong 518083, China

Funds:

We thank L.-F.K. (Southwest University) for the myc2 double mutants and MYC2b-OE lines. We thank Key Laboratory of Genomics, Ministry of Agriculture, BGI-Shenzhen, Shenzhen 518120, China and Guangdong Provincial Key Laboratory of core collection of crop genetic resources research and application, BGI-Shenzhen, Shenzhen 518120, China for data sequencing. This work was supported by the National Key Research and Development Program of China (2021YFD2201100), the National Natural Science Foundation of China (32322057, and 32071732), and Fundamental Research Funds for the Central Universities (2020SCUNL207, SCU2021D006 and 2020SCUNL103) to T. M and Y.-Z. J.

Received Date: 2025-10-09
Accepted Date: 2026-03-16
Rev Recd Date: 2026-03-10

Available Online: 2026-03-24

Abstract

Abstract

Leaf adaxial-abaxial polarity is fundamental for plant morphogenesis and environmental adaptation through asymmetric cell differentiation. Emerging evidence reveals dorsoventral metabolic gradients act downstream of transcriptional networks to fine-tune cellular specialization. While conserved transcription factors (e.g., HD-ZIP III, KANADI) establish initial polarity, the molecular networks driving position-specific cellular differentiation and their integration with metabolic adaptation remain unclear. Leveraging single-nucleus and spatial transcriptomics, we resolve major cell classes (mesophyll, epidermal, and vascular-associated) and their adaxial-abaxial subtypes, revealing dorsoventral polarity in transcriptional profiles and metabolic pathways. Adaxial cells are enriched in phenylpropanoid/flavonoid biosynthesis, while abaxial cells show preferential activation of stress and hormone signaling. Notably, we identify MYC2 as a key regulator of adaxial cuticle biosynthesis, binding to promoters of lipid biosynthetic and transport genes (e.g., CER10, LTPG1) and promoting cuticle thickening. Our study uncovers how positional identity shapes transcriptional and metabolic polarity in leaves, with MYC2 emerging as a central regulator coordinating organ-specific adaptations. These findings provide insights into the spatial regulation of plant development and stress resilience, offering potential strategies for engineering stress-tolerant woody crops.
- Leaf adaxial-abaxial polarity,
- Spatial transcriptomics,
- Single-nucleus transcriptomics,
- Transcriptional regulation,
- CUT&,
- Tag-seq

FullText(HTML)

References(44)

References

Abe, H., Urao, T., Ito, T., Seki, M., Shinozaki, K., Yamaguchi-Shinozaki, K., 2003. Arabidopsis AtMYC2 (bHLH) and AtMYB2 (MYB) function as transcriptional activators in abscisic acid signaling. Plant Cell 15, 63-78.

Agati, G., Brunetti, C., Di Ferdinando, M., Ferrini, F., Pollastri, S., Tattini, M., 2013. Functional roles of flavonoids in photoprotection: new evidence, lessons from the past. Plant Physiol. Biochem. 72, 35-45.

Aharoni, A., Dixit, S., Jetter, R., Thoenes, E., Van Arkel, G., Pereira, A., 2004. The SHINE clade of AP2 domain transcription factors activates wax biosynthesis, alters cuticle properties, and confers drought tolerance when overexpressed in Arabidopsis. Plant Cell 16, 2463-2480.

Bird, D., Beisson, F., Brigham, A., Shin, J., Greer, S., Jetter, R., Kunst, L., Wu, X., Yephremov, A., Samuels, L., et al., 2007. Characterization of Arabidopsis ABCG11/WBC11, an ATP binding cassette (ABC) transporter that is required for cuticular lipid secretion. Plant J. 52, 485-498.

Birkenbihl, R.P., Diezel, C., Somssich, I.E., 2012. Arabidopsis WRKY33 is a key transcriptional regulator of hormonal and metabolic responses toward Botrytis cinerea infection. Plant Physiol. 159, 266-285.

Bowman, J.L., Eshed, Y., Baum, S.F., 2002. Establishment of polarity in angiosperm lateral organs. Trends Genet. 18, 134-141.

Broun, P., Poindexter, P., Osborne, E., Jiang, C.Z., Riechmann, J.L., 2004. WIN1, a transcriptional activator of epidermal wax accumulation in Arabidopsis. Proc. Natl. Acad. Sci. U. S. A. 101, 4706-4711.

Caarls, L., Elberse, J., Awwanah, M., Ludwig, N.R., De Vries, M., Zeilmaker, T., Van Wees, S.C., Schuurink, R.C., Van den Ackerveken, G., 2017. Arabidopsis jasmonate-induced oxygenases down-regulate plant immunity by hydroxylation and inactivation of the hormone jasmonic acid. Proc. Natl. Acad. Sci. U. S. A. 114, 6388-6393.

