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dc.contributor.advisorLaura C. Alonso
dc.contributor.authorStamateris, Rachel E.
dc.date2022-08-11T08:08:39.000
dc.date.accessioned2022-08-23T16:02:50Z
dc.date.available2022-08-23T16:02:50Z
dc.date.issued2021-05-13
dc.date.submitted2021-06-08
dc.identifier.doi10.13028/padg-ny31
dc.identifier.urihttp://hdl.handle.net/20.500.14038/31368
dc.description.abstractStrategies aimed at expanding functional beta cell mass remain a prime goal of diabetes research. Both the insulin signaling pathway, as well as the G1/S transition of the cell cycle are critically important for the maintenance of beta cell mass. We previously demonstrated in a mouse model of diabetes, insulin receptor substrate 2 (Irs2) deficient mice, that beta cell failure was attributed to reduced islet expression of Cyclin D2, and that overexpressing Cyclin D2 rescued proliferation in Irs2 deficient beta cells in vitro. Since Cyclin D2 partners with CDK4 to drive cell cycle progression, we hypothesized that an activated form of CDK4, Cdk4-R24C (resistant to inhibition by the INK4A cell cycle inhibitor p16), would rescue the in vivo proliferation defect in Irs2 deficient mice. Interestingly, Irs2 knockout mice with the active Cdk4 R24C allele, displayed rescued blood glucose, and normalized glucose tolerance, without affecting peripheral insulin resistance. I found that both and beta cell mass and proliferation were rescued in vivo, contributing to the rescue of glucose tolerance. Interestingly, the dedifferentiated phenotype of Irs2 knockout islets (ALDH1A3+ cells, nuclear FOXO1 and suppressed PDX1) was completely restored with the active Cdk4 allele, suggesting that CDK4 may play a role in promoting beta cell differentiation. Utilizing various in vitro models where FOXO1 represses Pdx1, overexpression of CDK4/CyclinD2 was consistently able to rescue the FOXO1-mediated repression of Pdx1, without significant impacts on FOXO1 subcellular localization. These results suggested that FOXO1 regulation in the beta cell is more complex than previously described, and also suggested that CDK4/Cyclin D2 may be instead modulating the acetylation status of FOXO1, impacting its transcriptional activity. To this end, inhibiting histone acetylate transferases (HATs) partially rescued FOXO1-mediated Pdx1 suppression, while inhibiting histone deacetylase enzymes (HDACs) showed the reverse effect of trending towards blocking the Cyclin D2/CDK4-mediated rescue of Pdx1. Finally, I found that CDK4/Cyclin D2 increases phosphorylation of sirtuin 1 (SIRT1), an HDAC that modulates the acetylation status, and transcriptional activity of FOXO1, and that CDK4/Cyclin D2 promotes FOXO1 degradation. In sum, we conclude that activated CDK4 rescues beta cell failure due to IRS2 deficiency through multiple mechanisms related to not only cell cycle regulation but also to beta cell differentiation status, primarily through modulation of FOXO1 transcriptional activity.
dc.language.isoen_US
dc.rightsCopyright is held by the author, with all rights reserved.
dc.subjectbeta cell
dc.subjectdiabetes
dc.subjectinsulin
dc.subjectproliferation
dc.subjectdifferentiation
dc.subjectIRS2
dc.subjectFOXO1
dc.subjectPDX1
dc.subjectHDACs
dc.subjectSIRT1
dc.subjectBiology
dc.subjectCell Biology
dc.subjectEndocrine System Diseases
dc.titleCDK4 Rescues Diabetes in IRS2-Deficient Mice: Exploring Novel Roles of a Cell Cycle Regulator in Promoting Beta Cell Differentiation
dc.typeDoctoral Dissertation
dc.identifier.legacyfulltexthttps://escholarship.umassmed.edu/cgi/viewcontent.cgi?article=2148&context=gsbs_diss&unstamped=1
dc.identifier.legacycoverpagehttps://escholarship.umassmed.edu/gsbs_diss/1138
dc.legacy.embargo2023-06-08T00:00:00-07:00
dc.identifier.contextkey23263005
html.description.abstract<p>Strategies aimed at expanding functional beta cell mass remain a prime goal of diabetes research. Both the insulin signaling pathway, as well as the G1/S transition of the cell cycle are critically important for the maintenance of beta cell mass. We previously demonstrated in a mouse model of diabetes, insulin receptor substrate 2 (<em>Irs2</em>) deficient mice, that beta cell failure was attributed to reduced islet expression of Cyclin D2, and that overexpressing Cyclin D2 rescued proliferation in <em>Irs2</em> deficient beta cells <em>in vitro</em>. Since Cyclin D2 partners with CDK4 to drive cell cycle progression, we hypothesized that an activated form of CDK4, <em>Cdk4-R24C</em> (resistant to inhibition by the INK<sup>4A</sup> cell cycle inhibitor p16), would rescue the <em>in vivo</em> proliferation defect in <em>Irs2</em> deficient mice.<em> </em>Interestingly, <em>Irs2</em> knockout mice with the active <em>Cdk4 R24C</em> allele, displayed rescued blood glucose, and normalized glucose tolerance, without affecting peripheral insulin resistance. I found that both and beta cell mass and proliferation were rescued <em>in vivo</em>, contributing to the rescue of glucose tolerance. Interestingly, the dedifferentiated phenotype of <em>Irs2 </em>knockout islets (ALDH1A3+ cells, nuclear FOXO1 and suppressed PDX1) was completely restored with the active <em>Cdk4 </em>allele, suggesting that CDK4 may play a role in promoting beta cell differentiation. Utilizing various <em>in vitro </em>models where FOXO1 represses <em>Pdx1</em>, overexpression of CDK4/CyclinD2 was consistently able to rescue the FOXO1-mediated repression of <em>Pdx1</em>, without significant impacts on FOXO1 subcellular localization. These results suggested that FOXO1 regulation in the beta cell is more complex than previously described, and also suggested that CDK4/Cyclin D2 may be instead modulating the acetylation status of FOXO1, impacting its transcriptional activity. To this end, inhibiting histone acetylate transferases (HATs) partially rescued FOXO1-mediated <em>Pdx1 </em>suppression, while inhibiting histone deacetylase enzymes (HDACs) showed the reverse effect of trending towards blocking the Cyclin D2/CDK4-mediated rescue of <em>Pdx1.</em> Finally, I found that CDK4/Cyclin D2 increases phosphorylation of sirtuin 1 (SIRT1), an HDAC that modulates the acetylation status, and transcriptional activity of FOXO1, and that CDK4/Cyclin D2 promotes FOXO1 degradation. In sum, we conclude that activated CDK4 rescues beta cell failure due to IRS2 deficiency through multiple mechanisms related to not only cell cycle regulation but also to beta cell differentiation status, primarily through modulation of FOXO1 transcriptional activity.</p>
dc.identifier.submissionpathgsbs_diss/1138
dc.contributor.departmentMedicine/Diabetes
dc.description.thesisprogramInterdisciplinary Graduate Program
dc.identifier.orcid0000-0003-2030-1487


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