Methylation-Dependent Binding of the Epstein-Barr Virus BZLF1 Protein to Viral Promoters

The switch between latent and lytic Epstein-Barr virus (EBV) infection is mediated by the viral immediate-early (IE) protein, BZLF1 (Z). Z, a homologue of c-jun that binds to AP1-like motifs (ZREs), induces expression of the BRLF1 (R) and BRRF1 (Na) viral proteins, which cooperatively activate transcription of the Z promoter and thereby establish a positive autoregulatory loop. A unique feature of Z is its ability to preferentially bind to, and activate, the methylated form of the BRLF1 promoter (Rp). To date, however, Rp is the only EBV promoter known to be regulated in this unusual manner. We now demonstrate that the promoter driving transcription of the early BRRF1 gene (Nap) has two CpG-containing ZREs (ACGCTCA and TCGCCCG) that are only bound by Z in the methylated state. Both Nap ZREs are highly methylated in cells with latent EBV infection. Z efficiently activates the methylated, but not unmethylated, form of Nap in reporter gene assays, and both ZREs are required. Z serine residue 186, which was previously shown to be required for Z binding to methylated ZREs in Rp, but not for Z binding to the AP1 site, is required for Z binding to methylated Nap ZREs. The Z(S186A) mutant cannot activate methylated Nap in reporter gene assays and does not induce Na expression in cells with latent EBV infection. Molecular modeling studies of Z bound to the methylated Nap ZREs help to explain why methylation is required for Z binding, and the role of the Z Ser186 residue. Methylation-dependent Z binding to critical viral promoters may enhance lytic reactivation in latently infected cells, where the viral genome is heavily methylated. Conversely, since the incoming viral genome is initially unmethylated, methylation-dependent Z activation may also help the virus to establish latency following infection

Efeitos de diferentes frações inspiradas de oxigênio no índice biespectral em cães submetidos à infusão contínua de propofol Effects of several inspired oxygen fractions on the bispectral index in dogs submitted to continuous infusion of propofol

Avaliaram-se os efeitos do fornecimento de diferentes frações inspiradas de oxigênio (FiO2) sobre o índice biespectral (BIS) em cães submetidos a infusão contínua de propofol e mantidos em ventilação espontânea. Oito cães foram submetidos a cinco anestesias, diferenciando-se uma da outra pela FiO2 fornecida. Formaram-se cinco grupos denominados G100 (FiO2 = 1); G80 (FiO2 = 0,8); G60 (FiO2 = 0,6); G40 (FiO2 = 0,4) e G20 (FiO2 = 0,21). Os animais foram induzidos à anestesia com propofol na dose necessária para intubação, e, ato contínuo, iniciaram-se a infusão do fármaco e o fornecimento de oxigênio, conforme a FiO2 determinada para cada grupo. As primeiras mensurações (M0) foram efetuadas 30 minutos após o início da infusão do anestésico e, depois, em intervalos de 15 minutos (M15, M30, M45 e M60). A pressão parcial de oxigênio no sangue arterial (PaO2) variou conforme a FiO2, ou seja, quanto maior a FiO2, maior foi a PaO2. Para a pressão parcial de dióxido de carbono no sangue arterial (PaCO2), foram registradas diferenças em M30, no qual G100 foi maior que G20. Não foram observadas diferenças significativas nas variáveis estudadas do BIS. Os intervalos de médias registrados para o BIS foram, para G100, de 68 a 62; G80, de 71 a 58; G60, de 72 a 62; G40, de 76 a 68; e G20, de 77 a 68. Conclui-se que as variáveis relacionadas ao BIS não são afetadas pelo emprego de diferentes FiO2, e sugere-se que o monitoramento pelo BIS foi capaz de detectar alterações no equilíbrio do fluxo sangüíneo cerebral, oriundas das alterações ocasionadas na dinâmica respiratória pelo emprego de diferentes frações inspiradas de oxigênio.<br>The effects of several inspired oxygen fractions (FiO2) on the bispectral index in spontaneously breathing dogs submitted to continuous infusion of propofol were evaluated. Eight adult mongrel dogs were used. Each animal was submitted to five anesthesias. In each procedure, the patient was allowed to breathe a different FiO2, thereby resulting in five groups, namely: G100 (FiO2 = 1), G80 (FiO2 = 0.8), G60 (FiO2 =0.6), G40 (FiO2 = 0.4), and G20 (FiO2 = 0.21). To induce anesthesia, propofol was given until the animals allowed orotracheal intubation, followed by immediate continuous infusion of drug. The initial measurement (M0) was recorded 30 minutes after the infusion of propofol onset. Additional recordings were performed at 15-minute intervals during 60 minutes (M15, M30, M45, and M60). No significant differences on BIS parameters were recorded. Regarding arterial partial pressure of carbon dioxide (PaCO2), the mean of G100 was greater than G20 at M30, whereas arterial partial pressure of oxygen (PaO2) varied according to the changes in oxygen. The mean intervals of BIS were: for G100, from 68 to 62; for G80, from 71 to 58; for G60, from 72 to 62; for G40 from 76 to 68; and for G20, from 77 to 68. In conclusion, different FiO2 does not impair BIS parameters. However, it is suggested that BIS was able to detect changes in the balance of cerebral blood flow, which was a result of changes in respiratory dynamic by the use of several inspired fractions