Cai, J., Panda, S., Kazachkova, Y., Amzallag, E., Li, Z., Meir, S., Rogachev, I., Aharoni, A., 2024. A NAC triad modulates plant immunity by negatively regulating N-hydroxy pipecolic acid biosynthesis. Nat. Commun. 15, 7212.

Chen, S., Zhou, Y., Chen, Y., Gu, J., 2018. Fastp: an ultra-fast all-in-one FASTQ preprocessor. Bioinformatics 34, i884-i890.

Chen, T.W., Li, H.P., Lee, C.C., Gan, R.C., Huang, P.J., Wu, T.H., Lee, C.Y., Chang, Y., Tang, P., 2014. ChIPseek, a web-based analysis tool for ChIP data. BMC Genomics 15, 539.

Chen, Y., Tong, S., Jiang, Y., Ai, F., Feng, Y., Zhang, J., Gong, J., Qin, J., Zhang, Y., Zhu, Y., et al., 2021. Transcriptional landscape of highly lignified poplar stems at single-cell resolution. Genome Biol. 22, 319.

Chini, A., Fonseca, S., Fernandez, G., Adie, B., Chico, J. M., Lorenzo, O., Garcia-Casado, G., Lopez-Vidriero, I., Lozano, F. M., Ponce, M. R., et al., 2007. The JAZ family of repressors is the missing link in jasmonate signalling. Nature 448, 666-671.

Choe, Y., Pleasure, S.J., Mira, H., 2015. Control of adult neurogenesis by short-range morphogenic-signaling molecules. Cold Spring Harb. Perspect. Biol. 8, a018887.

Cui, F., Brosche, M., Lehtonen, M.T., Amiryousefi, A., Xu, E., Punkkinen, M., Valkonen, J.P.T., Fujii, H., Overmyer, K., 2016. Dissecting abscisic acid signaling pathways involved in cuticle formation. Mol. Plant 9, 926-938.

Dominguez, E., Heredia-Guerrero, J.A., Heredia, A., 2011. The biophysical design of plant cuticles: an overview. New Phytol. 189, 938-949.

Emery, J.F., Floyd, S.K., Alvarez, J., Eshed, Y., Hawker, N.P., Izhaki, A., Baum, S.F., Bowman, J.L., 2003. Radial patterning of Arabidopsis shoots by class III HD-ZIP and KANADI genes. Curr. Biol. 13, 1768-1774.

Erickson, B.J., Staples, N.C., Hess, N., Staples, M.A., Weissert, C., Finkelstein, R.R., Cooper, J.B., 2020. PRPs localized to the middle lamellae are required for cortical tissue integrity in Medicago truncatula roots. Plant Mol. Biol. 102, 571-588.

Eshed, Y., Izhaki, A., Baum, S.F., Floyd, S.K., Bowman, J.L., 2004. Asymmetric leaf development and blade expansion in Arabidopsis are mediated by KANADI and YABBY activities. Development 131, 2997-3006.

Fabre, G., Garroum, I., Mazurek, S., Daraspe, J., Mucciolo, A., Sankar, M., Humbel, B.M., Nawrath, C., 2016. The ABCG transporter PEC1/ABCG32 is required for the formation of the developing leaf cuticle in Arabidopsis. New Phytol. 209, 192-201.

Fukaki, H., Wysocka-Diller, J., Kato, T., Fujisawa, H., Benfey, P.N., Tasaka, M., 1998. Genetic evidence that the endodermis is essential for shoot gravitropism in Arabidopsis thaliana. Plant J. 14, 425-430.

Fukuda, N., Oshima, Y., Ariga, H., Kajino, T., Koyama, T., Yaguchi, Y., Tanaka, K., Yotsui, I., Sakata, Y., Taji, T., 2022. ECERIFERUM 10 encoding an enoyl-CoA reductase plays a crucial role in osmotolerance and cuticular wax loading in Arabidopsis. Front. Plant Sci. 13, 898317.

Godoy, M., Franco-Zorrilla, J.M., Perez-Perez, J., Oliveros, J.C., Lorenzo, O., Solano, R., 2011. Improved protein-binding microarrays for the identification of DNA-binding specificities of transcription factors. Plant J. 66, 700-711.

He, Z., Webster, S., He, S.Y., 2022. Growth-defense trade-offs in plants. Curr. Biol. 32, R634-R639.

Huang, H., Wang, Y., Yang, P., Zhao, H., Jenks, M.A., Lu, S., Yang, X., 2024. The Arabidopsis cytochrome P450 enzyme CYP96A4 is involved in the wound-induced biosynthesis of cuticular wax and cutin monomers. Plant J. 118, 1619-1634.

