Phytophthora infestans effector PexRD54 associates with host Rab GTPase Rab8-1 to reprogram endomembrane transport

Lead Research Organisation: Imperial College London
Department Name: Life Sciences

Abstract

Filamentous plant pathogens are a group of eukaryotic pathogens including oomycete genus Phytophthora, as well as rust fungi, and Powdery mildew fungi, which cause most destructive plant diseases and threaten global food security. These pathogens cost billions of dollars annual looses to modern agriculture and severely impact subsistence agriculture in developing countries. These microorganisms are accommodated within the host cells through specialized cellular structures. However, little is known about molecular mechanisms underlying microbial accommodation inside the plant cells. The proposed research aims to characterize the host processes required for accommodation of filamentous plant pathogens inside the plant cells with a specific focus on illustrating the role of plant endomembrane transport system in this process. Among the oomycetes, Phytophthora spp. cause some of the most destructive plant diseases in the world and cause enormous economic damage to important crop species such as potato, tomato, and soybean, as well as environmental damage in natural ecosystems. Phytophthora infestans, the Irish potato famine pathogen, which causes late blight of potato and tomato, is one of the most important biotic threats to global food production leading to worldwide economic losses exceeding $5 billion annually. Unfortunately, late blight of potato is a reemerging destructive disease, which caused severe epidemics in potato farms across the UK lately. Our long-term objective is to dissect the molecular mechanisms underlying plant cell autonomous immunity and the role of endomembrane traffic in this process by particularly using P. infestans as a model pathogen. Like some other filamentous pathogens that form destructive plant diseases, P. infestans is accommodated inside the host cells by forming specialized compartments termed haustoria, which are separated from the invaded plant cells by newly synthesized host derived membranes with unknown origin and composition. This interface is critical for development of parasitic infection by enabling efficient macromolecule exchange. Therefore, understanding the regulation of macromolecule exchange processes at host pathogen interface is critical to develop novel strategies to engineer disease resistance in plants. Unfortunately, despite being discovered decades ago, our current knowledge about haustoria is limited. Deciphering the cellular and biochemical activities at this interface is critical for understanding the mechanisms of pathogenesis. Like other pathogens, P. infestans secretes a battery of proteins, termed effectors that suppress plant immunity and enable parasitic infection. Recently we discovered that some of these effectors specifically accumulate at the host pathogen interface inside the infected plant cells. In this study we aimed to identify host components of focal immunity using one such effector, termed PexRD54, as a molecular probe. Our preliminary work unraveled that around haustoria, PexRD54 associated with a host protein named Rab8-1, a member of Rab GTPases family of eukaryotic vesicle trafficking regulators. Our goal is to functionally characterize PexRD54 and Rab8-1 in order to establish the roles of these molecules at host pathogen interface. This study will help to establish functional characterization of plant endomembrane transport processes perturbed by pathogens and help understanding the origin, function and composition of the host pathogen interface. In turn, a detailed knowledge of PexRD54 manipulation of Rab8-1 will improve our understanding of the microbial accommodation inside the plant cells and will allow for creating renewed opportunities for engineering disease resistance in crops.

Technical Summary

Some filamentous plant pathogens associate with plant cells through specialized infection structures termed haustoria to deliver effector proteins or uptake nutrients. Haustorium is separated from the plant cell by a host-derived membrane known as the extrahaustorial membrane (EHM), with unknown origin and composition. Plant cells generally respond to pathogen penetration with a spatially confined cell-autonomous immune response known as focal immunity. Unfortunately, despite the documented importance of haustoria formation and focal immunity, our current knowledge in these processes is limited. These processes have been difficult to dissect using standard genetic approaches due to overlap between focal immunity and plant development. Pathogen secreted effectors accumulating around haustoria (perihaustorial effectors) provided an excellent alternative to study focal immunity. In addition to the reported P. infestans perihaustorial effectors, we identified a novel perihaustorial effector named PexRD54. By deploying PexRD54 as a molecular probe, we identified a host Rab GTPase that accumulated at the EHM named Rab8-1. The overall aim of this proposal is to determine how PexRD54 accumulates at haustoria and modulates host endomembrane transport by targeting Rab8-1. We will use genetic, biochemical, and cell imaging methods to functionally characterize PexRD54 and Rab8-1 association and elucidate their role in immunity. This research program is both timely and innovative because little is known about molecular mechanisms underlying microbial accommodation inside the plant cells. At the end of this study, we expect to dissect a unique type of effector targeted vesicle transport process and identify clues for the biogenesis and function of the EHM. The proposed research will uncover basic knowledge in endomembrane transport systems by assigning molecular functions to key regulators of this process and shed light on the microbial accommodation inside the plant cells.

