Life Sciences Research for Lifelong Health

Nicholas Ktistakis

Research Summary

Autophagy (from the Greek self-eating) is a cellular mechanism which generates nutrients for the cell, primarily during times of starvation. Autophagy is also used to eliminate cell material that becomes damaged, leading to a periodic clean-up of the cell interior. Although it is a response by single cells, it is also very important for the health of an organism.

When autophagy is suppressed cells exhibit signs of oxidative damage because their dysfunctional mitochondria cannot be removed and continue to produce reactive oxygen species. Similarly, suppression of autophagy causes the build-up of mutant proteins that cause neurodegenerative disorders.

Autophagy is also critical for the neonatal period: animals which lack autophagy die soon after birth because they cannot generate nutrients during that time. Finally, autophagy is critical for the extension of lifespan in all organisms studied, and is therefore a significant factor that affects healthy ageing. The pathway of autophagy starts when a novel double membrane vesicle called an autophagosome is formed in the cell interior.

We have shown that one of the signals for formation of autophagosomes is the synthesis of a lipid called PI3P which leads to formation of omegasomes. These are membrane extensions of the endoplasmic reticulum, from which some autophagosomes emerge. We are studying exactly how this happens, both in terms of signals and of how the intermediate structures eventually lead to an autophagosome.

Latest Publications

Phospholipase D activity couples plasma membrane endocytosis with retromer dependent recycling.
Thakur R, Panda A, Coessens E, Raj N, Yadav S, Balakrishnan S, Zhang Q, Georgiev P, Basak B, Pasricha R, Wakelam MJ, Ktistakis NT, Padinjat R

During illumination, the light sensitive plasma membrane (rhabdomere) of Drosophila photoreceptors undergoes turnover with consequent changes in size and composition. However the mechanism by which illumination is coupled to rhabdomere turnover remains unclear. We find that photoreceptors contain a light-dependent phospholipase D (PLD) activity. During illumination, loss of PLD resulted in an enhanced reduction in rhabdomere size, accumulation of Rab7 positive, rhodopsin1-containing vesicles (RLVs) in the cell body and reduced rhodopsin protein. These phenotypes were associated with reduced levels of phosphatidic acid, the product of PLD activity and were rescued by reconstitution with catalytically active PLD. In wild type photoreceptors, during illumination, enhanced PLD activity was sufficient to clear RLVs from the cell body by a process dependent on Arf1-GTP levels and retromer complex function. Thus, during illumination, PLD activity couples endocytosis of RLVs with their recycling to the plasma membrane thus maintaining plasma membrane size and composition.

+ View Abstract

eLife, 5, 2050-084X, , 2016

PMID: 27848911

Assembly of early machinery for autophagy induction: novel insights from high resolution microscopy.
Ktistakis NT, Walker SA, Karanasios E

Oncotarget, , 1949-2553, , 2016

PMID: 27829241

Dynamics of mTORC1 activation in response to amino acids.
Manifava M, Smith M, Rotondo S, Walker S, Niewczas I, Zoncu R, Clark J, Ktistakis NT

Amino acids are essential activators of mTORC1 via a complex containing RAG GTPases, RAGULATOR and the vacuolar ATPase. Sensing of amino acids causes translocation of mTORC1 to lysosomes, an obligate step for activation. To examine the spatial and temporal dynamics of this translocation, we used live imaging of the mTORC1 component RAPTOR and a cell permeant fluorescent analogue of di-leucine methyl ester. Translocation to lysosomes is a transient event, occurring within 2 min of aa addition and peaking within 5 min. It is temporally coupled with fluorescent leucine appearance in lysosomes and is sustained in comparison to aa stimulation. Sestrin2 and the vacuolar ATPase are negative and positive regulators of mTORC1 activity in our experimental system. Of note, phosphorylation of canonical mTORC1 targets is delayed compared to lysosomal translocation suggesting a dynamic and transient passage of mTORC1 from the lysosomal surface before targetting its substrates elsewhere.

+ View Abstract

eLife, 5, 2050-084X, , 2016

PMID: 27725083

 

Group Members

Latest Publications

Assembly of early machinery for autophagy induction: novel insights from high resolution microscopy.

Ktistakis NT, Walker SA, Karanasios E

Oncotarget
1949-2553: (2016)

PMID: 27829241

Dynamics of mTORC1 activation in response to amino acids.

Manifava M, Smith M, Rotondo S

eLife
5 2050-084X: (2016)

PMID: 27725083

Characterization of Atg38 and NRBF2, a fifth subunit of the autophagic Vps34/PIK3C3 complex.

Ohashi Y, Soler N, García Ortegón M

Autophagy
1554-8635:0 (2016)

PMID: 27630019

Autophagy initiation by ULK complex assembly on ER tubulovesicular regions marked by ATG9 vesicles.

Karanasios E, Walker SA, Okkenhaug H

Nature communications
7 2041-1723:12420 (2016)

PMID: 27510922

Digesting the Expanding Mechanisms of Autophagy.

Ktistakis NT, Tooze SA

Trends in cell biology
1879-3088: (2016)

PMID: 27050762

Guidelines for the use and interpretation of assays for monitoring autophagy (3rd edition).

Klionsky DJ, Abdelmohsen K, Abe A

Autophagy
12 1554-8635:1-222 (2016)

PMID: 26799652

Structure and flexibility of the endosomal Vps34 complex reveals the basis of its function on membranes.

Rostislavleva K, Soler N, Ohashi Y

Science (New York, N.Y.)
350 1095-9203:aac7365 (2015)

PMID: 26450213

ERES: sites for autophagosome biogenesis and maturation?

Sanchez-Wandelmer J, Ktistakis NT, Reggiori F

Journal of cell science
128 1477-9137:185-192 (2015)

PMID: 25568152

Live-cell imaging for the assessment of the dynamics of autophagosome formation: Focus on early steps.

Karanasios E, Ktistakis NT

Methods (San Diego, Calif.)
1095-9130: (2014)

PMID: 25498007

Dynamics of autophagosome formation: a pulse and a sequence of waves.

Ktistakis NT, Karanasios E, Manifava M

Biochemical Society transactions
42 1470-8752:1389-95 (2014)

PMID: 25233420