Javelle, M., Vernoud, V., Rogowsky, P.M., Ingram, G.C., 2011. Epidermis: the formation and functions of a fundamental plant tissue. New Phytol. 189, 17-39.

Kazan, K., Manners, J.M., 2013. MYC2: the master in action. Mol. Plant 6, 686-703.

Kim, D., Paggi, J.M., Park, C., Bennett, C., Salzberg, S.L., 2019. Graph-based genome alignment and genotyping with HISAT2 and HISAT-genotype. Nat. Biotechnol. 37, 907-915.

Kunst, L., Samuels, A.L., 2003. Biosynthesis and secretion of plant cuticular wax. Prog. Lipid Res. 42, 51-80.

Lee, S. B., Go, Y. S., Bae, H. J., Park, J. H., Cho, S. H., Cho, H. J., Lee, D. S., Park, O. K., Hwang, I., Suh, M. C., 2009. Disruption of glycosylphosphatidylinositol-anchored lipid transfer protein gene altered cuticular lipid composition, increased plastoglobules, and enhanced susceptibility to infection by the fungal pathogen Alternaria brassicicola. Plant Physiol. 150, 42-54.

Lewandowska, M., Keyl, A., Feussner, I., 2020. Wax biosynthesis in response to danger: its regulation upon abiotic and biotic stress. New Phytol. 227, 698-713.

Li, H., Handsaker, B., Wysoker, A., Fennell, T., Ruan, J., Homer, N., Marth, G., Abecasis, G., Durbin, R., 1000 Genome Project Data Processing Subgroup, 2009. The sequence alignment/map format and SAMtools. Bioinformatics 25, 2078-2079.

Li, J., Brader, G., Palva, E.T., 2004. The WRKY70 transcription factor: a node of convergence for jasmonate-mediated and salicylate-mediated signals in plant defense. Plant Cell 16, 319-331.

Li, Y., Beisson, F., Koo, A.J., Molina, I., Pollard, M., Ohlrogge, J., 2007. Identification of acyltransferases required for cutin biosynthesis and production of cutin with suberin-like monomers. Proc. Natl. Acad. Sci. U. S. A. 104, 18339-18344.

Pertea, M., Pertea, G.M., Antonescu, C.M., Chang, T.C., Mendell, J.T., Salzberg, S.L., 2015. StringTie enables improved reconstruction of a transcriptome from RNA-seq reads. Nat. Biotechnol. 33, 290-295.

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

Satija, R., Farrell, J.A., Gennert, D., Schier, A.F., Regev, A., 2015. Spatial reconstruction of single-cell gene expression data. Nat. Biotechnol. 33, 495-502.

Wu, T., Hu, E., Xu, S., Chen, M., Guo, P., Dai, Z., Feng, T., Zhou, L., Tang, W., Zhan, li., et al., 2021. clusterProfiler 4.0: a universal enrichment tool for interpreting omics data. Innovation (Camb.) 2, 100141.

Xia, K., Sun, H. X., Li, J., Li, J., Zhao, Y., Chen, L., Qin, C., Chen, R., Chen, Z., Liu, G., et al., 2022. The single-cell stereo-seq reveals region-specific cell subtypes and transcriptome profiling in Arabidopsis leaves. Dev. Cell 57, 1299-1310.

Yeats, T., Rose, J.K.C., 2013. The formation and function of plant cuticles. Plant Physiol. 163, 5-20.

Zander, M., Lewsey, M. G., Clark, N. M., Yin, L., Bartlett, A., Saldierna Guzman, J. P., Hann, E., et al., Langford, A. E., Jow, B., et al., 2020. Integrated multi-omics framework of the plant response to jasmonic acid. Nat. Plants 6, 290-302.

Zhang, X., Zhu, Z., An, F., Hao, D., Li, P., Song, J., Yi, C., Guo, H., 2014. Jasmonate-activated MYC2 represses ETHYLENE INSENSITIVE3 activity to antagonize ethylene-promoted apical hook formation in Arabidopsis. Plant Cell 26, 1105-1117.

Zhang, Y., Liu, T., Meyer, C. A., Eeckhoute, J., Johnson, D. S., Bernstein, B. E., Nusbaum, C., Myers, R. M., Li, W., et al., 2008. Model-based analysis of ChIP-seq (MACS). Genome Biol. 9, R137.

Zeier, J., Schreiber, L., 1998. Ontogenetic and seasonal development of wax composition and cuticular transpiration of ivy (Hedera helix L.) sun and shade leaves. Planta 207, 67-75.

Relative Articles

Supplements(0)

Cited By

Proportional views