Functional dissection of Condensin and Cohesin in atypical mitosis and meiosis in Plasmodium
Lead Research Organisation:
University of Nottingham
Department Name: School of Life Sciences
Abstract
Plasmodium is the causative agent of malaria and is responsible for about 584,000 human deaths annually, mostly children under the age of 5. The increasing resistance of the parasite to existing drugs and the lack of an efficient vaccine represent the main obstacles to combat this burden. Therefore, there is an urgent need for the development of innovative therapeutics. Plasmodium has a complex life cycle with diverse host environments, changes in cell shape, size and motility and an atypical mode of cell proliferation. The symptoms of the disease are manifest when malaria parasites invade host red blood cells, wherein the parasite divides and multiplies many times (endomitosis), eventually leading to destruction of the host cell. Some of the parasite cells may cease to divide and they become precursor sex cells (male and female gametocytes). When a female mosquito bites an infected person they ingest parasites along with the blood and this acts as a trigger to activate the precursor sex cells within the mosquito gut. The male gametocytes undergo rapid cell division (endoreduplication) to produce eight male gametes, which then fertilise the female gametes and the parasite life cycle continues in the mosquito gut. After further development and multiplication the parasite moves to the mosquito's salivary glands and is passed again to a new human host when the mosquito feeds.
The process of Plasmodium cell division in host red blood cells, and during sexual development in the mosquito vector is very different, but both are essential for parasite growth and transmission. If any of these stages are blocked then both proliferation and transmission are curtailed. Therefore, it is critically important to understand how the parasite multiplies and divides at these stages so that we can devise ways to interfere with them by developing appropriate drugs.
The molecules that control these types of atypical cell division in the parasite are very poorly understood. The project proposed here is to study the role of two important protein complexes: condensin and cohesin. Although these complexes are known to be involved in cell division in many model systems, there is no knowledge of how they function during malaria parasite cell division and proliferation. We can start by using the knowledge gathered in model systems and applying this to the malaria parasite. For example, we have recently identified one such protein (CDC20) in the malaria parasite and showed that it has a key role in regulating male gamete formation. In preliminary work showing that our approach is feasible, we have obtained evidence for the presence of condensin and cohesin in the parasite multiplying within the red blood cell, in the male sex cell and in the meiotic cell (an ookinete). Recent advances in analysing genes in malaria allow us to study the function of these molecules. For example, by taking away the gene, we can see what happens when the proteins are no longer made, and if they are tagged experimentally with a fluorescent marker we can see under the microscope where they are located in the parasite. We will also study how these molecules interact with other proteins during various processes such as chromosome segregation and cytokinesis, to name two. We can also purify these protein complexes and study their interactions using mass spectroscopy-based approaches. Therefore, we are now in a position where we can explore how cell division in malaria is controlled by these complexes and we can study how these molecules regulate different stages of cell division.
Our study may identify molecular targets important in parasite cell division in host red blood cells and in the cells dividing within the mosquito vector. As a result, we may identify potential targets for drugs or vaccines that could in the future be used to block parasite replication in the blood and transmission from one individual to another through the mosquito.
The process of Plasmodium cell division in host red blood cells, and during sexual development in the mosquito vector is very different, but both are essential for parasite growth and transmission. If any of these stages are blocked then both proliferation and transmission are curtailed. Therefore, it is critically important to understand how the parasite multiplies and divides at these stages so that we can devise ways to interfere with them by developing appropriate drugs.
The molecules that control these types of atypical cell division in the parasite are very poorly understood. The project proposed here is to study the role of two important protein complexes: condensin and cohesin. Although these complexes are known to be involved in cell division in many model systems, there is no knowledge of how they function during malaria parasite cell division and proliferation. We can start by using the knowledge gathered in model systems and applying this to the malaria parasite. For example, we have recently identified one such protein (CDC20) in the malaria parasite and showed that it has a key role in regulating male gamete formation. In preliminary work showing that our approach is feasible, we have obtained evidence for the presence of condensin and cohesin in the parasite multiplying within the red blood cell, in the male sex cell and in the meiotic cell (an ookinete). Recent advances in analysing genes in malaria allow us to study the function of these molecules. For example, by taking away the gene, we can see what happens when the proteins are no longer made, and if they are tagged experimentally with a fluorescent marker we can see under the microscope where they are located in the parasite. We will also study how these molecules interact with other proteins during various processes such as chromosome segregation and cytokinesis, to name two. We can also purify these protein complexes and study their interactions using mass spectroscopy-based approaches. Therefore, we are now in a position where we can explore how cell division in malaria is controlled by these complexes and we can study how these molecules regulate different stages of cell division.
