Plant Cell Responses to Nematode-Driven Mechanical Breach

Research


Muhammad Shahzad Anjam is standing in a greenhouse wearing a blue coat, only his head and sholders are visible and some small green plants in the back, he is looking straight into the cameraMuhammad Shahzad Anjam (photo: Maria Kidwai)

Plants are constantly exposed to biotic and abiotic stresses, resulting in considerable yield losses in agriculture. Among the most significant threats are plant-parasitic nematodes, microscopic organisms that attack plant roots and contribute to global agricultural losses of $157 billion annually. This research project aims to explore how plants detect and respond to mechanical damage caused by endoparasitic cyst nematodes as they migrate through plant cells.

Plants are always encountered by biotic and abiotic stresses which causes significant yield losses. While growing in the soil, plant roots undergo frequent mechanical injuries from soil texture and invasive pathogens such as nematodes. Plant-parasitic nematodes are microscopic organisms with exceptionally broad host range that pose major challenge to global agriculture accounting for $157 billion each year. The losses are further enhanced when they form disease-complexes with other microbes.

Endoparasitic cyst nematodes penetrate successive cell files and reach the vasculature, where they establish specialized cellular feeding structures. During this migration phase, they move intracellularly while leaving behind a trail of dead cells (Figure 1). How plant cells sense and respond to such mechanical injuries are investigated in current research project.

Two microscope images beside each other: on the left side a brightfield image of a part of a root that is infected with a nematode; the nematode is labeled with dotted lines; a fluorescence image of the same root is displayed on the right side; the damage of the cells caused by the nematode infection is highlighted with small yellow arrows Figure 1: Cellular damage in Arabidopsis root during migration phase of cyst nematode (illustration: Muhammad Shahzad Anjam)

Cell wall: Structural barrier

The plant cell wall, a first physical barrier, is a dynamic and complex structure that is continuously monitored and maintained to ensure its functionality during growth and environmental interactions. It is mainly composed of cellulose, hemicellulose and pectin whereas additionally lignin is also deposited in the secondary cell wall of vascular tissues. The lignin is rigid compound and provides mechanical strength, stiffness, hydrophobicity and protection against invaders (Figure 2). In Arabidopsis roots, the lignin is found in the xylem vessel and in the endodermal cells (Figure 2, left). The endodermis contains lignified casparian strip as well as suberin lamellae that potentially contributes to defence against parasitic nematodes. Among other plant-parasitic nematodes, the cyst nematodes such as Heterodera spp. are known for breaching the endodermis cell layer to reach vascular tissues at any root zone. As nematodes only rely on reserved food in their body for reaching the target cell and developing the feeding site, strengthening the cell layers with local ectopic lignification and suberization is potential strong barrier for reducing physical penetration of nematodes to the root central cylinder.

Left: Schematic illustration of the different layers of the cell wall including the middle lamella, the primary cell wall, the secondary cell wall with its different components (cellulose, pectin, hemicellulose, lignin) and the cell membrane; Right: Cross section of an infected Arabidopsis root taken with a microscope: the cell walls are seen as white surroundings; their diameter becomes tinier towards the center of the root; som parts are labelled with red colour to illustrate damaged cell wallsFigure 2:
Plant’s secondary cell wall contains lignin, pectin, cellulose and hemicellulose shown in red, yellow, green and blue respectively (right). Left: Orthogonal structure of Arabidopsis root, stained with basic fuchsin (red) for lignin and calcofluor white (grey) for cell wall visibility.

We have found that plants have evolved a powerful signalling mechanism to detect defects in the endodermis barrier and can induce over-lignification, which is also associated with increased resistance to a variety of soil-borne pathogens. Our findings show that Arabidopsis roots deposit localized ectopic lignification in response to cyst nematode damage (Figure 3).

