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Georgios Tzelepis

Georgios Tzelepis
I work as an Assistant Professor (Associate Senior Lecturer) at Swedish University of Agricultural Sciences (SLU), Department of Forest Mycologist and Plant Pathology, on Prof Magnus Karlsson research group. My research interests are focused on soil-borne plant pathogens such as Rhizoctonia solani and Verticillium longisporum and the quality control fungi deploy to cope with ER stress caused by accumulation of misfolded N-glycoproteins.

Presentation

I study the mechanisms that these organisms utilize (effector proteins) to overcome the plant immune system, how plants respond to this attack, and how pathogens sense the environment in rhizosphere in order to establish a successful infection. I am also interested in studying how the deglycosylation process of misfolded N-glycoproteins affects fungal physiology and virulence in plant and human pathogens. I am a new group leader and my group consists of a postdoctoral researcher and a graduate student. In 2021, I promoted to Docent in Biology at SLU.

 

Teaching

I am involved in teaching at different Mycology and Plant Pathology courses  (MSc and PhD level) at SLU and Uppsala University.

Research

The protein quality control in filamentous fungal species

Eukaryotic cells produce a huge amount of proteins; the majority of them are glycoproteins, which are proteins contain a sugar chain (N-glycan). N-glycans play a significant role in protein function and stability, protecting them from proteolysis, facilitating the protein folding and secretion, affecting the protein solubility and the protein localization in the cells. It is also known that N-glycoproteins are structural componets of plasma membranes and fungal cell wall. N-glycoproteins are synthezised in the endoplasmic reticulum (ER) and Golgi apparatus and their glycan structures are processed as they proceed through the secretory pathway. Glycoprotein folding is one of the most important post-translational modifications in eukaryotic cells. When proteins consistently fail to be folded properly are trimmed by the ER-degradation enhancing α-mannosidase proteins (EDEMs) and are retrotranslocated to cytosol for further degradation by the ERAD pathway.

When misfolded glycoproteins are exported to cytosol the glycan chain must be cleaved from the protein in order to be degraded efficiently by the proteasome (10). This removal is conducted by the action of the ubiquitous, cytoplasmic peptide:N-glycanase (PNGase), releasing free N-glycans to the cytosol (Figure 1).

Proteins, illustration.

Figure 1. The deglycosylation activity of misfolded glycoproteins in eukaryotic cells.

 


This enzyme cleaves the amide bond in the side chain of glycosylated–asparagine residue generating free N-glycans with a chitobiose structure at the reducing terminus (GN2). Enzymatically active PNGases have been reported in a wide range of eukaryotic cells from mammals to yeasts. But the situation seems to be different in filamentous fungi, since data claim that the enzymatic function of the cytosolic neutral PNGases has been abolished. However, an acidic PNGase has been identified in many filamentous species but its function is completely unknown. When free N-glycans are created in cytosol they are further catabolized in order to be utilized possibly as a sugar source by cells. In cytosol two enzymes are responsible for free N-glycans degradation: the endo-β-N-acetyloglucosaminidases (ENGases) and the α-mannosidases. The role of ENGases in filamentous fungi seems to be crucial, since deletion of the cytosolic ones has a severe impact in fungal phenotype affecting among others the hyphal growth, conidiation, tolerance to abiotic stress, secretion etc. Despite the fact that ENGases have an important contribution in fungal biology many aspects remains unknown.

We study the mechanisms that filamentous fungi deploy in order to cope with ER stress caused by accumulation of misfolded glycoproteins. Our previous results showed that these organisms possibly utilize different mechanisms as compared to mammalian cells and to yeasts. The model fungal species Aspergillus nidulans is used in this project

Effector proteins in soilborne pathogens

Plant pathogens are categorized in biotrophs, necrotrophs and hemibiotrophs, depending on their interaction established with their hosts. Biotrophy defined the growth on living plant cells; hemibiotrophic pathogens establish first a biotrophic stage and then switch to necrotrophic, while necrotrophic pathogens grow on dead plant material. However, many necrotrophs rely on an initial short biotrophic phase in order to establish a successful infection. It is known that pathogens deploy small-secreted proteins termed as effectors to manipulate plant defense responses. These proteins are usually host even lineage- specific. There are also some effectors, which are more conserved, such as the necrosis and ethylene–inducing-like proteins (NLPs) and the LySM effectors, which are present in a broad range of organisms.

