p25/Cdk5-mediated retinoblastoma phosphorylation is an early event in neuronal cell death.
In large models of neuronal cell death, there is a tight correlation between Cdk5 deregulation and cell-cycle dysfunction. However, pathways that link Cdk5 to the cell cycle during neuronal death are still unclear. We have investigated the molecular events that precede p25/Cdk5-triggered neuronal death using a neuronal cell line that allows inducible p25 expression. In this system, no sign of apoptosis was seen before 24 hours of p25 induction. Thus, at that time, cell-cycle-regulatory proteins were analysed by immunoblotting and some of them showed a significant deregulation. Interestingly, after time-course experiments, the earliest feature correlated with p25 expression was the phosphorylation of the retinoblastoma protein (Rb). Indeed, this phosphorylation was observed 6 hours after p25 induction and was abolished in the presence of a Cdk5 inhibitor, roscovitine, which does not inhibit the usual Rb cyclin-D kinases Cdk4 and Cdk6. Furthermore, analyses of levels and subcellular localization of Cdk-related cyclins did not reveal any change following Cdk5 activation, arguing for a direct effect of Cdk5 activity on Rb protein. This latter result was clearly demonstrated by in vitro kinase assays showing that the p25-Cdk5 complex in our cell system phosphorylates Rb directly without the need for any intermediary kinase activity. Hence, Rb might be an appropriate candidate that connects Cdk5 to cell-cycle deregulation during neuronal cell death.
Hamdane
Malika
M
INSERM U422, Institut de Médecine Prédictive et Recherche Thérapeutique, Université de Lille 2, Place de Verdun, 59045 Lille Cedex, France.
Bretteville
Alexis
A
Sambo
Anne-Véronique
AV
Schindowski
Katharina
K
Bégard
Séverine
S
Delacourte
André
A
Bertrand
Philippe
P
Buée
Luc
L
D017209:Apoptosis; D015153:Blotting, Western; D002453:Cell Cycle; D018797:Cell Cycle Proteins / Q000378:metabolism; D016923:Cell Death; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D051360:Cyclin-Dependent Kinase 5; D018844:Cyclin-Dependent Kinases / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D050684:E2F Transcription Factors; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D020932:Nerve Growth Factor / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D009474:Neurons / Q000378:metabolism / Q000473:pathology*; D010766:Phosphorylation; D011687:Purines / Q000494:pharmacology; D016160:Retinoblastoma Protein / Q000378:metabolism*; D000077546:Roscovitine; D013997:Time Factors; D014157:Transcription Factors / Q000378:metabolism
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D009419:Nerve Tissue Proteins; D011687:Purines; D016160:Retinoblastoma Protein; D014157:Transcription Factors; C089495:neuronal Cdk5 activator (p25-p35); D000077546:Roscovitine; D020932:Nerve Growth Factor; D051360:Cyclin-Dependent Kinase 5; C495905:CDK5 protein, human; D018844:Cyclin-Dependent Kinases