Laser Ablation: a tool to mimic nematode-induced injury

The nematodes constantly move in the root during the migration phase, damaging various cell files. Therefore analysing samples under confocal microscope in certain time and targeted region, is highly challenging. To overcome this, we employed laser ablation method to mimic nematode-induced injury in Arabidopsis roots (Figure 3). In our initial experiments have revealed lignification responses to mechanical injury with laser ablation in the context of the current project. Therefore, laser ablation-assisted cell wounding was employed to investigate lignin mediated defense responses.

Six microscope images displayed next to each other showing parts of three different plant roots; a brightfield image is always displayed left to an image in which lignin is lableled in red Figure 3: Localized lignin and suberin deposition in Arabidopsis root. A-C: 5-day old roots stained with basic fuchsin (red color) to visualize lignin. A: Control root. B: Nematodes damaging roots cells during migration phase. An outline has been drawn around nematode body to visualize nematodes. C: A single cortex cell damaged using laser ablation. The red color in figures A-C represents lignin stained with basic fuchsin.

Elucidating translational changes in cortex and endodermis cells in response to laser ablation and nematode associated damages

Our preliminary experiments showed that lignin deposition in response to mechanical damage is cell specific. Therefore, we aim to further investigate the cell-specific responses by using lignin staining, activation of phenylpropanoid and lignin biosynthetic pathway. For this purpose, highly sensitive nuclear localized, promoter specific fluorescent marker lines will be used in spatial and temporal manner following by mechanical injuries.

Identifying upstream transcription factors regulating the phenylpropanoid pathway specific to cell damage

Phenylpropanoid is complex pathway which produces phenolic precursors which are later on used consumed in lignin biosynthetic pathway to synthesize basic monolignol units, Syringyl (S), Guaiacyl (G) and p-cuomory hydroxyphenol (H). These monolignols polymerizes in the cell wall using peroxidases and laccases. The ratio of these monolignols determine the lignin composition and mechanical properties. The lignin induction is regulated strongly by internal and external environmental cues. Therefore, a fine regulation of the mechanism is required by the plant and thereby more than 20 MYB transcription factors are involved in the regulation. We sought, to identify the transcription factors regulating under mechanical injury. We shall analyse RNAseq data of Arabidopsis roots after infection with nematodes. The identified candidates will be further confirmed using highly sensitive, nuclear localized promoter specific Arabidopsis lines in spatio-temporal manner.

Revealing the nematode-specific molecular signalling of lignin biosynthesis and polymeric composition

The composition of lignin deposited in response to environmental cues is specific. The change in lignin composition is linked to the activation of plant basal defense mechanisms. Previously, monolignol G have been shown increasing in response to bacterial infection (REF). The purpose is to explore the lignin composition when employed mechanical wounding.

Exploring the potential of identified transcription factors in reducing nematode’s access to vasculature

The transcription factors regulating lignin deposition under infection conditions, could be potentially exploited to develop resistant genotypes. The aim is validating identified candidate MYB transcription factors if they play a role in nematode parasitism or resistance. The crispr/cas9 generated and t-DNA insertion mutant and overexpression lines under 35S promoter will be used for assessment of nematode infection under invitro conditions.

Selected Publications

Ca2+ Waves and Ethylene/JA Crosstalk Orchestrate Wound Responses in Arabidopsis Roots. Ma X., Hasan S., Anjam M.S., Mahmud S., Bhattacharyya S., Vothknecht U., Mendy B., Grundler F., and Marhavy P. EMBO Reports 2025. https://doi.org/10.1038/s44319-025-00471-z

Damage activates EXG1 and RLP44 to suppress vascular differentiation during regeneration in Arabidopsis. Mazumdar, S., Augstein, F., Zhang, A., Musseau, C., Anjam, M. S., Marhavy, P., & Melnyk, C. W. Plant Communications, 6(4): 101256. 2025. http://doi.org/10.1016/j.xplc.2025.101256

Cellular damage triggers mechano-chemical control of cell wall dynamics and patterned cell divisions in plant healing. Di Fino, L. M., Anjam, M. S., Besten, M., Mentzelopoulou, A., Papadakis, V., Zahid, N., Baez, L. A., Trozzi, N., Majda, M., Ma, X., Hamann, T., Sprakel, J., Moschou, P. N., Smith, R. S., & Marhavý, P. Developmental Cell. J 2025. http://doi.org/10.1016/j.devcel.2024.12.032

