Hepatocyte differentiation is controlled by liver-enriched transcription factors (LETFs). We investigated whether LETFs control microRNA expression during development and whether this control is required for hepatocyte differentiation.

Using in vivo DNA binding assays, we identified miR-122 as a direct target of the LETF hepatocyte nuclear factor (HNF) 6. The role and mechanisms of the HNF6–miR-122 gene cascade in hepatocyte differentiation were studied in vivo and in vitro by gain-of-function and loss-of-function experiments, using developing mice and zebrafish as model organisms.

HNF6 and its paralog Onecut2 are strong transcriptional stimulators of miR-122 expression. Specific levels of miR-122 were required for proper progression of hepatocyte differentiation; miR-122 stimulated the expression of hepatocyte-specific genes and most LETFs, including HNF6. This indicates that HNF6 and miR-122 form a positive feedback loop. Stimulation of hepatocyte differentiation by miR-122 was lost in HNF6-null mice, revealing that a transcription factor can mediate microRNA function. All hepatocyte-specific genes whose expression was stimulated by miR-122 bound HNF6 in vivo, confirming their direct regulation by this factor.

Hepatocyte differentiation is directed by a positive feedback loop that includes a transcription factor (HNF6) and a microRNA (miR-122) that are specifically expressed in liver. These findings could lead to methods to induce differentiation of hepatocytes in vitro and improve our understanding of liver cell dedifferentiation in pathologic conditions.