Proliferation is the major component for maintenance of β-cell mass in adult animals. Activation of phosphoinositide 3-kinase/Akt-kinase pathway is a critical regulator of β-cell mass. Pancreatic β-cell overexpression of constitutively active Akt in mice (caAktTg) resulted in marked expansion of β-cell mass by increase in β-cell proliferation and size. The current studies provide new insights into the molecular mechanisms involved in β-cell proliferation by Akt. Proliferation of β-cells in caAktTg was associated with increased cyclin D1, cyclin D2, and p21 levels and cyclin-dependent kinase-4 (cdk4) activity. To determine the role of cdk4 in β-cell proliferation induced by Akt, we generated caAktTg mice that were homozygous, heterozygous, or nullizygous for cdk4. The results of these studies showed that deletion of one cdk4 allele significantly reduced β-cell expansion in caAktTg mice by decreased proliferation. CaAktTg mice deficient in cdk4 developed β-cell failure and diabetes. These experiments suggest that Akt induces β-cell proliferation in a cdk4-dependent manner by regulation of cyclin D1, cyclin D2, and p21 levels. These data also indicate that alteration in levels of these cell cycle components could affect the maintenance of β-cell mass in basal states and the adaptation of β-cells to pathological states resulting in diabetes.