RNA Isolation from Nematode-Induced Feeding Sites in Arabidopsis Roots Using Laser Capture Microdissection. Anjam, M. S., Siddique, S., & Marhavý, P. In Duque, P., & Szakonyi, D., editor(s), Environmental Responses in Plants: Methods and Protocols, of Methods in Molecular Biology, pages 313–324. Springer US, New York, NY, 2022. https://doi.org/10.1007/978-1-0716-2297-1_22

Host factors influence the sex of nematodes parasitizing roots of Arabidopsis thaliana. Anjam, M. S., Shah, S. J., Matera, C., Różańska, E., Sobczak, M., Siddique, S., & Grundler, F. M. W. Plant, Cell & Environment, 43(5): 1160–1174. 2020. https://doi.org/10.1111/pce.13728

Signal Transduction in Plant–Nematode Interactions. Ali, M. A., Anjam, M. S., Nawaz, M. A., Lam, H., & Chung, G. International Journal of Molecular Sciences, 19(6): 1648. June 2018. https://doi.org/10.3390/ijms19061648

Arabidopsis HIPP27 is a host susceptibility gene for the beet cyst nematode Heterodera schachtii. Radakovic, Z. S., Anjam, M. S., Escobar, E., Chopra, D., Cabrera, J., Silva, A. C., Escobar, C., Sobczak, M., Grundler, F. M. W., & Siddique, S. Molecular Plant Pathology, 19(8): 1917–1928. 2018. https://doi.org/10.1111/mpp.12668

Genome-wide association study uncovers a novel QTL allele of AtS40-3 that affects the sex ratio of cyst nematodes in Arabidopsis. Anwer, M. A., Anjam, M. S., Shah, S. J., Hasan, M S., Naz, A. A, Grundler, F. M W, & Siddique, S. Journal of Experimental Botany, 69(7): 1805–1814. March 2018. Genome-wide association study uncovers a novel QTL allele of AtS40-3 that affects the sex ratio of cyst nematodes in Arabidopsis https://doi.org/10.1093/jxb/ery019

Damage-associated responses of the host contribute to defence against cyst nematodes but not root-knot nematodes. Shah, S. J., Anjam, M. S., Mendy, B., Anwer, M. A., Habash, S. S, Lozano-Torres, J. L, Grundler, F. M W, & Siddique, S. Journal of Experimental Botany, 68(21-22): 5949–5960. December 2017. Damage-associated responses of the host contribute to defence against cyst nematodes but not root-knot nematodes.https://doi.org/10.1093/jxb/erx374

An improved procedure for isolation of high-quality RNA from nematode-infected Arabidopsis roots through laser capture microdissection. Anjam, M. S., Ludwig, Y., Hochholdinger, F., Miyaura, C., Inada, M., Siddique, S., & Grundler, F. M. W. Plant Methods, 12(1): 25. April 2016. https://doi.org/10.1186/s13007-016-0123-9

A parasitic nematode releases cytokinin that controls cell division and orchestrates feeding site formation in host plants. Siddique, S., Radakovic, Z. S., De La Torre, C. M., Chronis, D., Novák, O., Ramireddy, E., Holbein, J., Matera, C., Hütten, M., Gutbrod, P., Anjam, M. S., Rozanska, E., Habash, S., Elashry, A., Sobczak, M., Kakimoto, T., Strnad, M., Schmülling, T., Mitchum, M. G., & Grundler, F. M. W. Proceedings of the National Academy of Sciences, 112(41): 12669–12674. October 2015. A parasitic nematode releases cytokinin that controls cell division and orchestrates feeding site formation in host plants.https://doi.org/10.1073/pnas.1503657112


Logo of Marie Skłodowska-Curie ActionsThis project has received funding from the European Union’s Horizon 2020 research and innovation programme under the Marie Sklodowska-Curie grant agreement No 101066035.