TISSUE TRANSGLUTAMINASE AND APOPTOSIS - SENSE AND ANTISENSE TRANSFECTION STUDIES WITH HUMAN NEUROBLASTOMA-CELLS

In this report, we show that the overexpression of tissue transglutami nase (tTG) in the human neuroblastoma cell line SK-N-BE(2) renders the se neural crest-derived cells highly susceptible to death by apoptosis . Cells transfected with a full-length tTG cDNA, under the control of a constitutive promoter, show a drastic reduction in proliferative cap acity paralleled by a large increase in cell death rate. The dying tTG -transfected cells exhibit both cytoplasmic and nuclear changes charac teristic of cells undergoing apoptosis. The tTG-transfected cells expr ess high Bcl-2 protein levels as well as phenotypic neural cell adhesi on molecule markers (NCAM and neurofilaments) of cells differentiating along the neuronal pathway. In keeping with these findings, transfect ion of neuroblastoma cells with an expression vector containing segmen ts of the human tTG cDNA in antisense orientation resulted in a pronou nced decrease of both spontaneous and retinoic acid (RA)-induced apopt osis. We also present evidence that (i) the apoptotic program of these neuroectodermal cells is strictly regulated by RA and (ii) cell death by apoptosis in the human neuroblastoma SK-N-BE(2) cells preferential ly occurs in the substrate-adherent phenotype. For the first time, we report here a direct effect of tTG in the phenotypic maturation toward apoptosis. These results indicate that the tTG-dependent irreversible cross-linking of intracellular protein represents an important bioche mical event in the induction of the structural changes featuring cells dying by apoptosis.