Cell cycle arrest by human cytomegalovirus 86-kDa IE2 protein resembles premature senescence

Primary human embryo lung fibroblasts and adult diploid fibroblasts infected by the human cytomegalovirus (HCMV) display beta-galactosidase (beta-Gal) activity at neutral pH (senescence-associated beta-Gal [SA-beta-Gal] activity) and overexpression of the plasminogen activator inhibitor type 1 (PAI-1) gene, two widely recognized markers of the process designated premature cell senescence. This activity is higher when cells are serum starved for 48 h before infection, a process that speeds and facilitates HCMV infection but that is insufficient by itself to induce senescence. Fibroblasts infected by HCMV do not incorporate bromodeoxyuridine, a prerequisite for the formal definition of senescence. At the molecular level, cells infected by HCMV, beside the accumulation of large amounts of the cell cycle regulators p53 and pRb, the latter in its hyperphosphorylated form, display a strong induction of the cyclin-dependent kinase inhibitor (cdki) p16(INK4a), a direct effector of the senescence phenotype in fibroblasts, and a decrease of the cdki p21(CIP1/WAF). Finally, a replicative senescence state in the early phases of infection significantly increased the number of cells permissive to virus infection and enhanced HCMV replication. HCMV infection assays carried out in the presence of phosphonoformic acid, which inhibits the virus DNA polymerase and the expression of downstream genes, indicated that immediate-early and/or early (alpha) genes are sufficient for the induction of SA-beta-Gal activity. When baculovirus vectors expressing HCMV IE1-72 or IE2-86 proteins were inoculated into fibroblasts, the increase of p16(INK4a) (observed predominantly with IE2-86) was similar to that observed with the whole virus, as was the induction of SA-beta-Gal activity, suggesting that the viral IE2 gene leads infected cells into senescence. Altogether our results demonstrate for the first time that HCMV, after arresting the cell cycle and inhibiting apoptosis, triggers the cellular senescence program, probably through the p16(INK4a) and p53 pathways.     

Validity of the reduced version of the Morningness-Eveningness Questionnaire

Sleep and Biological Rhythms - The aim of the present work was to validate the reduced version of the Morningness-Eveningness Questionnaire (MEQr) using circadian motor activity as external...     

Isolation and properties of a thialysine-resistant clone of CHO cells

A variant clone of Chinese hamster ovary (CHO) cells resistant to thialysine has been isolated. It maintains the phenotypic properties even after 250 generations in medium without thialysine. Growth rate, cell viability and protein synthesis rate of the variant are much less affected by thialysine than the parental strain. In both the parental strain and the variant, thialysine acts in competition with lysine as indicated by the fact that all thialysine effects can be completely reversed by lysine.     

Melanins production from enkephalins by tyrosinase.

Leu-enkephalin and Met-enkephalin are oxidized in vitro by mushroom and sepia tyrosinase giving rise to synthetic melanins whose production is dependent on incubation time and on enzyme concentration. The Enk-melanins formed are acid-insoluble brownish or reddish pigments showing a continuous absorbance in the visible region when dissolved in basic solution. The presence of the amino acid chain makes them fully soluble in pH 7.4 0.05 M phosphate buffer and methanol.     

Platelet adhesiveness and aggregation in combined factor V and factor VIII deficiency and in combined factor VII and factor VIII deficiency.

Platelet aggregation and adhesiveness were studied in 3 patients with combined factor V and factor VIII deficiency and in 3 patients with combined factor VII and factor VIII deficiency. The first three patients belonged to three different kindreds whereas the second group belonged to the same kindred. Serotonin C14 uptake and release was also found to be normal in these patients. These studies indicate that platelet function is normal in combined defects of factor VIII. These findings were in agreement with the presence of a normal bleeding time and a normal factor VIII antigen level in all these patients.