UC San Diego

A prevailing hypothesis in type 1 Diabetes (T1D) research is that replacing pancreatic β cells destroyed by autoimmune attack will restore euglycemia. Human embryonic stem cells (hESCs) have the potential to differentiate into any somatic cell type, including β cells, making them ideal candidates for cell replacement therapies. Although β cells derived from hESCs are currently being tested in T1D patients, a detailed understanding of the molecular mechanisms that regulate hESC differentiation at key points remains elusive. MicroRNAs are small, regulatory non-coding RNAs that play an essential role in directing hESC differentiation. Previously, microRNA653 (miR-653) has been shown to be upregulated during the transition of hESCs from pluripotency to definitive endoderm (DE). In the current study, the beta isoform of protein kinase C (PKCβ) was identified as a target of miR-653 in hESC. Consistent with its role in other cells, PKCβ was found to localize to centromeres in dividing hESCs and phosphorylate numerous substrates. Misexpression of miR-653 in hESCs resulted in decreased expression of PKCβ, increased proliferation, and decreased expression of the transcription factor Oct4 (Octamer-binding protein 4). Subsequently, we found that Oct4 and PKCβII, an isozyme of PKCβ, directly interact and that Oct4 is a PKCβII substrate. Overexpression of miR-653 in early hESC differentiation resulted in a loss of expression of pancreatic endocrine markers, suggesting that the PKCβII/Oct4 axis plays an important role in determining cell fate. Taken together, these studies provide insight into the role of miR-653 in driving hESC fate decisions.