A microRNA-21 surge facilitates rapid cyclin D1 translation and cell cycle progression in mouse liver regeneration
Raymond Ng, Guisheng Song, Garrett R. Roll, Niels M. Frandsen, Holger Willenbring
Raymond Ng, Guisheng Song, Garrett R. Roll, Niels M. Frandsen, Holger Willenbring
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Research Article Hepatology

A microRNA-21 surge facilitates rapid cyclin D1 translation and cell cycle progression in mouse liver regeneration

Abstract

MicroRNA-21 (miR-21) is thought to be an oncomir because it promotes cancer cell proliferation, migration, and survival. miR-21 is also expressed in normal cells, but its physiological role is poorly understood. Recently, it has been found that miR-21 expression is rapidly induced in rodent hepatocytes during liver regeneration after two-thirds partial hepatectomy (2/3 PH). Here, we investigated the function of miR-21 in regenerating mouse hepatocytes by inhibiting it with an antisense oligonucleotide. To maintain normal hepatocyte viability and function, we antagonized the miR-21 surge induced by 2/3 PH while preserving baseline expression. We found that knockdown of miR-21 impaired progression of hepatocytes into S phase of the cell cycle, mainly through a decrease in levels of cyclin D1 protein, but not Ccnd1 mRNA. Mechanistically, we discovered that increased miR-21 expression facilitated cyclin D1 translation in the early phase of liver regeneration by relieving Akt1/mTOR complex 1 signaling (and thus eIF-4F–mediated translation initiation) from suppression by Rhob. Our findings reveal that miR-21 enables rapid hepatocyte proliferation during liver regeneration by accelerating cyclin D1 translation.

Authors

Raymond Ng, Guisheng Song, Garrett R. Roll, Niels M. Frandsen, Holger Willenbring

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Figure 5

miR-21 induction in liver regeneration decreases Rhob expression by direct targeting.

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miR-21 induction in liver regeneration decreases Rhob expression by dire...
(A and B) Inverse correlation of miR-21 and Rhob expression levels after 2/3 PH. miR-21 induction after 2/3 PH was associated with decreased Rhob mRNA (A) and Rhob protein (B) levels. Numbers indicate protein level relative to time point 0 hours after 2/3 PH. Gapdh was analyzed as a loading control. (C) High prediction score and favorable binding energy suggested that miR-21 targets the 3′ UTR of Rhob. The complementary sequence in the Rhob 3′ UTR and the seed region of miR-21 (red letters) is conserved among mammalian species. (D) Liver Rhob mRNA levels increased 6 hours after miR-21–ASO injection. Control mice were injected with carrier. At least 3 mice were analyzed per time point and treatment. (E–H) Direct inhibition of Rhob by miR-21. (E) Transfection of miR-21 mimic into Hepa1,6 cells decreased Rhob mRNA. (F) The activity of a luciferase reporter gene linked to the Rhob 3′ UTR was inhibited by miR-21 mimic in a dose-dependent fashion. Mutation of the Rhob 3′ UTR sequence complimentary to the miR-21 seed sequence prevented inhibition by miR-21 mimic. (G) Transfection of miR-21 inhibitor into Hepa1,6 cells increased Rhob mRNA. (H) miR-21 inhibitor effectively restored the activity of a luciferase reporter gene linked to the Rhob 3′ UTR in Hepa1,6 cells transfected with miR-21 mimic. Mutation of the Rhob 3′ UTR sequence complimentary to the miR-21 seed sequence prevented this effect. Data represent mean ± SEM. *P < 0.05.