A GSK-3/b-CATENIN/ARHGAP axis regulates glioblastoma invasion

Anke Bruning-Richardson, Michel Mittelbronn, Matthew Humphries, Filomena Esteves, Marjorie Boissinot, Daniel Tams, Alastair Droop, Julia V. Cockle, Sophie Taylor, Ruth Morton, Azzam Ismail, Sean Lawler, Georgia Mavria, Susan C Short

Research output: Contribution to journalMeeting Abstract

Abstract

INTRODUCTION
Glycogen synthase kinase-3 (GSK-3) has been implicated as a potential target in glioblastoma migration and invasion. Here we investigated the effects of GSK-3 inhibition on downstream effector genes and discovered a novel GSK-3 /b-catenin/ARHGAP migration axis that drives glioblastoma invasion.

METHODS
Expression arrays of glioblastoma cells treated with the GSK-3 inhibitor 6-bromoindirubin-3’-oxime (BIO) were performed. Functional validation was carried out using 2D and 3D migration assays, siRNA knockdown and orthotopic xenograft models. Protein expression was investigated in patient glioblastoma samples.

RESULTS
Microarray data analysis identified deregulation of expression of members of the ARHGAP gene family of GTPase regulators after BIO treatment of U251 cells. This was confirmed at the protein level by Western blotting and immunostaining. siRNA treatment indicated a specific promigratory role for ARHGAP29. U251 cells exposed to BIO displayed a decrease in migration as evidenced by loss of polarity and reduction in cell velocity. This was associated with a rearrangement of actin filaments and b-catenin translocation from the cell surface to the nucleus, where it is known to act as a transcription factor downstream of GSK-3 inhibition. We then found that ICG001, an inhibitor of b-catenin-induced transcription, essentially mimicking GSK-3 activation, induced transcription of ARHGAP29 and led to a partial increase of ARHGAP29 protein expression, reduction of b-catenin target gene expression and the ability of cells to migrate. In vivo studies, as well as data from glioblastoma patients, confirmed a pro-migratory role for ARHGAP29 as evidenced by elevated protein levels in migratory cells at the invasive front in orthotopic xenograft models and its association with disease recurrence.

DISCUSSION
We demonstrate for the first time signalling events downstream of GSK-3 activation and b-catenin nuclear translocation which reveal a new pathway regulating glioblastoma migration through the transcription of the GTPase regulator ARHGAP29.
Original languageEnglish
Pages (from-to)24
Number of pages1
JournalNeuro-Oncology
Volume19
Issue numberS6
DOIs
Publication statusPublished - Nov 2017
Externally publishedYes
Event22nd Annual Scientific Meeting and Education Day of the Society for Neuro-Oncology - San Francisco, United States
Duration: 16 Nov 201719 Nov 2017
Conference number: 22

Fingerprint

Glycogen Synthase Kinase 3
Glioblastoma
Catenins
Oximes
GTP Phosphohydrolases
Heterografts
Small Interfering RNA
Proteins
Actin Cytoskeleton
Transcriptional Activation
Genes
Transcription Factors
Phosphotransferases
Western Blotting
Gene Expression
Recurrence
Therapeutics