Verticillium longisporum is a soil borne pathogen, infecting plants in Brassicaceae family, such as oilseed rape (Brassica napus), cauliflower (Brassica oleracea), turnip (Brassica rapa subsp rapa) etc (Figure 2). According to the data available by now, V. longisporum is assumed to be a hemibiotrophic pathogen.

Plants in pots, photo.

Figure 2. Symptoms on Brassica napus plants infected by Verticillium longisporum (left) compared to healthy plants (right).

This pathogen is responsible for severe yield losses worldwide. Any attempt to control this disease has a limited success since this pathogen forms special resting structures, able to survive in soil for a long time and under harsh environmental conditions. Furthermore, no resistant to V. longisporum cultivars have been developed by now.Our previous data show that V. longisporum contains a certain number of genes encoding putative effector proteins, but nothing is known about their contribution in virulence and pathogenicity. The main questions arisen in this project are:

  • How the candidate effector proteins are regulated?
  • What is their functional role in this pathosystem?

Rhizoctonia solani is a soilborne Basidiomycete causing dumping-off disease in seedlings (Figure 3). It is assumed as a nectroph.

Plants in pots, photo.

Figure 3. Infected sugar beet seedlings with Rhizoctonia solani (right) compared to healthy plants (left).

It forms microsclerotia, which are able to survive to the soil for long periods and under unfavourable environmental conditions. This pathogen is considered to have a necrotrophic lifestyle and only few effectors, promoting necrosis, have been identified. We focus on effectors that do not cause necrosis, and are induced on early-infection stages, studying their roles and their structure. A LysM effector recently has been characterized in this pathogen and it works as a chitin-binding protein, suppressing chitin-triggered immunity, similar to filamentous hemibiotrophic Ascomycetes 

Different techniques are used, such as construction of deletion strains, heterologous expression of proteins, X-ray crystallography, MS/MS spectrometry, electron and confocal microscopy, RNA-seq etc.

Cooperation

I am in close cooperation with Prof. Hanna Johannesson at Stockholm University, investigating the genome complex of the soilborne pathogen Verticillium longisporum.

I also work together with scientists at Aristotle University of Thessaloniki, Greece, studying the multidrug resistance mechanisms in the plant pathogen Botrytis cinerea.

Background

Research experience

  • Associate senior lecturer (2019- now)
    Swedish University of Agricultural Sciences, Uppsala, Sweden.
  • Researcher in Plant Microbe Interactions (2017 - 2019).
    •  Swedish University of Agricultural Sciences, Department of Plant Biology, Uppsala BioCenter, Linnean Center for Plant Biology
    • Wageningen University, Laboratory of Phytopathology, the Netherlands
    • University of Crete, Department of Biology and Institute of Molecular Biology and Biotechnology Foundation of Research and Technology, Crete
  • Post-Doctoral fellow (2015 - 2017).
    • Swedish University of Agricultural Sciences, Department of Plant Biology, Uppsala BioCenter, Linnean Center for Plant Biology. 

Degrees and Education

  • PhD in Biology, (2010 - 2014): Swedish University of Agricultural Sciences (SLU), Uppsala Biocentrum, Department of Forest Mycology and Plant Pathology. Subject of Thesis: Functional differentiation of glycoside hydrolases family 18 in filamentous Ascomycetes.
  • MSc Degree in Crop Protection, (2006 – 2008) specialization in Plant Pathology. Aristotle University of Thessaloniki, School of Agriculture, Plant Pathology Laboratory.
  • BSc Degree in Agricultural Sciences (2001 – 2006): Aristotle University of Thessaloniki, School of Agriculture, specialization: Crop Protection.

Supervision

I have co-supervised five PhD students and eleven graduate and undergraduate students at SLU and Wageningen University.