Our study may identify molecular targets important in parasite cell division in host red blood cells and in the cells dividing within the mosquito vector. As a result, we may identify potential targets for drugs or vaccines that could in the future be used to block parasite replication in the blood and transmission from one individual to another through the mosquito.
Technical Summary
Plasmodium is a unicellular parasite and the causative agent of malaria. These parasites have an atypical cell division that differs from that of the host they invade. They divide within red blood cells (schizogony) in a process resembling endomitosis where the nuclear envelope remains intact and a single cell becomes multinucleated. Moreover, during this stage their chromosomes do not condense. The parasite also shows endo-reduplication during male gametocyte differentiation, as well as a short meiotic stage within the mosquito vector. The molecular mechanisms that regulate these atypical cell divisions in Plasmodium are not well studied.
The main aim of this project is to dissect out the molecular mechanisms controlling two important components, Condensin and Cohesin in these types of atypical cell division and establish how they differ from the classical cell division of host cells, ultimately to identify intervention targets.
Condensin and cohesin belong to an important family of proteins that have crucial roles in chromosome dynamics and gene regulation, both in mitosis and meiosis in most eukaryotes. This has been widely studied in some cells, yet nothing is known of their localisation, function, regulation or the complexes associated with them during atypical cell division in Plasmodium.
Complementary approaches will be used to achieve three main objectives, using Plasmodium berghei as a model system: (1) To identify the subcellular location of condensin and cohesin at various stages of parasite cell division. 2) To study the function and transcriptional regulation of cohesin and condensin during parasite cell division and (3) To characterise the cohesion and condensin complexes using proteomic approaches.
We will use reverse genetics, state of the art cell biology methods with fluorescence, confocal, and electron microscopy, and protein biochemistry with proteomics to achieve our goals.
The main aim of this project is to dissect out the molecular mechanisms controlling two important components, Condensin and Cohesin in these types of atypical cell division and establish how they differ from the classical cell division of host cells, ultimately to identify intervention targets.
Condensin and cohesin belong to an important family of proteins that have crucial roles in chromosome dynamics and gene regulation, both in mitosis and meiosis in most eukaryotes. This has been widely studied in some cells, yet nothing is known of their localisation, function, regulation or the complexes associated with them during atypical cell division in Plasmodium.
Complementary approaches will be used to achieve three main objectives, using Plasmodium berghei as a model system: (1) To identify the subcellular location of condensin and cohesin at various stages of parasite cell division. 2) To study the function and transcriptional regulation of cohesin and condensin during parasite cell division and (3) To characterise the cohesion and condensin complexes using proteomic approaches.
We will use reverse genetics, state of the art cell biology methods with fluorescence, confocal, and electron microscopy, and protein biochemistry with proteomics to achieve our goals.
Planned Impact
This proposal fits well within the strategic aim of the MRC towards Global health, Research to people and picking research that delivers. We plan to address the role of Condensin and Cohesin in the core stages of atypical cell division during the malaria parasite life cycle, in both mammalian host and mosquito vector. To achieve this we have chosen an experimentally tractable rodent malaria parasite, Plasmodium berghei, to enable us to examine these atypical stages during both the sexual and asexual stages of the life cycle.
Cell division is a prerequisite for any organism to proliferate and develop during its life cycle. To understand how chromosomes are duplicated and segregated, and how cell division occurs in a proper manner are basic questions in cell biology. Although various systems have been used to provide our understanding of these processes, including studies in yeast, human and plant cells, very little is known about such processes and their regulation in unicellular parasitic protozoans like Plasmodium. Malaria is the third largest global health problem caused by a single infectious agent after HIV and TB, affecting millions of people, and resulting in one million deaths annually.
Understanding the molecular mechanisms controlling multiplication of the parasite following atypical cell division is crucial to help develop intervention and control strategies. Our research will give us a better understanding of the molecules that are involved in these processes of chromosome biology and nuclear division; thereby controlling cell division and hence the identification of probable targets for intervention both to treat the disease state and to prevent transmission.
This research will provide further fundamental knowledge on the role of these conserved condensin and cohesin complexes whilst the parasite develops and proliferates using a very different mode of cell division and closed mitosis, compared with the classical processes observed in mammalian cells. It will be informative for both pure research in cell and developmental biology, and for understanding the evolution of these molecules and their function.
Our multifaceted approach will employ technologies that are at the cutting edge of research in the areas of parasite genetics, cell biology, protein chemistry and proteomics. This study will further strengthen the research potential of the malaria and wider community studying other parasites. Most of the resources generated during the course of this project will be deposited in research databases such as PlasmoDB or the RMgm database. The research on malaria parasites will hopefully also impact on our understanding of cell division in other parasites such as Trypanosoma, Giardia and others.
