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Induction of 2,5 oas gene expression and activity is not sufficient for IFN‐γ‐induced neuroblastoma cell differentiation
Author(s) -
Corrias Maria Valeria,
Gribaudo Giorgio,
Guarnaccia Francesco,
Ponzoni Mirco
Publication year - 1995
Publication title -
international journal of cancer
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.475
H-Index - 234
eISSN - 1097-0215
pISSN - 0020-7136
DOI - 10.1002/ijc.2910620219
Subject(s) - gene isoform , messenger rna , microbiology and biotechnology , biology , neuroblastoma , cell culture , gene expression , interferon , gene , cellular differentiation , adenylate kinase , enzyme , biochemistry , immunology , genetics
We showed earlier that interferon‐γ is a powerful inducer of differentiation of human neuroblastoma (NB) cells. Although 2′,5′oligo‐adenylate synthetase (2,5 OAS) may play a role in mediating the anti‐proliferative and/or differentiative effects of interferons (IFNs), direct evidence is lacking. We have investigated gene and protein expression of the 4 different 2,5 OAS isoforms and their cumulative enzymatic activity in a previously characterized IFN‐γ‐sensitive human NB cell line, LAN‐5. Analysis of total and poly(A) + RNA by Northern Mot and RT‐PCR indicated that expression of the mRNA coding for the 40‐, 46‐and 69‐kDa Isoforms was induced in a time‐ and dose‐dependent manner, reaching a maximum after a 36‐hr treatment with 1000 IU/ml of IFN‐γ. In the absence of treatment, only the mRNA for the 69‐kDa isoform was detectable by RT‐PCR. Inhibition of transcription with actinomycin D showed that 2,5 OAS mRNA was quite stable, with a half‐life of about 4 hr. With respect to the protein content, no 2,5 OAS isoform was present in proliferating LAN‐5 cells; following IFN‐γ treatment, the 100‐, 69‐ and 46‐kDa isoforms became detectable. Accordingly, 2,5 OAS enzymatic activity, virtually undetectable in untreated LAN‐5 cells, increased up to 132 pmol oligoadenylate/μg protein/hr after 48 hr of treatment, then slowly decreased, remaining detectable up to 96 hr. However, the 2,5 OAS proteins required an exogenous activation by synthetic dsRNA to exert enzymatic activity. It is therefore conceivable that they do not play a biological role in NB cell functions. Moreover, an increase in 2,5 OAS enzymatic activity was also observed in NB cells resistant to the differentiation‐promoting activity of IFN‐γ, further suggesting that 2,5 OAS induction was not sufficient to trigger IFN‐γ‐dependent neuronal maturation. Furthermore, other differentiation‐inducing agents, such as retinoic acid and cytosine arabinoside, or complete proliferative arrest produced by serum deprivation, failed to enhance 2,5 OAS activity, thus indicating that the 2,5 OAS system is not directly involved in mediating other differentiative pathways of NB cells. © 1995 Wiley‐Liss, Inc.