RESUMO
Interferons exert their biological function mainly through the activation of interferon-stimulated genes (ISGs). ISG12 (originally designated p27) belongs to a family of small, interferon alpha inducible genes of unknown function. We have determined the 5' end sequence of ISG12 cDNA from the human cell lines HeLa and AMA by RACE. Comparing this sequence to ISG12 sequences in the expressed sequence tag (EST) database revealed the presence of two alternative splice variants of ISG12 in human cells exhibiting the same open reading frame. We have sequenced the promoter region of the ISG12 gene and found ISRE, IRF1/IRF2, and STAT elements correlating to the interferon alpha inducibility of the gene. Subsequently, we have expressed human ISG12, a 12-kDa hydrophobic protein in the baculovirus expression system and with a C-terminal FLAG-tag in the human cell line 293. Recombinant ISG12 sediments in the nuclear envelope in both cell types. Finally, we have been able to demonstrate the prevalence of the ISG12 gene product in the nuclear envelope of HeLa cells treated with interferon alpha by immunocytochemical analyses. ISG12 is the first interferon induced protein found localizing to the nuclear envelope.
Assuntos
Núcleo Celular/metabolismo , Interferons/metabolismo , Proteínas/metabolismo , Sequência de Aminoácidos , Baculoviridae/genética , Sequência de Bases , Fracionamento Celular , Primers do DNA , DNA Complementar , Imunofluorescência , Células HeLa , Humanos , Membranas Intracelulares/metabolismo , Proteínas de Membrana , Regiões Promotoras Genéticas , Proteínas/genética , RNA Mensageiro/química , RNA Mensageiro/genética , RNA Mensageiro/metabolismoRESUMO
The gene for ISG12 (originally designated p27) was isolated as an oestrogen-induced gene. The authors undertook a comprehensive study using quantitative RT-PCR, in which we delineate the regulation of ISG12 by seven different cytokines including interferons and poly(I). poly(C) in seven human cell lines of different origin. In all cell lines ISG12 is strongly induced by IFN-alpha and only slightly by IFNgamma. Poly(I).poly(C) induces ISG12 in a cell line-dependent manner, whereas none of the other cytokines tested elicited a response. Comparing the induction pattern of ISG12 to that of 6-16 a high degree of similarity was found. The induction levels varied, however, between cell lines.