Close
Close

preLights is supported by…

Related posts

Related preLists

Profiling the surface proteome identifies actionable biology for TSC1 mutant cells beyond mTORC1 signaling

Junnian Wei, Kevin K. Leung, Charles Truillet, Davide Ruggero, James A. Wells, Michael J. Evans

Preprint posted on 1 August 2018 https://www.biorxiv.org/content/early/2018/08/01/382929

Cell surface proteomics reveals new mTORC1-independent functions of TSC1 and suggests new avenues to develop therapies for TSC1-mutant tumours.

Selected by Rob Hynds

Categories: cancer biology, cell biology

Background

Tuberous sclerosis is an incurable autosomal dominant disorder characterised by skin lesions and the formation of benign tumours in multiple organs, particularly the central nervous system. The disease is caused by mutations in the tumour suppressor genes TSC1 (tuberous sclerosis 1) and TSC2, which function in the PI3K/mTOR signalling cascade and are also mutated in some cancers.

Class 1 PI3K activation recruits Akt to cell membranes via PIP3, where it is phosphorylated by mTORC2 and PDK1. Activated Akt phosphorylates TSC2, which exists in complex with TSC1. In this state, the GTPase-activating protein (GAP) function of TSC2 is inhibited, leading to the accumulation of active, GTP-bound Rheb and mTORC1 activation via downstream mediators such as p70S6K and 4E-BP1. As such, mutations in TSC1/TSC2 lead to hyperactivation of mTORC1.

Everolimus, an mTORC1 inhibitor, is approved for the control of tuberous sclerosis but TSC1/TSC2 mutant cancers are frequently not susceptible to mTORC1 inhibition, suggesting that TSC1/TSC2 mutations might have pro-tumorigenic effects that are mTORC1-independent.

Schematic representation of the role of the TSC complex in mTORC1 signalling.

Key findings

In their preprint, Wei, Leung and their colleagues report a liquid chromatography-mass spectrometry experiment comparing Tsc1-/-mouse embryonic fibroblasts (MEFs) with isogenic Tsc1+/+cells. After stable isotope labelling with amino acids in culture, N-linked glycosylation enrichment allowed interrogation of the cell surface proteome. 20 proteins were identified as upregulated in Tsc1-/-MEFs; reassuringly, these included proteins whose expression is known to be upregulated by mTORC1 activation and multiple proteins involved in cell adhesion, which TSC1 is known to regulate.

After flow cytometry validation, the top two hits were neprilysin (NEP) and aminopeptidase N (APN/CD13). TSC1 knockdown by shRNA or deletion using CRISPR-Cas9 led to increased NEP and APN expression in human cancer cell lines. Moreover, suppression of NEP and APN using siRNA led to large reductions in cell line proliferation. Existing inhibitors of these proteins had mixed results: LBQ657, a NEP inhibitor, and bestatin, an APN inhibitor, had no impact on proliferation but a more specific APN inhibitor, CHR2797, was effective through the induction of apoptosis. Counterintuitively, CHR2797 did not affect mTORC1 substrate phosphorylation and APN expression was not reduced by CHR2797 in TSC1-null cell lines, suggesting that TSC1 loss upregulates APN independently of mTORC1 signalling. Combinations of mTORC1 inhibitors and CHR2797 were synergistic, suggesting a novel therapeutic strategy for TSC1-mutant disease.

Conclusions and Outlook

Since Tsc1-/- MEFs do not have higher expression of APN and APN upregulation is independent of mTORC1, these data suggest that TSC1 mutations have effects that are independent of mTORC1. As such, the current strategy to treat TSC-mutant tumours with mTORC1 inhibitors may be insufficient and these might be susceptible to combination therapies. It’s interesting too that TSC2-deficient tumours can respond to checkpoint inhibition so we will have the opportunity to decipher the best combination of inhibitors of mTORC1, inhibitors of these novel targets and immunotherapies over the coming years.