Selected publications

  1. Rafiei V, Vélëz H, Piombo E, Dubey M, Tzelepis G (2023). Verticillium longisporum phospholipase VlsPLA2 is a virulence factor that targets host nuclei and modulates plant immunity. Molecular Plant Pathology 24:1078-1092. 
  2.  Rafiei V, Vélëz H, Dixelius C, Tzelepis G (2023).Advances in molecular interactions on the Rhizoctonia solani-sugar beet pathosystem. Fungal Biology Reviews 44: 100297.
  3. Rafiei V., Ruffino A., Persson Hodén C., Tornkvist A., Mozuraitis R., Dubey M., Tzelepis G. (2022). A Verticillium longisporum pleiotropic drug transporter determines tolerance to the plant host β-pinene monoterpene. Molecular Plant Pathology. 23: 291-303.
  4. Rafiei V., Velez H., Tzelepis G. (2021). The role of glycoside hydrolases in phytopathogenic fungi and oomycetes virulence. International Journal of Molecular Sciences. 22:9359.
  5. Samaras A., Karaoglanidis G., Tzelepis G. (2021). Insights into the multitrophic interactions between the biocontrol agent Bacillus subtilis MBI 600, the pathogen Botrytis cinerea and their plant host. Microbiological Research 248, 126752. 
  6. Tzelepis G., Dölfors F., Holmquist L., Dixelius C. (2021). Plant mitochondria and chloroplasts are targeted by the Rhizoctonia solani RsCRP1 effector. Biochemical and Biophysical Research Communications 544, 86–90.
  7. Charova, S., Dölfors, F., Holmquist, L., Moschou, P., Dixelius, C., Tzelepis, G. (2020). The RsRlpA Effector Is a Protease Inhibitor Promoting Rhizoctonia solani Virulence through Suppression of the Hypersensitive Response. International Journal of Molecular Sciences 21: 8070
  8. Tzelepis G., Persson-Hodèn K., Fogelqvist J., Åsman A., Vetukuri R.R., Dixelius C. (2019). Dominance of mating type A1 and possible epigenetic effects during mating in Phytophthora infestans. Frontiers in Microbiology 11:252.
  9. Dolfors F., Holmquist L., Dixelius C., Tzelepis G. (2019). A LysM effector protein from the basidiomycete Rhizoctonia solani contributes to virulence through suppresion of chitin-triggered immunity. Molecular Genetics and Genomics. 294: 1211-1218.
  10. Tzelepis G. and Karlsson M. (2019). Killer toxin-like chitinases in filamentous fungi: Structure, regulation and potential roles in fungal biology. Fungal Biology Reviews. 33: 123-132.
  11. Fogelqvist J., Tzelepis G., Bejai., Ilbäck J., Schwelm A. and Dixelius C. (2018).  Analysis of the hybrid genomes of two field isolates of the soil- borne fungal   species Verticillium longisporum. BMC Genomics. 19:14
  12. Tzelepis G., Karlsson M. and Suzuki T. (2017). Deglycosylating enzymes acting on N-glycans in fungi: Insights from genome survey. BBA General Subjects, 1861:2551–255.
  13. Wibberg D., Andersson L., Tzelepis G., Rupp O. et al. (2016). Genome analysis of the sugar beet pathogen Rhizoctonia solani AG2-2IIIB revealed high numbers in secreted proteins and cell wall degrading enzymes. BMC Genomics, 17:245.
  14. Tzelepis G., Dubey M., Jensen D. F and Karlsson M. (2015). Identifying glycoside hydrolase family 18 genes in the mycoparasitic fungal species Clonostachys rosea. Microbiology-Sgm, 161: 1407-1419.
  15. Karlsson M., Brandström-Durling M., Choi J., Lackner G, Tzelepis G., et al (2015). Insights on the evolution of mycoparasitism from the genome of Clonostachys rosea. Genome Biology and Evolution. 7 (2): 465-480.
  16. Tzelepis G., Hosomi A., Hossain J.T., Hirayama H., Dubey M., Jensen D. F., Suzuki T. and Karlsson, M. (2014).  Endo-β-N-acetylglucosamidases (ENGases) in the fungus Trichoderma atroviride: Possible involvement of the filamentous fungi-specific cytosolic ENGase in the ERAD process. Biochemical and Biophysical Research Communications 449: 256-261.
  17. Tzelepis G., Melin P., Jensen D. F, Stenlid J. and Karlsson M. (2014). Functional analysis of the C-II subgroup killer toxin-like chitinases in the filamentous ascomycete Aspergillus nidulans. Fungal Genetics and Biology 64:58-66.
  18. Tzelepis G., Melin P, Jensen D. F, Stenlid J. and Karlsson M. (2012). Functional analysis of glycoside hydrolase family 18 and 20 genes in Neurospora crassa. Fungal Genetics and Biology. 49: 717-730.

Book chapters

  1. Tzelepis G. and Karlsson M. (2020). The fungal chitinases. Encyclopedia of Mycology, (eds) Elsevier, pp 23-31. 

 

 

 

 

 

 

 

 

 


Contact

Senior Lecturer at the Department of Forest Mycology and Plant Pathology; Division of Plant Pathology
Telephone: +4618-671503
Postal address:
Skoglig mykologi och växtpatologi , Box 7026
750 07 UPPSALA
Visiting address: Almas Allé 5, Uppsala