Planned Impact

Plant diseases directly impact global food security, one of the most important problems for growing human population. While many people in developing countries suffer from starvation and related diseases, food prizes gradually increase in developed countries. Filamentous plant pathogens cause the most devastating plants diseases and lead to vast economic losses on important crop species as well as environmental damage in natural ecosystems. Among the oomycetes, Phytophthora spp. cause some of the most destructive plant diseases in the world and are arguably the most devastating pathogens of dicot plants. Phytophthora species cause enormous economic damage to important crop species such as potato, tomato, and soybean, as well as environmental damage in natural ecosystems. The most notable species is Phytophthora infestans, the Irish potato famine pathogen, which causes late blight of potato and tomato causing worldwide potato production losses exceeding $5 billion annually, making P. infestans one of the most important biotic threats to global food production. Currently, management of plant diseases such as the late blight disease is mainly achieved by applications of chemicals, some of which are likely to be banned in the near future. Understanding the biology of host colonization by P. infestans and other filamentous pathogens is not only critical for improving genetic control that can replace chemical control of plant diseases, but also for devising innovative disease management strategies and maintaining healthy natural ecosystems. The proposed project will shed light on the unknown nature of the role of endomembrane transport in plant immunity. This will address one of the major unknowns in this scientific field, and lead to the creation of novel strategies for manipulating plants towards resistance to filamentous pathogens. Such knowledge will also contribute to the leadership of the UK in the scientific field of molecular plant pathology. Consequently, we expect that our research proposal will potentially have an impact on non-academic units such as plant biotechnology corporates, the agricultural society, and policy makers.
The PI will manage the impact plan. The plan will be revised at monthly project meetings. Data generated will be publicly released in line with the BBSRC policy on data sharing. We will ensure the maximum availability of the data and resources produced to allow for investigation and exploitation by others. PI has excellent track record in communicating the output of his research to a broad audience and experienced at writing scientific articles. Data generated in this proposal will be made publicly available via open access publishing where it does not interfere with potential exploitation, as well as via presentations at various national/international meetings. In addition, Imperial College has an extensive series of Open Days and Summer Schemes for schools organized by the Outreach department (imperial.ac.uk/outreach) where PI can regularly presenting his research outcomes. Postdoctoral researcher will be encouraged to develop his communication skills within both the academic and non-academic community in order to better understand the wider impact of his research.
Imperial Innovations Group provides technology commercialisation services and technology development funding for Imperial College London academic staff. We will use Imperial Innovations to engage with industrial partners concerning any commercialization of our research. This will ensure rapid translation of cutting-edge fundamental discoveries into practical solutions and assist global improvement of agriculture. The PI will oversee the impact activities and will seek the assistance of other project members as well as expert staff at The Imperial College, when necessary. Relevant offices at Imperial College will be involved whenever impact activities include technology transfer or outreach/press releases.
 
Description Plants and animals can digest and recycle parts of their own cells in order to survive stress and starvation by a process known as autophagy. Autophagy also contributes to immunity in plants and animals. We discovered that pathogens have evolved a novel mechanism to counteract the plant's autophagy-related defences. We showed that the Irish famine pathogen Phytophthora infestans secretes a protein inside the host cells to manipulate autophagy for its own benefit. Using this pathogen secreted protein as a molecular tool, we recently discovered new host molecules that are required for regulation of autophagy in plants and our current project focused on identifying the roles of these components in autophagy and plant immunity. In a follow-up work, we demonstrate that the autophagy pathway is diverted towards sites where the microbial-pathogens try to penetrate the plant cells.
Exploitation Route Autophagy is a conserved process shred by both plants and animals. It is implicated in many processes including ageing, cancer, cellular starvation etc. Autophagy also contributes to immunity in both systems. Future work could explore whether other secreted-proteins from disease-causing microbes work in a similar way. The pathogen-secreted protein we identified that stimulates autophagy can be used as an inducer of autophagy in different systems to study autophagy and its role in infectious diseases as well as other autophagy related processes. Furthermore, we have generated new information in molecular mechanisms of defence related autophagy and generated molecular tools that might be useful to a wider community of scientists working with other plant pathogens.
Sectors Agriculture, Food and Drink,Environment,Pharmaceuticals and Medical Biotechnology

URL http://elifesciences.org/content/5/e10856v2
 
Description BBSRC IAA Imperial College London
Amount £300,000 (GBP)
Funding ID BB/S506667/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 03/2018 
End 03/2021
 
Description BBSRC Impact Acceleration Award
Amount £80,000 (GBP)
Funding ID BB/GCRF-IAA/17/10 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 09/2017 
End 02/2018
 