The preprint suggests new avenues for basic research as many of the 20 proteins found to be upregulated are poorly understood. It also adds to recent progress on the mTORC1-independent roles of the TSC1/TSC2 complex: both TSC1 and TBC1D7, a third subunit of the TSC1/TSC2 complex that binds to and stabilises TSC1 but is not expressed in all tissues, are tumour suppressors in canonical mTORC1 signalling. However, disruption of their interaction seems to be growth promoting in lung cancer cell lines, supportive of the idea that these proteins participate in other pathways independently of TSC2. To date, TSC1 has been shown to participate in TGFbeta-induced Smad2/3 signalling (and consequently affect the Hedgehog pathway) and TBC1D7 operates independently in insulin signalling. As such, the elucidation of the mechanism by which APN and NEP are upregulated in TSC1-/- cells will be noteworthy.

Further reading

Rosset, C., Netto, C.B.O. and Ashton-Prolla, P. TSC1 and TSC2 gene mutations and their implications for treatment in Tuberous Sclerosis Complex: a review. Genetics and Molecular Biology, 2017. 40(1): p. 69–79.

Questions for Authors

  1. The authors performed their discovery experiment in MEFs. How significant might species differences be? Might there be additional targetable surface proteins in TSC1-mutant human cells?
  2. AGRL2 is mentioned as an upregulated gene that has low expression in normal tissues – could this (or other hits) be a target for CAR T cell therapy in TSC1-mutant disease, independently of the success of small molecule inhibitor-based approaches?

Tags: cancer, mass spectrometry, proteomics, tuberous sclerosis

Posted on: 9 August 2018

doi: https://doi.org/10.1242/prelights.4241

Read preprint (No Ratings Yet)

Author's Response

Junnian Wei, Jim Wells and Mike Evans shared

Thanks for taking the time to highlight our work.

Q1. This is an important point.  We chose to study MEFs as complete TSC1 knockout cells already existed, and were previously studied to glean important insights about the biology associated with TSC1/2 loss and mTORC1 signaling. While APN and NEP are both upregulated in human cell lines due to TSC1 loss, it is possible that there are human-specific targets associated with TSC1 that our proteomics screen did not capture. We are planning proteomics experiments in human bladder cancer cell lines isogenic with respect to TSC1 expression to address this question.

Q2. Certainly this is a possibility, assuming potent and selective monoclonal antibodies can be raised to an extracellular epitope.  AGRL2 is also a GPCR, so small molecule agonists or antagonists could also have bioactivity.  We are conducting further studies to understand the role of AGRL2 expression and/or activity to the survival of cells with TSC1 mutations.

Have your say

Your email address will not be published. Required fields are marked *

This site uses Akismet to reduce spam. Learn how your comment data is processed.

Sign up to customise the site to your preferences and to receive alerts

Register here

Also in the cancer biology category:

Spatial transcriptomics elucidates medulla niche supporting germinal center response in myasthenia gravis thymoma

Yoshiaki Yasumizu, Makoto Kinoshita, Martin Jinye Zhang, et al.



Selected by Jessica Chevallier

Invasion of glioma cells through confined space requires membrane tension regulation and mechano-electrical coupling via Plexin-B2

Chrystian Junqueira Alves, Theodore Hannah, Sita Sadia, et al.



Selected by Jade Chan

Modeling transcriptional profiles of gene perturbation with a deep neural network

Wenke Liu, Xuya Wang, D R Mani, et al.



Selected by Raquel Moya

Also in the cell biology category:

Clusters of lineage-specific genes are anchored by ZNF274 in repressive perinucleolar compartments

Martina Begnis, Julien Duc, Sandra Offner, et al.



Selected by Silvia Carvalho

Structural basis of respiratory complexes adaptation to cold temperatures

Young-Cheul Shin, Pedro Latorre-Muro, Amina Djurabekova, et al.