Cite this

Bruning-Richardson, A., Mittelbronn, M., Humphries, M., Esteves, F., Boissinot, M., Tams, D., ... Short, S. C. (2017). A GSK-3/b-CATENIN/ARHGAP axis regulates glioblastoma invasion. Neuro-Oncology, 19(S6), 24. https://doi.org/10.1093/neuonc/nox168.092
Bruning-Richardson, Anke ; Mittelbronn, Michel ; Humphries, Matthew ; Esteves, Filomena ; Boissinot, Marjorie ; Tams, Daniel ; Droop, Alastair ; Cockle, Julia V. ; Taylor, Sophie ; Morton, Ruth ; Ismail, Azzam ; Lawler, Sean ; Mavria, Georgia ; Short, Susan C. / A GSK-3/b-CATENIN/ARHGAP axis regulates glioblastoma invasion. In: Neuro-Oncology. 2017 ; Vol. 19, No. S6. pp. 24.
@article{5cd3dcc065a9495c966ea256334529bd,
title = "A GSK-3/b-CATENIN/ARHGAP axis regulates glioblastoma invasion",
abstract = "INTRODUCTIONGlycogen synthase kinase-3 (GSK-3) has been implicated as a potential target in glioblastoma migration and invasion. Here we investigated the effects of GSK-3 inhibition on downstream effector genes and discovered a novel GSK-3 /b-catenin/ARHGAP migration axis that drives glioblastoma invasion.METHODSExpression arrays of glioblastoma cells treated with the GSK-3 inhibitor 6-bromoindirubin-3’-oxime (BIO) were performed. Functional validation was carried out using 2D and 3D migration assays, siRNA knockdown and orthotopic xenograft models. Protein expression was investigated in patient glioblastoma samples.RESULTSMicroarray data analysis identified deregulation of expression of members of the ARHGAP gene family of GTPase regulators after BIO treatment of U251 cells. This was confirmed at the protein level by Western blotting and immunostaining. siRNA treatment indicated a specific promigratory role for ARHGAP29. U251 cells exposed to BIO displayed a decrease in migration as evidenced by loss of polarity and reduction in cell velocity. This was associated with a rearrangement of actin filaments and b-catenin translocation from the cell surface to the nucleus, where it is known to act as a transcription factor downstream of GSK-3 inhibition. We then found that ICG001, an inhibitor of b-catenin-induced transcription, essentially mimicking GSK-3 activation, induced transcription of ARHGAP29 and led to a partial increase of ARHGAP29 protein expression, reduction of b-catenin target gene expression and the ability of cells to migrate. In vivo studies, as well as data from glioblastoma patients, confirmed a pro-migratory role for ARHGAP29 as evidenced by elevated protein levels in migratory cells at the invasive front in orthotopic xenograft models and its association with disease recurrence.DISCUSSIONWe demonstrate for the first time signalling events downstream of GSK-3 activation and b-catenin nuclear translocation which reveal a new pathway regulating glioblastoma migration through the transcription of the GTPase regulator ARHGAP29.",
author = "Anke Bruning-Richardson and Michel Mittelbronn and Matthew Humphries and Filomena Esteves and Marjorie Boissinot and Daniel Tams and Alastair Droop and Cockle, {Julia V.} and Sophie Taylor and Ruth Morton and Azzam Ismail and Sean Lawler and Georgia Mavria and Short, {Susan C}",
year = "2017",
month = "11",
doi = "10.1093/neuonc/nox168.092",
language = "English",
volume = "19",
pages = "24",
journal = "Neuro-Oncology",
issn = "1522-8517",
publisher = "Oxford University Press",
number = "S6",

}

Bruning-Richardson, A, Mittelbronn, M, Humphries, M, Esteves, F, Boissinot, M, Tams, D, Droop, A, Cockle, JV, Taylor, S, Morton, R, Ismail, A, Lawler, S, Mavria, G & Short, SC 2017, 'A GSK-3/b-CATENIN/ARHGAP axis regulates glioblastoma invasion', Neuro-Oncology, vol. 19, no. S6, pp. 24. https://doi.org/10.1093/neuonc/nox168.092

A GSK-3/b-CATENIN/ARHGAP axis regulates glioblastoma invasion. / Bruning-Richardson, Anke; Mittelbronn, Michel; Humphries, Matthew; Esteves, Filomena; Boissinot, Marjorie; Tams, Daniel; Droop, Alastair; Cockle, Julia V.; Taylor, Sophie; Morton, Ruth; Ismail, Azzam; Lawler, Sean; Mavria, Georgia; Short, Susan C.

In: Neuro-Oncology, Vol. 19, No. S6, 11.2017, p. 24.

Research output: Contribution to journalMeeting Abstract

TY - JOUR

T1 - A GSK-3/b-CATENIN/ARHGAP axis regulates glioblastoma invasion

AU - Bruning-Richardson, Anke

AU - Mittelbronn, Michel

AU - Humphries, Matthew

AU - Esteves, Filomena

AU - Boissinot, Marjorie

AU - Tams, Daniel

AU - Droop, Alastair

AU - Cockle, Julia V.

