Human cytomegalovirus infection blocks cell cycle progression in primary human fibroblasts. The arrest occurs at multiple phases of the cell cycle, including a point late in the G1 compartment. While studying cell cycle perturbation induced by HCMV, we have observed that human embryonic lung fibroblasts infected by HCMV are strongly positive for neutral ß-galactosidase (SA-ß-gal), a widely accepted marker of senescence. Infectious foci became intensely blue compared to uninfected cells that appeared not stained. BrdU incorporation experiments revealed that infected cells were growth arrested, showing only a non uniform BrdU staining typical of infected cells. The expression of several cell cycle regulatory proteins was examined by both immunoblotting and immunofluorescence analysis. Infected cells accumulated high levels of both p53 and pRb. Among the cyclin dependent kinase inhibitors, a strong induction of p16INK4 was observed, with no variation of p21CIP/WAF1 expression, compared to mock infected cells. Interestingly, p21CIP/WAF1 staining was predominantly in the cytoplasm, indicating a potential cytoplasmic translocation or retention of nuclear p21CIP/WAF1. The induction of a senescence-like phenotype seems to be typical of primary diploid fibroblasts cells, since no SA-ß-gal activity was detected in infected astrocytoma U373 cells, a tumor cell line permissive for HCMV infection. In these cells p53, pRb and p16INK4 were not upregulated following HCMV infection, although cytoplasmic staining for p21CIP/WAF1 was also observed. Altogether these results demonstrate a premature senescence-like phenotype following HCMV infection that may have important implications in the proliferation process of normal cells.
A premature senescence-like phenotype is induced by HCMV infection.
Noris E;
2001
Abstract
Human cytomegalovirus infection blocks cell cycle progression in primary human fibroblasts. The arrest occurs at multiple phases of the cell cycle, including a point late in the G1 compartment. While studying cell cycle perturbation induced by HCMV, we have observed that human embryonic lung fibroblasts infected by HCMV are strongly positive for neutral ß-galactosidase (SA-ß-gal), a widely accepted marker of senescence. Infectious foci became intensely blue compared to uninfected cells that appeared not stained. BrdU incorporation experiments revealed that infected cells were growth arrested, showing only a non uniform BrdU staining typical of infected cells. The expression of several cell cycle regulatory proteins was examined by both immunoblotting and immunofluorescence analysis. Infected cells accumulated high levels of both p53 and pRb. Among the cyclin dependent kinase inhibitors, a strong induction of p16INK4 was observed, with no variation of p21CIP/WAF1 expression, compared to mock infected cells. Interestingly, p21CIP/WAF1 staining was predominantly in the cytoplasm, indicating a potential cytoplasmic translocation or retention of nuclear p21CIP/WAF1. The induction of a senescence-like phenotype seems to be typical of primary diploid fibroblasts cells, since no SA-ß-gal activity was detected in infected astrocytoma U373 cells, a tumor cell line permissive for HCMV infection. In these cells p53, pRb and p16INK4 were not upregulated following HCMV infection, although cytoplasmic staining for p21CIP/WAF1 was also observed. Altogether these results demonstrate a premature senescence-like phenotype following HCMV infection that may have important implications in the proliferation process of normal cells.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
