Characterisation of the PI3P-dependent signalling network responsible for nutrient sensing and autophagy
Lead Research Organisation:
Babraham Institute
Department Name: Signalling
Abstract
One of the most important requirements for healthy cellular growth is the ability to sense and respond to extracellular nutrients. When nutrient supply is plentiful, cells need only to fine tune their anabolic and catabolic rates so that their energy requirements match the available resources. When nutrients become limiting or are completely unavailable, a much stronger course of action is required to allow the cells to survive. During such times of nutrient withdrawal, the cells respond by initiating autophagy, a degradative pathway that allows the breakdown of intracellular proteins into amino acids that can be used subsequently for new protein synthesis or energy generation. An important question in autophagy concerns the mechanisms by which cells sense their extracellular nutrient content. Recent work from many groups has indicated that a small lipid molecule termed phosphatidylinositol 3 phosphate (PI3P) is an important signal for nutrient sensing. Our own work also indicates that autophagy is induced partially as a result of the formation of PI3P in specialised membrane compartments called omegasomes. Therefore, the mechanisms and signals that generate PI3P early during autophagy are likely to provide important information on the control of autophagy by nutrient sensing. The aim of this grant is to identify all human genes that are involved in this pathway. We plan to systematically silence all known human genes and then look at the effect that this will have in formation of PI3P during autophagy. By identifying all such genes we will be able to construct a wiring diagram of the cellular pathways that are implicated in nutrient sensing and respond during nutrient limitation by the induction of autophagy.
Technical Summary
During nutrient withdrawal, cells respond by initiating autophagy, a degradative pathway that allows the breakdown of intracellular proteins into amino acids that can be used subsequently for new protein synthesis or ATP generation. In addition to this role in nutrient sensing, autophagy is important for elimination of unneeded organelles, for the defence against pathogens, for the pathogenesis of various diseases and for healthy ageing. Our recent work has suggested that at least some autophagosomes (double membrane vesicles mediating autophagy) are formed in novel membrane specializations termed omegasomes which are connected to the endoplasmic reticulum and are enriched in a phosphatidylinositol 3-phosphate (PI3P). This is consistent with earlier work showing that formation of PI3P is essential for the induction of autophagy. However, the signalling pathways leading from nutrient sensing to PI3P formation and autophagy induction are still unknown. The aim of this grant is to identify the main gene products whose activity is important for the PI3P-dependent induction of autophagy. We will use an siRNA screen of all known human genes, to identify those whose down-regulation (a) leads to omegasome formation in normal nutrient conditions or (b) inhibits amino acid starvation-induced formation of omegasomes. Based on our preliminary data we expect the following classes of molecules to be identified: (1) Those involved in amino acid sensing [AA sensor(s)] (2) Those involved in signalling to calcium following amino acid sensing (3) Those involved in signalling from calcium to Vps34 (4) Those involved in AA-dependent mTOR activation (5) Those involved in omegasome formation, expansion and collapse (5) Those involved in termination of PI3P signal (6) Those involved in feedback inhibition of autophagosome biogenesis. The ultimate aim of this grant is to construct a signalling diagram leading from nutrient sensing to PI3P formation and autophagy induction.
People |
ORCID iD |
| Nicholas Ktistakis (Principal Investigator) |
Publications
Burman C
(2010)
Autophagosome formation in mammalian cells.
in Seminars in immunopathology
Cottam EM
(2011)
Coronavirus nsp6 proteins generate autophagosomes from the endoplasmic reticulum via an omegasome intermediate.
in Autophagy
Deretic V
(2019)
Autophagy, Inflammation, and Metabolism (AIM) Center in its second year.
in Autophagy
Fraser J
(2019)
Targeting of early endosomes by autophagy facilitates EGFR recycling and signalling.
in EMBO reports
Galluzzi L
(2017)
Molecular definitions of autophagy and related processes.
in The EMBO journal
Karanasios E
(2013)
Dynamic association of the ULK1 complex with omegasomes during autophagy induction.
in Journal of cell science
Karanasios E
(2014)
Imaging autophagy.
in Current protocols in cytometry
Kishi-Itakura C
(2020)
Ultrastructural insights into pathogen clearance by autophagy.
in Traffic (Copenhagen, Denmark)
Ktistakis NT
(2012)
How phosphoinositide 3-phosphate controls growth downstream of amino acids and autophagy downstream of amino acid withdrawal.
in Biochemical Society transactions
Ktistakis NT
(2019)
Who plays the ferryman: ATG2 channels lipids into the forming autophagosome.
in The Journal of cell biology
Ktistakis NT
(2015)
Monitoring the Localization of MAP1LC3B by Indirect Immunofluorescence.
