Silencing of the microtubule-associated proteins doublecortin-like and doublecortin-like kinase-long induces apoptosis in neuroblastoma cells.
Doublecortin-like kinase-long (DCLK-long) and doublecortin-like (DCL) are two splice variants of DCLK gene. DCL and DCLK-long are microtubule-associated proteins with specific expression in proliferative neural progenitor cells. We have tested the hypothesis that knockdown of DCL/DCLK-long by RNA interference technology will induce cell death in neuroblastoma (NB) cells. First, we analyzed the expression of DCL and DCLK-long in several human neuroblastic tumors, other tumors, and normal tissues, revealing high expression of both DCL and DCLK-long in NB and glioma. Secondly, gene expression profiling revealed numerous differentially expressed genes indicating apoptosis induction after DCL/DCLK-long knockdown in NB cells. Finally, apoptosis was confirmed by time-lapse imaging of phosphatidylserine translocation, caspase-3 activation, live/dead double staining assays, and fluorescence-activated cell sorting. Together, our results suggest that silencing DCL/DCLK-long induces apoptosis in NB cells.
Leiden/Amsterdam Center for Drug Research, Leiden, The Netherlands.
Carla S
CS
Verissimo
Jan J
JJ
Molenaar
John
J
Meerman
Jordi Carreras
JC
Puigvert
Fieke
F
Lamers
Jan
J
Koster
Erik H J
EH
Danen
Bob
B
van de Water
Rogier
R
Versteeg
Carlos P
CP
Fitzsimons
Erno
E
Vreugdenhil
D000818:Animals; D017209:Apoptosis / Q000235:genetics*; D015153:Blotting, Western; D053148:Caspase 3 / Q000235:genetics / Q000378:metabolism; D002452:Cell Count; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D000089742:Doublecortin-Like Kinases; D005434:Flow Cytometry; D020869:Gene Expression Profiling; D006801:Humans; D007091:Image Processing, Computer-Assisted; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D009474:Neurons / Q000378:metabolism*; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000235:genetics* / Q000378:metabolism; D034622:RNA Interference; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013234:Stem Cells / Q000166:cytology / Q000378:metabolism
D047908:Intracellular Signaling Peptides and Proteins; D020033:Protein Isoforms; D012333:RNA, Messenger; D034741:RNA, Small Interfering; C117347:DCLK1 protein, human; D000089742:Doublecortin-Like Kinases; D017346:Protein Serine-Threonine Kinases; D053148:Caspase 3