AU - Taylor, Sophie

AU - Morton, Ruth

AU - Ismail, Azzam

AU - Lawler, Sean

AU - Mavria, Georgia

AU - Short, Susan C

PY - 2017/11

Y1 - 2017/11

N2 - INTRODUCTIONGlycogen synthase kinase-3 (GSK-3) has been implicated as a potential target in glioblastoma migration and invasion. Here we investigated the effects of GSK-3 inhibition on downstream effector genes and discovered a novel GSK-3 /b-catenin/ARHGAP migration axis that drives glioblastoma invasion.METHODSExpression arrays of glioblastoma cells treated with the GSK-3 inhibitor 6-bromoindirubin-3’-oxime (BIO) were performed. Functional validation was carried out using 2D and 3D migration assays, siRNA knockdown and orthotopic xenograft models. Protein expression was investigated in patient glioblastoma samples.RESULTSMicroarray data analysis identified deregulation of expression of members of the ARHGAP gene family of GTPase regulators after BIO treatment of U251 cells. This was confirmed at the protein level by Western blotting and immunostaining. siRNA treatment indicated a specific promigratory role for ARHGAP29. U251 cells exposed to BIO displayed a decrease in migration as evidenced by loss of polarity and reduction in cell velocity. This was associated with a rearrangement of actin filaments and b-catenin translocation from the cell surface to the nucleus, where it is known to act as a transcription factor downstream of GSK-3 inhibition. We then found that ICG001, an inhibitor of b-catenin-induced transcription, essentially mimicking GSK-3 activation, induced transcription of ARHGAP29 and led to a partial increase of ARHGAP29 protein expression, reduction of b-catenin target gene expression and the ability of cells to migrate. In vivo studies, as well as data from glioblastoma patients, confirmed a pro-migratory role for ARHGAP29 as evidenced by elevated protein levels in migratory cells at the invasive front in orthotopic xenograft models and its association with disease recurrence.DISCUSSIONWe demonstrate for the first time signalling events downstream of GSK-3 activation and b-catenin nuclear translocation which reveal a new pathway regulating glioblastoma migration through the transcription of the GTPase regulator ARHGAP29.

AB - INTRODUCTIONGlycogen synthase kinase-3 (GSK-3) has been implicated as a potential target in glioblastoma migration and invasion. Here we investigated the effects of GSK-3 inhibition on downstream effector genes and discovered a novel GSK-3 /b-catenin/ARHGAP migration axis that drives glioblastoma invasion.METHODSExpression arrays of glioblastoma cells treated with the GSK-3 inhibitor 6-bromoindirubin-3’-oxime (BIO) were performed. Functional validation was carried out using 2D and 3D migration assays, siRNA knockdown and orthotopic xenograft models. Protein expression was investigated in patient glioblastoma samples.RESULTSMicroarray data analysis identified deregulation of expression of members of the ARHGAP gene family of GTPase regulators after BIO treatment of U251 cells. This was confirmed at the protein level by Western blotting and immunostaining. siRNA treatment indicated a specific promigratory role for ARHGAP29. U251 cells exposed to BIO displayed a decrease in migration as evidenced by loss of polarity and reduction in cell velocity. This was associated with a rearrangement of actin filaments and b-catenin translocation from the cell surface to the nucleus, where it is known to act as a transcription factor downstream of GSK-3 inhibition. We then found that ICG001, an inhibitor of b-catenin-induced transcription, essentially mimicking GSK-3 activation, induced transcription of ARHGAP29 and led to a partial increase of ARHGAP29 protein expression, reduction of b-catenin target gene expression and the ability of cells to migrate. In vivo studies, as well as data from glioblastoma patients, confirmed a pro-migratory role for ARHGAP29 as evidenced by elevated protein levels in migratory cells at the invasive front in orthotopic xenograft models and its association with disease recurrence.DISCUSSIONWe demonstrate for the first time signalling events downstream of GSK-3 activation and b-catenin nuclear translocation which reveal a new pathway regulating glioblastoma migration through the transcription of the GTPase regulator ARHGAP29.

U2 - 10.1093/neuonc/nox168.092

DO - 10.1093/neuonc/nox168.092

M3 - Meeting Abstract

VL - 19

SP - 24

JO - Neuro-Oncology

JF - Neuro-Oncology

SN - 1522-8517

IS - S6

ER -

Bruning-Richardson A, Mittelbronn M, Humphries M, Esteves F, Boissinot M, Tams D et al. A GSK-3/b-CATENIN/ARHGAP axis regulates glioblastoma invasion. Neuro-Oncology. 2017 Nov;19(S6):24. https://doi.org/10.1093/neuonc/nox168.092