Cell division is a prerequisite for any organism to proliferate and develop during its life cycle. To understand how chromosomes are duplicated and segregated, and how cell division occurs in a proper manner are basic questions in cell biology. Although various systems have been used to provide our understanding of these processes, including studies in yeast, human and plant cells, very little is known about such processes and their regulation in unicellular parasitic protozoans like Plasmodium. Malaria is the third largest global health problem caused by a single infectious agent after HIV and TB, affecting millions of people, and resulting in one million deaths annually.
Understanding the molecular mechanisms controlling multiplication of the parasite following atypical cell division is crucial to help develop intervention and control strategies. Our research will give us a better understanding of the molecules that are involved in these processes of chromosome biology and nuclear division; thereby controlling cell division and hence the identification of probable targets for intervention both to treat the disease state and to prevent transmission.
This research will provide further fundamental knowledge on the role of these conserved condensin and cohesin complexes whilst the parasite develops and proliferates using a very different mode of cell division and closed mitosis, compared with the classical processes observed in mammalian cells. It will be informative for both pure research in cell and developmental biology, and for understanding the evolution of these molecules and their function.
Our multifaceted approach will employ technologies that are at the cutting edge of research in the areas of parasite genetics, cell biology, protein chemistry and proteomics. This study will further strengthen the research potential of the malaria and wider community studying other parasites. Most of the resources generated during the course of this project will be deposited in research databases such as PlasmoDB or the RMgm database. The research on malaria parasites will hopefully also impact on our understanding of cell division in other parasites such as Trypanosoma, Giardia and others.
Organisations
- University of Nottingham (Lead Research Organisation)
- University of Geneva (Collaboration)
- University College London (Collaboration)
- University of California, Riverside (Collaboration)
- UNIVERSITY OF LEICESTER (Collaboration)
- University of Groningen (Collaboration)
- Birkbeck, University of London (Collaboration)
Publications
Bunnik EM
(2019)
Comparative 3D genome organization in apicomplexan parasites.
in Proceedings of the National Academy of Sciences of the United States of America
Green JL
(2017)
Compositional and expression analyses of the glideosome during the Plasmodium life cycle reveal an additional myosin light chain required for maximum motility.
in The Journal of biological chemistry
Guttery DS
(2020)
Plasmodium DEH is ER-localized and crucial for oocyst mitotic division during malaria transmission.
in Life science alliance
Jacot D
(2016)
An Apicomplexan Actin-Binding Protein Serves as a Connector and Lipid Sensor to Coordinate Motility and Invasion.
in Cell host & microbe
Mancio-Silva L
(2017)
Nutrient sensing modulates malaria parasite virulence.
in Nature
Pandey R
(2018)
High throughput in silico identification and characterization of Plasmodium falciparum PRL phosphatase inhibitors.
in Journal of biomolecular structure & dynamics
Rea E
(2017)
Plasmodium Peekaboo: PK4 Mediates Parasite Latency.
in Cell host & microbe
Rea E
(2018)
Sex in Plasmodium falciparum: Silence Play between GDV1 and HP1.
in Trends in parasitology
Roques M
(2019)
Plasmodium centrin PbCEN-4 localizes to the putative MTOC and is dispensable for malaria parasite proliferation.
in Biology open
Saini E
(2017)
Photosensitized INA-Labelled protein 1 (PhIL1) is novel component of the inner membrane complex and is required for Plasmodium parasite development.
in Scientific reports
Wall RJ
(2018)
Author Correction: Plasmodium APC3 mediates chromosome condensation and cytokinesis during atypical mitosis in male gametogenesis.
in Scientific reports
Wall RJ
(2018)
Plasmodium APC3 mediates chromosome condensation and cytokinesis during atypical mitosis in male gametogenesis.
in Scientific reports
Wall RJ
(2019)
Systematic analysis of Plasmodium myosins reveals differential expression, localisation, and function in invasive and proliferative parasite stages.
in Cellular microbiology
Zeeshan M
(2020)
Plasmodium berghei Kinesin-5 Associates With the Spindle Apparatus During Cell Division and Is Important for Efficient Production of Infectious Sporozoites
in Frontiers in Cellular and Infection Microbiology
Zeeshan M
(2019)
Kinesin-8B controls basal body function and flagellum formation and is key to malaria transmission.
