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Targeting the cell cycle inhibitor p57Kip2 promotes adult human ? cell replication.

ABSTRACT: Children with focal hyperinsulinism of infancy display a dramatic, non-neoplastic clonal expansion of ? cells that have undergone mitotic recombination, resulting in paternal disomy of part of chromosome 11. This disomic region contains imprinted genes, including the gene encoding the cell cycle inhibitor p57Kip2 (CDKN1C), which is silenced as a consequence of the recombination event. We hypothesized that targeting p57Kip2 could stimulate adult human ? cell replication. Indeed, when we suppressed CDKN1C expression in human islets obtained from deceased adult organ donors and transplanted them into hyperglycemic, immunodeficient mice, ? cell replication increased more than 3-fold. The newly replicated cells retained properties of mature ? cells, including the expression of ? cell markers such as insulin, PDX1, and NKX6.1. Importantly, these newly replicated cells demonstrated normal glucose-induced calcium influx, further indicating ? cell functionality. These findings provide a molecular explanation for the massive ? cell replication that occurs in children with focal hyperinsulinism. These data also provided evidence that ? cells from older humans, in which baseline replication is negligible, can be coaxed to re-enter and complete the cell cycle while maintaining mature ? cell properties. Thus, controlled manipulation of this pathway holds promise for the expansion of ? cells in patients with type 2 diabetes.

SUBMITTER: Avrahami D

PROVIDER: S-EPMC3904605 | biostudies-literature | 2014 Feb

REPOSITORIES: biostudies-literature

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Targeting the cell cycle inhibitor p57Kip2 promotes adult human β cell replication.

Avrahami Dana D Li Changhong C Yu Ming M Jiao Yang Y Zhang Jia J Naji Ali A Ziaie Seyed S Glaser Benjamin B Kaestner Klaus H KH

The Journal of clinical investigation 20140116 2

Children with focal hyperinsulinism of infancy display a dramatic, non-neoplastic clonal expansion of β cells that have undergone mitotic recombination, resulting in paternal disomy of part of chromosome 11. This disomic region contains imprinted genes, including the gene encoding the cell cycle inhibitor p57Kip2 (CDKN1C), which is silenced as a consequence of the recombination event. We hypothesized that targeting p57Kip2 could stimulate adult human β cell replication. Indeed, when we suppresse ...[more]

PMID: 24430183

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Project description:DNA replication of E1-deleted first-generation adenoviruses (AdV) in cultured cancer cells has been reported repeatedly and it was suggested that certain cellular proteins could functionally compensate for E1A, leading to the expression of the early region 2 (E2)-encoded proteins and subsequently virus replication. Referring to this, the observation was named E1A-like activity. In this study, we investigated different cell cycle inhibitors with respect to their ability to increase viral DNA replication of dl70-3, an E1-deleted adenovirus. Our analyses of this issue revealed that in particular inhibition of cyclin-dependent kinases 4/6 (CDK4/6i) increased E1-independent adenovirus E2-expression and viral DNA replication. Detailed analysis of the E2-expression in dl70-3 infected cells by RT-qPCR showed that the increase in E2-expression originated from the E2-early promoter. Mutations of the two E2F-binding sites in the E2-early promoter (pE2early-LucM) caused a significant reduction in E2-early promoter activity in trans-activation assays. Accordingly, mutations of the E2F-binding sites in the E2-early promoter in a virus named dl70-3/E2Fm completely abolished CDK4/6i induced viral DNA replication. Thus, our data show that E2F-binding sites in the E2-early promoter are crucial for E1A independent adenoviral DNA replication of E1-deleted vectors in cancer cells. IMPORTANCE E1-deleted AdV vectors are considered replication deficient and are important tools for the study of virus biology, gene therapy, and large-scale vaccine development. However, deletion of the E1 genes does not completely abolish viral DNA replication in cancer cells. Here, we report, that the two E2F-binding sites in the adenoviral E2-early promoter contribute substantially to the so-called E1A-like activity in tumor cells. With this finding, on the one hand, the safety profile of viral vaccine vectors can be increased and, on the other hand, the oncolytic property for cancer therapy might be improved through targeted manipulation of the host cell.

Dataset Information

Targeting the cell cycle inhibitor p57Kip2 promotes adult human ? cell replication.

Publications

Targeting the cell cycle inhibitor p57Kip2 promotes adult human β cell replication.

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