Stable interference of EWS-FLI1 in an Ewing sarcoma cell line impairs IGF-1/IGF-1R signalling and reveals TOPK as a new target.
BACKGROUND
Ewing sarcoma is a paradigm of solid tumour -bearing chromosomal translocations resulting in fusion proteins that act as deregulated transcription factors. Ewing sarcoma translocations fuse the EWS gene with an ETS transcription factor, mainly FLI1. Most of the EWS-FLI1 target genes still remain unknown and many have been identified in heterologous model systems.


METHODS
We have developed a stable RNA interference model knocking down EWS-FLI1 in the Ewing sarcoma cell line TC71. Gene expression analyses were performed to study the effect of RNA interference on the genetic signature of EWS-FLI1 and to identify genes that could contribute to tumourigenesis.


RESULTS
EWS-FLI1 inhibition induced apoptosis, reduced cell migratory and tumourigenic capacities, and caused reduction in tumour growth. IGF-1 was downregulated and the IGF-1/IGF-1R signalling pathway was impaired. PBK/TOPK (T-LAK cell-originated protein kinase) expression was decreased because of EWS-FLI1 inhibition. We showed that TOPK is a new target gene of EWS-FLI1. TOPK inhibition prompted a decrease in the proliferation rate and a dramatic change in the cell's ability to grow in coalescence.


CONCLUSIONS
This is the first report of TOPK activity in Ewing sarcoma and suggests a significant role of this MAPKK-like protein kinase in the Ewing sarcoma biology.
Herrero-Martín
D
D
Molecular Pathology Program, Centro de Investigación del Cáncer-IBMCC (USAL-CSIC), Campus Unamuno s/n, Salamanca, Spain. david.herrero@kispi.uzh.ch
Osuna
D
D
Ordóñez
J L
JL
Sevillano
V
V
Martins
A S
AS
Mackintosh
C
C
Campos
M
M
Madoz-Gúrpide
J
J
Otero-Motta
A P
AP
Caballero
G
G
Amaral
A T
AT
Wai
D H
DH
Braun
Y
Y
Eisenacher
M
M
Schaefer
K-L
KL
Poremba
C
C
de Alava
E
E
D000818:Animals; D017209:Apoptosis / Q000502:physiology; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000502:physiology; D015536:Down-Regulation; D005260:Female; D006801:Humans; D007334:Insulin-Like Growth Factor I / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D020929:Mitogen-Activated Protein Kinase Kinases; D015514:Oncogene Proteins, Fusion / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D051789:Proto-Oncogene Protein c-fli-1; D034622:RNA Interference; D034802:RNA-Binding Protein EWS; D017526:Receptor, IGF Type 1 / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D012512:Sarcoma, Ewing / Q000201:enzymology / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D015398:Signal Transduction; D014157:Transcription Factors / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism
C106031:EWS-FLI fusion protein; D015514:Oncogene Proteins, Fusion; D051789:Proto-Oncogene Protein c-fli-1; D034802:RNA-Binding Protein EWS; D014157:Transcription Factors; D007334:Insulin-Like Growth Factor I; D017526:Receptor, IGF Type 1; D017346:Protein Serine-Threonine Kinases; D020929:Mitogen-Activated Protein Kinase Kinases; C409022:PDZ-binding kinase