in Life science alliance
Zeeshan M
(2020)
Real-time dynamics of Plasmodium NDC80 reveals unusual modes of chromosome segregation during parasite proliferation.
in Journal of cell science
Zeeshan M
(2021)
Protein phosphatase 1 regulates atypical mitotic and meiotic division in Plasmodium sexual stages
in Communications Biology
Description | We observed the atypical complex of condensin in two stages of parasite proliferation. THese finding are now finding in an open access journal Cell Reports |
Exploitation Route | Thse finding have now led to new research to indenfy the partners of these complex components and identify their function in two stages of parasite proliferation |
Sectors | Education Manufacturing including Industrial Biotechology |
URL | https://www.cell.com/cell-reports/pdf/S2211-1247(20)30048-6.pdf |
Title | Live cell imagine for anaysing spindle dynamics |
Description | T study rapid spindle dynamics and basal body formation in male gametocyte of malaria parasite |
Type Of Material | Model of mechanisms or symptoms - in vitro |
Year Produced | 2019 |
Provided To Others? | Yes |
Impact | This has been shared with many lab nows |
Title | Protein interaction map |
Description | We have developed the protocol for parasite gametocyte purified using GFP transgenic lines and studying the protein interaction map. THis is mainly to understand the chrosome segregation in this divergent parasite and could be used for blocking cell proliferation |
Type Of Material | Technology assay or reagent |
Year Produced | 2020 |
Provided To Others? | Yes |
Impact | It is useful to study the kinetochore and spindle dynamics of the unique parasite cell that is necessary for male gamete formation and can be information for transmission blocking. |
Title | Superresolution microscopy |
Description | We have developed the superresolution technology for analysing the sexual stages of parasites especially for the spindle and kinetochore biology |
Type Of Material | Technology assay or reagent |
Year Produced | 2022 |
Provided To Others? | Yes |
Impact | This is important as the kinetochore biology was not understood in malaria parasite |
URL | https://www.life-science-alliance.org/content/5/9/e202101329 |
Title | transgenic parasite lines |
Description | THese are transgenic parasite lines for functional of kinesin in Plasmodium |
Type Of Material | Technology assay or reagent |
Provided To Others? | No |
Impact | No |
Title | PRIDE |
Description | This is the research data base for Mass spec data produced during the protein interaction studies |
Type Of Material | Database/Collection of data |
Year Produced | 2020 |
Provided To Others? | Yes |
Impact | It is open data base and can be used by anyone |
Description | Anaphase promoting complex |
Organisation | University College London |
Department | UCL Cancer Institute |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | This collaboration is with Prof Hiro Yamano to understand the anaphase promoting complex |
Collaborator Contribution | Prf Hiro Yamano has contributed immensly for undersyanding of cell division in yeat and mammalian system and provided us with various resources. |
Impact | We have published one paper together in Plos Pathogenes in 2015 |
Start Year | 2014 |
Description | Atypical celldivision in male gametogony |
Organisation | University of Geneva |
Country | Switzerland |
Sector | Academic/University |
PI Contribution | We have been contributing the resources in the lab in terms of tagged and knockdown parasites to his group |
Collaborator Contribution | He has been able to help us with phosphoproteome |
Impact | Nat Commun. 2023 Sep 13;14(1):5652. doi: 10.1038/s41467-023-41395-3. PMID: 37704606 PLoS Biol. 2022 Jul 28;20(7):e3001704. doi: 10.1371/journal.pbio.3001704. eCollection 2022 Jul.PMID: 35900985 |
Start Year | 2018 |
Description | Bioinformatic and evolutionary Cell Biology |
Organisation | University of Groningen |
Department | Groningen Biomolecular Sciences and Biotechnology Institute (GBB) |
Country | Netherlands |
Sector | Academic/University |
PI Contribution | Evolutionary Biology of divergent kinases, motor protein and condensin |
Collaborator Contribution | Evolutionary Biology of divergent kinases, motor protein and condensin |
Impact | Nat Commun. 2023 Sep 13;14(1):5652. doi: 10.1038/s41467-023-41395-3.PMID: 37704606 Trends Parasitol. 2023 Oct;39(10):812-821. doi: 10.1016/j.pt.2023.07.002. Epub 2023 Aug 2.PMID: 37541799 |
Start Year | 2019 |
Description | Cell Division |
Organisation | University College London |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | We collaborate with Prof Hiro Yamano for cyclin and APC project |
Collaborator Contribution | He is an expert in APC in Xenopus and yeast system |
Impact | mutidisciplinary |
Start Year | 2012 |
Description | Condensin in Plasmodium |
Organisation | University of California, Riverside |
Country | United States |
Sector | Academic/University |