Selected by Pamela Ornelas

RIPK3 coordinates RHIM domain-dependent inflammatory transcription in neurons

Sigal B. Kofman, Lan H. Chu, Joshua M. Ames, et al.



Selected by Zoie Magri

preLists in the cancer biology category:

CSHL 87th Symposium: Stem Cells

Preprints mentioned by speakers at the #CSHLsymp23

 



List by Alex Eve

Journal of Cell Science meeting ‘Imaging Cell Dynamics’

This preList highlights the preprints discussed at the JCS meeting 'Imaging Cell Dynamics'. The meeting was held from 14 - 17 May 2023 in Lisbon, Portugal and was organised by Erika Holzbaur, Jennifer Lippincott-Schwartz, Rob Parton and Michael Way.

 



List by Helen Zenner

CellBio 2022 – An ASCB/EMBO Meeting

This preLists features preprints that were discussed and presented during the CellBio 2022 meeting in Washington, DC in December 2022.

 



List by Nadja Hümpfer et al.

Fibroblasts

The advances in fibroblast biology preList explores the recent discoveries and preprints of the fibroblast world. Get ready to immerse yourself with this list created for fibroblasts aficionados and lovers, and beyond. Here, my goal is to include preprints of fibroblast biology, heterogeneity, fate, extracellular matrix, behavior, topography, single-cell atlases, spatial transcriptomics, and their matrix!

 



List by Osvaldo Contreras

Single Cell Biology 2020

A list of preprints mentioned at the Wellcome Genome Campus Single Cell Biology 2020 meeting.

 



List by Alex Eve

ASCB EMBO Annual Meeting 2019

A collection of preprints presented at the 2019 ASCB EMBO Meeting in Washington, DC (December 7-11)

 



List by Madhuja Samaddar et al.

Lung Disease and Regeneration

This preprint list compiles highlights from the field of lung biology.

 



List by Rob Hynds

Anticancer agents: Discovery and clinical use

Preprints that describe the discovery of anticancer agents and their clinical use. Includes both small molecules and macromolecules like biologics.

 



List by Zhang-He Goh

Biophysical Society Annual Meeting 2019

Few of the preprints that were discussed in the recent BPS annual meeting at Baltimore, USA

 



List by Joseph Jose Thottacherry

Also in the cell biology category:

‘In preprints’ from Development 2022-2023

A list of the preprints featured in Development's 'In preprints' articles between 2022-2023

 



List by Alex Eve, Katherine Brown

preLights peer support – preprints of interest

This is a preprint repository to organise the preprints and preLights covered through the 'preLights peer support' initiative.

 



List by preLights peer support

The Society for Developmental Biology 82nd Annual Meeting

This preList is made up of the preprints discussed during the Society for Developmental Biology 82nd Annual Meeting that took place in Chicago in July 2023.

 



List by Joyce Yu, Katherine Brown

CSHL 87th Symposium: Stem Cells

Preprints mentioned by speakers at the #CSHLsymp23

 



List by Alex Eve

Journal of Cell Science meeting ‘Imaging Cell Dynamics’

This preList highlights the preprints discussed at the JCS meeting 'Imaging Cell Dynamics'. The meeting was held from 14 - 17 May 2023 in Lisbon, Portugal and was organised by Erika Holzbaur, Jennifer Lippincott-Schwartz, Rob Parton and Michael Way.

 



List by Helen Zenner

9th International Symposium on the Biology of Vertebrate Sex Determination

This preList contains preprints discussed during the 9th International Symposium on the Biology of Vertebrate Sex Determination. This conference was held in Kona, Hawaii from April 17th to 21st 2023.

 



List by Martin Estermann

Alumni picks – preLights 5th Birthday

This preList contains preprints that were picked and highlighted by preLights Alumni - an initiative that was set up to mark preLights 5th birthday. More entries will follow throughout February and March 2023.

