Divergent modulation of neuronal differentiation by caspase-2 and -9.
Human Ntera2/cl.D1 (NT2) cells treated with retinoic acid (RA) differentiate towards a well characterized neuronal phenotype sharing many features with human fetal neurons. In view of the emerging role of caspases in murine stem cell/neural precursor differentiation, caspases activity was evaluated during RA differentiation. Caspase-2, -3 and -9 activity was transiently and selectively increased in differentiating and non-apoptotic NT2-cells. SiRNA-mediated selective silencing of either caspase-2 (si-Casp2) or -9 (si-Casp9) was implemented in order to dissect the role of distinct caspases. The RA-induced expression of neuronal markers, i.e. neural cell adhesion molecule (NCAM), microtubule associated protein-2 (MAP2) and tyrosine hydroxylase (TH) mRNAs and proteins, was decreased in si-Casp9, but markedly increased in si-Casp2 cells. During RA-induced NT2 differentiation, the class III histone deacetylase Sirt1, a putative caspase substrate implicated in the regulation of the proneural bHLH MASH1 gene expression, was cleaved to a ∼100 kDa fragment. Sirt1 cleavage was markedly reduced in si-Casp9 cells, even though caspase-3 was normally activated, but was not affected (still cleaved) in si-Casp2 cells, despite a marked reduction of caspase-3 activity. The expression of MASH1 mRNA was higher and occurred earlier in si-Casp2 cells, while was reduced at early time points during differentiation in si-Casp9 cells. Thus, caspase-2 and -9 may perform opposite functions during RA-induced NT2 neuronal differentiation. While caspase-9 activation is relevant for proper neuronal differentiation, likely through the fine tuning of Sirt1 function, caspase-2 activation appears to hinder the RA-induced neuronal differentiation of NT2 cells.
Department of Neuroscience, University of Rome Tor Vergata-Medical School, Rome, Italy. pistritto@med.uniroma2.it
Giuseppa
G
Pistritto
Veruska
V
Papaleo
Pilar
P
Sanchez
Claudia
C
Ceci
Maria Luisa
ML
Barbaccia
D017209:Apoptosis / Q000502:physiology; D053143:Caspase 2 / Q000378:metabolism*; D053148:Caspase 3 / Q000502:physiology; D053453:Caspase 9 / Q000378:metabolism*; D002454:Cell Differentiation / Q000502:physiology*; D045744:Cell Line, Tumor; D006801:Humans; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D019006:Neural Cell Adhesion Molecules / Q000378:metabolism; D009474:Neurons / Q000502:physiology*; D034741:RNA, Small Interfering; D014162:Transfection; D014446:Tyrosine 3-Monooxygenase / Q000378:metabolism
C489788:MAP2 protein, human; D008869:Microtubule-Associated Proteins; D019006:Neural Cell Adhesion Molecules; D034741:RNA, Small Interfering; D014446:Tyrosine 3-Monooxygenase; D053143:Caspase 2; D053148:Caspase 3; D053453:Caspase 9