PI Contribution | We have provided the transgenic parasite and ptoteomics approaches for the condensin copmplex subunit and their charecterisation during parasite life cycle mainly in the mosquito stages |
Collaborator Contribution | Prof Karine Le Roch has provided the support fr genome wide approaches like Chipseq and RNa seq for this project |
Impact | We have published four paper together in Cell Reports and Plos Pathogens and Nature Communication . This collaboration is multidiscplinary. PMID: 32501284;J Cell Sci. 2020 Jun 30;134(5):jcs245753. doi: 10.1242/jcs.245753. PMID: 32049018; Cell Rep. 2020 Feb 11;30(6):1883-1897.e6. doi: 10.1016/j.celrep.2020.01.033. PMID: 31600347; PLoS Pathog. 2019 Oct 10;15(10):e1008048. doi: 10.1371/journal.ppat.1008048. eCollection 2019 Oct. PMID: 37704606 Nat Commun. 2023 Sep 13;14(1):5652. |
Start Year | 2018 |
Description | Kinesin in Plasmodium |
Organisation | Birkbeck, University of London |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | I am collaborating with Prof Carolyn Moores who is an expert on Kinesin in mammalian system and is a reputed Structural Biologist |
Collaborator Contribution | She will provide us the biochemistry of of the Kinesin 5 and Kininesin 13 |
Impact | We will be writing the paper at the end of the year. |
Start Year | 2014 |
Description | Kinesin in Plasmodium |
Organisation | Birkbeck, University of London |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | We colloborate with Prof Carolyn Moores who works with biochemistry and molecular dissection of kinesin motor using CryoEm |
Collaborator Contribution | Prof Carolyn will provide the biochemistry and structual side of the project to this study |
Impact | PLoS Biol. 2022 Jul 28;20(7):e3001704. doi: 10.1371/journal.pbio.3001704. eCollection 2022 Jul. PMID: 35900985 Nat Commun. 2022 Nov 16;13(1):6988. doi: 10.1038/s41467-022-34710-x.PMID: 36384964 |
Start Year | 2015 |
Description | Phosphorylation |
Organisation | University of Leicester |
Department | Department of Cell Physiology and Pharmacology |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | significant intellectual input |
Collaborator Contribution | To provide technical help with Global Phosphorylationtraining of staff |
Impact | Two manuscript in preparation |
Start Year | 2010 |
Description | Phosphorylation |
Organisation | University of Leicester |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | significant intellectual input |
Collaborator Contribution | To provide technical help with Global Phosphorylationtraining of staff |
Impact | Two manuscript in preparation |
Start Year | 2010 |
Description | A seminar at Mredical Erasmus University |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Study participants or study members |
Results and Impact | This helped me to meet scientist who are working with Cohesin in mammalian system. |
Year(s) Of Engagement Activity | 2017 |
Description | Interview with BBC Radio nottingham |
Form Of Engagement Activity | A press release, press conference or response to a media enquiry/interview |
Part Of Official Scheme? | No |
Geographic Reach | Regional |
Primary Audience | Media (as a channel to the public) |
Results and Impact | This was radio interview where I was interviewed about our work malaria cell division and how this research help in wider context. |
Year(s) Of Engagement Activity | 2019 |
Description | Organised a Midland Cel Cycle Club 2017 for the East Midland |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | Regional |
Primary Audience | Postgraduate students |
Results and Impact | This meeting made good network of scientist working in the area of Cel Cycle and Cytoskeleton in various model system like human, drosophila, yeat parasites etc. |
Year(s) Of Engagement Activity | 2017 |
Description | Organising the London Molecular Parasitology Club |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | National |
Primary Audience | Study participants or study members |
Results and Impact | Three meeting of London Molecular Parasitology are organised every year. The speakers range from any where in Uk or Europe. It benefots the scientist working in London and adjoining areas. |
Year(s) Of Engagement Activity | 2007,2008,2009,2010,2011,2012,2013,2014,2015,2016,2017,2018 |
Description | School student demonstration |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | Regional |
Primary Audience | Schools |
Results and Impact | This was mainly organised for school student around Nottinghamshire to make them aware of research studies and mainly to teach them about cell biology and microscopy detection of malaria parasite |
Year(s) Of Engagement Activity | 2019 |
Description | press release on role of condensin |
Form Of Engagement Activity | A press release, press conference or response to a media enquiry/interview |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Media (as a channel to the public) |
Results and Impact | This was the press release nby University of Nottingham and this was picked up in many countries like India, Belgium etc. |
Year(s) Of Engagement Activity | 2020 |