Type 1 and type 2 diabetes are ultimately characterized by depleted b-cell mass. Characterization of the molecular pathways that control b-cell proliferation could be harnessed to restore these cells. The homeobox b-cell transcription factor Nkx6.1 induces b-cell proliferation by activating the orphan nuclear receptors Nr4a1 and Nr4a3. Here, we demonstrate that Nkx6.1 localizes to the promoter of the mitotic kinase AURKA (Aurora Kinase A) and induces its expression. Adenovirus mediated overexpression of AURKA is sufficient to induce proliferation in primary rat islets while maintaining glucose stimulated insulin secretion. Furthermore, AURKA is necessary for Nkx6.1 mediated b-cell proliferation as demonstrated by shRNA mediated knock down and pharmacological inhibition of AURKA kinase activity. AURKA preferentially induces DNA replication in b-cells as measured by BrdU incorporation, and enhances the rate of histone H3 phosphorylation in primary b-cells, demonstrating that AURKA induces the replicative and mitotic cell cycle phases in rat b-cells. Finally, overexpression of AURKA results in phosphorylation of the cell cycle regulator p53, which targets p53 for degradation and permits cell cycle progression. These studies define a pathway by which AURKA upregulation by Nkx6.1 results in phosphorylation and degradation of p53, thus removing a key inhibitory factor and permitting engagement of the b-cell proliferation pathway.

Decreased β-cell mass is a hallmark of type 1 and type 2 diabetes. Islet transplantation as a method of diabetes therapy is hampered by the paucity of transplant ready islets. Understanding the pathways controlling islet proliferation may be used to increase functional β-cell mass through transplantation or by enhanced growth of endogenous β-cells. We have shown that the transcription factor Nkx6.1 induces β-cell proliferation by upregulating the orphan nuclear hormone receptors Nr4a1 and Nr4a3. Using expression analysis to define Nkx6.1-independent mechanisms by which Nr4a1 and Nr4a3 induce β-cell proliferation, we demonstrated that cyclin-dependent kinase 5 regulatory subunit 1 (Cdk5r1) is upregulated by Nr4a1 and Nr4a3 but not by Nkx6.1. Overexpression of Cdk5r1 is sufficient to induce primary rat β-cell proliferation while maintaining glucose stimulated insulin secretion. Overexpression of Cdk5r1 in β-cells confers protection against apoptosis induced by etoposide and thapsigargin, but not camptothecin. The Cdk5 kinase complex inhibitor roscovitine blocks islet proliferation, suggesting that Cdk5r1 mediated β-cell proliferation is a kinase dependent event. Overexpression of Cdk5r1 results in pRb phosphorylation, which is inhibited by roscovitine treatment. These data demonstrate that activation of the Cdk5 kinase complex is sufficient to induce β-cell proliferation while maintaining glucose stimulated insulin secretion.

Decreased β‐cell mass is a hallmark of type 1 and type 2 diabetes. The use of islet transplantation as a diabetes therapy is hampered by the relative paucity of transplant ready islets. Greater understanding of the proliferative pathways controlling islet proliferation may be harnessed to increase functional beta‐cell mass through transplantation or by enhanced growth of endogenous beta‐cells. We have shown that the β‐cell transcription factor Nkx6.1 induces β‐cell proliferation by upregulating the orphan nuclear hormone receptors Nr4a1 and Nr4a3. Using expression analysis to elucidate the Nkx6.1 independent mechanism by which Nr4a1 and Nr4a3 induce beta‐cell proliferation, we demonstrated that cyclin‐dependent kinase 5 regulatory subunit 1 (Cdk5r1) is upregulated by Nr4a1 and Nr4a3 and not Nkx6.1. Adenovirus mediated overexpression of Cdk5r1 is sufficient to induce proliferation in primary rat islets. The observed proliferation is primarily in beta‐cells. Glucose stimulated insulin secretion is maintained with Cdk5r1 overexpression. The Cdk5 inhibitor roscovitine blocks islet proliferation, suggesting that Nr4a mediated β‐cell proliferation is a kinase dependent event. Overexpression of Cdk5r1 results in pRb phosphorylation, which is inhibited by roscovitine treatment. These data demonstrate that activation of the cdk5 complex is sufficient to induce β‐cell proliferation while maintaining glucose stimulated insulin secretion.