in Cold Spring Harbor protocols
Ktistakis NT
(2017)
In praise of M. Anselmier who first used the term "autophagie" in 1859.
in Autophagy
Ktistakis NT
(2016)
Digesting the Expanding Mechanisms of Autophagy.
in Trends in cell biology
Ktistakis NT
(2020)
ER platforms mediating autophagosome generation.
in Biochimica et biophysica acta. Molecular and cell biology of lipids
Ktistakis NT
(2010)
Lipid signaling and homeostasis: PA- is better than PA-H, but what about those PIPs?
in Science signaling
Ktistakis NT
(2013)
PIPing on lysosome tubes.
in The EMBO journal
Matsunaga K
(2010)
Autophagy requires endoplasmic reticulum targeting of the PI3-kinase complex via Atg14L.
in The Journal of cell biology
Musiwaro P
(2013)
Characteristics and requirements of basal autophagy in HEK 293 cells.
in Autophagy
Proikas-Cezanne T
(2020)
Editorial: Autophagy and Ageing: Ideas, Methods, Molecules.
in Frontiers in cell and developmental biology
Roberts R
(2013)
Omegasomes: PI3P platforms that manufacture autophagosomes.
in Essays in biochemistry
Taguchi-Atarashi N
(2010)
Modulation of local PtdIns3P levels by the PI phosphatase MTMR3 regulates constitutive autophagy.
in Traffic (Copenhagen, Denmark)
Walker S
(2020)
Autophagosome Biogenesis Machinery
in Journal of Molecular Biology
Zachari M
(2019)
Selective Autophagy of Mitochondria on a Ubiquitin-Endoplasmic-Reticulum Platform.
in Developmental cell
| Description | Several genes affecting autophagy responses |
| Exploitation Route | Understand how autophagy is induced |
| Sectors | Chemicals,Healthcare |
| Description | Many scientists are using our reporter cell lines to follow autophagy |
| First Year Of Impact | 2011 |
| Sector | Chemicals,Healthcare |
| Description | Colaborration with Japanese |
| Organisation | University of Tokyo |
| Country | Japan |
| Sector | Academic/University |
| PI Contribution | Paper together with Prof Noboru Mizushima |
| Collaborator Contribution | Paper together with Prof Noboru Mizushima |
| Impact | paper together visit of scientist |
| Start Year | 2011 |
| Description | Roger Williams |
| Organisation | Medical Research Council (MRC) |
| Department | MRC Laboratory of Molecular Biology (LMB) |
| Country | United Kingdom |
| Sector | Academic/University |
| PI Contribution | Exchange of reagents and cell lines |
| Collaborator Contribution | Exchange of reagents and cell lines |
| Impact | Publications and grants |
| Start Year | 2012 |
| Description | Tamotsu |
| Organisation | Osaka University |
| Country | Japan |
| Sector | Academic/University |
| PI Contribution | Papers together |
| Collaborator Contribution | Papers together |
| Impact | Published 2 papers together |
| Start Year | 2009 |
| Title | DFCP1 cell line |
| Description | Reporter cell line for autophagy induction |
| IP Reference | |
| Protection | Protection not required |
| Year Protection Granted | 2010 |
| Licensed | Yes |
| Impact | WE have distributed this cell line to over 40 laboratories in UK, Europe, USA, China, Japan |
| Description | Cambridge Enterprise & Technology Club |
| Form Of Engagement Activity | A talk or presentation |
| Part Of Official Scheme? | No |
| Geographic Reach | Regional |
| Primary Audience | Professional Practitioners |
| Results and Impact | Talk on autophagy and degenerative diseases |
| Year(s) Of Engagement Activity | 2016 |
| URL | http://www.cetc.info/degenerative-diseases/ |
| Description | Cambridge Hellenic Learned Society |
| Form Of Engagement Activity | Participation in an activity, workshop or similar |
| Part Of Official Scheme? | No |
| Geographic Reach | Regional |
| Primary Audience | Professional Practitioners |
| Results and Impact | President of Society of Greek and Cypriot academics in Cambridge, charged with organizing events with well-known speakers in the fields of Science of Humanities and addressed to a lay/educated audience. So far we have organized 7 such events with average attendance of 90 people. |
| Year(s) Of Engagement Activity | 2014,2015,2016 |
| URL | https://camlearnhelsoc.wordpress.com/ |
| Description | Presentation at Instutute Ageing Conference |
| Form Of Engagement Activity | A formal working group, expert panel or dialogue |
| Part Of Official Scheme? | No |
| Geographic Reach | National |
| Primary Audience | Professional Practitioners |
| Results and Impact | Invited talk at Ageing conference, Babraham Institute |
| Year(s) Of Engagement Activity | 2017 |