Description Global Challenges Research Fund initiative (BBSRC GCRF IAA)
Amount £37,000 (GBP)
Funding ID BB/GCRF-IAA/10 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 12/2016 
End 04/2017
 
Description Yellow rust effector project collaboration with Turkey 
Organisation Middle East Technical University
Country Turkey 
Sector Academic/University 
PI Contribution In this project, we teamed-up with Prof. Akkaya's group (METU, Turkey) to study the functions of yellow rust pathogen of wheat using a non-host heterologous system (Nicotiana benthamiana plants). We provide insights into the poorly understood process of non-host disease resistance in plants. We discovered that some virulence factors secreted by fungal pathogens, which typically act to subvert immune responses of their adapted hosts, can and activate immunity in non-host plant genotypes (Dagvadorj et al., Sci Rep., 2017). We hosted the leading author (B. Dagvadorj) from Turkey in my lab and my team contributed to this work by carrying out cell biology and pathogen infection assays.
Collaborator Contribution Our partners carried out tasks including cloning of the virulence factors, gene silencing, and biochemical assays.
Impact doi:10.1038/s41598-017-01100-z
Start Year 2015
 
Description 36th New Phytologist meeting (Munich) on cell biology at the plant-microbe interface 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact The Activity was to disseminate our findings on reprogramming of host cell processes by plant pathogens
Year(s) Of Engagement Activity 2015
 
Description 3rd Autophagy UK Network Meeting 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Postgraduate students
Results and Impact I gave a talk to disseminate our findings funded by BSBRC.
Year(s) Of Engagement Activity 2017
URL http://autophagy.uk/2017-london-meeting-2/
 
Description 4th Annual and Final Conference of the Sustain COST Action 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact To disseminate findings of our ongoing BBSRC funded research
Year(s) Of Engagement Activity 2017
URL https://www.cost-sustain.org/Events-and-meetings/4th-Annual-and-Final-Conference-of-the-Sustain-COST...
 
Description B-DEBATE 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact This was an international meeting on plant stress entitled, "When Development meets Stress" which took place in CRAG, Barcelona, Spain.
Year(s) Of Engagement Activity 2018
URL https://www.bdebate.org/en/forum/when-development-meets-stress-understanding-developmental-reprogram...
 
Description BSPP presidential meeting 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact This was a conference organised by the British Society of Plant Pathology on Imaging plant-microbe interactions.
Year(s) Of Engagement Activity 2018
URL http://www.bspp.org.uk/meetings/bspppres2018.php
 
Description Biology Seminar at GMI - Gregor Mendel Institute (GMI) of Molecular Plant Biology, Austria, Vienna 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact I have shared our BBSRC funded reseach outcomes with an audience of postgraduate students and group leaders working in one of the world leading institutes (GMI vienna). My talk covered topics on how pathogens manipulate plant cellular processes for their own benefit. The activity was useful for establishing future collaborations and stimulating scientific interest.
Year(s) Of Engagement Activity 2017
URL http://seminars.viennabiocenter.org/seminars.php?display=seminar/2836&kal_monat=20
 
Description COST Action Plant Autophagy meeting 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact Dissemination of our BBSRC funded research on modulation of plant autophagy by the oomycete pathogens.
Year(s) Of Engagement Activity 2017
URL http://cost-transautophagy.eu/wp-content/uploads/2016/11/Report-WG3-Meeting-Madrid-March2017Pilar_Ca...
 
Description Imperial Fringe: Food for tomorrow 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact The Imperial Fringe is a series of free evening public events that explore the livelier side of science.
They provide a unique opportunity to share the wonder and importance of our research with the public,
the College and the wider College community. It also provides an opportunity to create links with new
research groups, collaborate with other institutes, and try out new ways to present research to new and
diverse audiences
Around 5-700 people attend each Fringe event, made up of families and general members of the public as well as
staff, students, alumni and neighbours, including local schools and workers.
We as a grop introduced our research on how to improve crop resistance against pathogens
Year(s) Of Engagement Activity 2016
URL http://www3.imperial.ac.uk/newsandeventspggrp/imperialcollege/newssummary/news_26-2-2016-10-16-55
 
Description Oomycete Molecular Genetics Network Annual Meeting 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact The purpose for attending this meeting was to disseminate our findings from our ongoing BBSRC grant
Year(s) Of Engagement Activity 2016
URL https://omgn.org/about/
 
Description The International Society for Molecular Plant-Microbe Interactions (IS-MPMI) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact The purpose was to disseminate the findings of our ongoing BBSRC grant
Year(s) Of Engagement Activity 2016
URL http://www.ismpmi.org/Congress/2016/Pages/default.aspx