 



List by Sergio Menchero et al.

CellBio 2022 – An ASCB/EMBO Meeting

This preLists features preprints that were discussed and presented during the CellBio 2022 meeting in Washington, DC in December 2022.

 



List by Nadja Hümpfer et al.

Fibroblasts

The advances in fibroblast biology preList explores the recent discoveries and preprints of the fibroblast world. Get ready to immerse yourself with this list created for fibroblasts aficionados and lovers, and beyond. Here, my goal is to include preprints of fibroblast biology, heterogeneity, fate, extracellular matrix, behavior, topography, single-cell atlases, spatial transcriptomics, and their matrix!

 



List by Osvaldo Contreras

EMBL Synthetic Morphogenesis: From Gene Circuits to Tissue Architecture (2021)

A list of preprints mentioned at the #EESmorphoG virtual meeting in 2021.

 



List by Alex Eve

FENS 2020

A collection of preprints presented during the virtual meeting of the Federation of European Neuroscience Societies (FENS) in 2020

 



List by Ana Dorrego-Rivas

Planar Cell Polarity – PCP

This preList contains preprints about the latest findings on Planar Cell Polarity (PCP) in various model organisms at the molecular, cellular and tissue levels.

 



List by Ana Dorrego-Rivas

BioMalPar XVI: Biology and Pathology of the Malaria Parasite

[under construction] Preprints presented at the (fully virtual) EMBL BioMalPar XVI, 17-18 May 2020 #emblmalaria

 



List by Dey Lab, Samantha Seah

1

Cell Polarity

Recent research from the field of cell polarity is summarized in this list of preprints. It comprises of studies focusing on various forms of cell polarity ranging from epithelial polarity, planar cell polarity to front-to-rear polarity.

 



List by Yamini Ravichandran

TAGC 2020

Preprints recently presented at the virtual Allied Genetics Conference, April 22-26, 2020. #TAGC20

 



List by Maiko Kitaoka et al.

3D Gastruloids

A curated list of preprints related to Gastruloids (in vitro models of early development obtained by 3D aggregation of embryonic cells). Updated until July 2021.

 



List by Paul Gerald L. Sanchez and Stefano Vianello

ECFG15 – Fungal biology

Preprints presented at 15th European Conference on Fungal Genetics 17-20 February 2020 Rome

 



List by Hiral Shah

ASCB EMBO Annual Meeting 2019

A collection of preprints presented at the 2019 ASCB EMBO Meeting in Washington, DC (December 7-11)

 



List by Madhuja Samaddar et al.

EMBL Seeing is Believing – Imaging the Molecular Processes of Life

Preprints discussed at the 2019 edition of Seeing is Believing, at EMBL Heidelberg from the 9th-12th October 2019

 



List by Dey Lab

Autophagy

Preprints on autophagy and lysosomal degradation and its role in neurodegeneration and disease. Includes molecular mechanisms, upstream signalling and regulation as well as studies on pharmaceutical interventions to upregulate the process.

 



List by Sandra Malmgren Hill

Lung Disease and Regeneration

This preprint list compiles highlights from the field of lung biology.

 



List by Rob Hynds

Cellular metabolism

A curated list of preprints related to cellular metabolism at Biorxiv by Pablo Ranea Robles from the Prelights community. Special interest on lipid metabolism, peroxisomes and mitochondria.

 



List by Pablo Ranea Robles

BSCB/BSDB Annual Meeting 2019

Preprints presented at the BSCB/BSDB Annual Meeting 2019

 



List by Dey Lab

MitoList

This list of preprints is focused on work expanding our knowledge on mitochondria in any organism, tissue or cell type, from the normal biology to the pathology.

 



List by Sandra Franco Iborra

ASCB/EMBO Annual Meeting 2018

This list relates to preprints that were discussed at the recent ASCB conference.

 



List by Dey Lab, Amanda Haage
Close