Hao Yin1, Roman L Bogorad1, Carmen Barnes2, Stephen Walsh1, Iris Zhuang3, Hidenori Nonaka4, Vera Ruda1, Satya Kuchimanchi2, Lubomir Nechev2, Akin Akinc2, Wen Xue5, Marino Zerial4, Robert Langer6, Daniel G Anderson7, Victor Koteliansky8. 1. David H. Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA 02139, USA. 2. Alnylam Pharmaceuticals, Cambridge, MA 02142, USA. 3. David H. Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA. 4. Max Planck Institute of Molecular Cell Biology and Genetics (MPI-CBG), Dresden 01307, Germany. 5. RNA Therapeutics Institute and Program in Molecular Medicine, University of Massachusetts Medical School, Worcester, MA 01605, USA. 6. David H. Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Harvard-MIT Division of Health Sciences & Technology, Cambridge, MA 02139, USA; Institute of Medical Engineering and Science, Massachusetts Institute of Technology, Cambridge, MA 02139, USA. 7. David H. Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Harvard-MIT Division of Health Sciences & Technology, Cambridge, MA 02139, USA; Institute of Medical Engineering and Science, Massachusetts Institute of Technology, Cambridge, MA 02139, USA. Electronic address: dgander@mit.edu. 8. Skolkovo Institute of Science and Technology, Skolkovo 143025, Russia; Department of Chemistry, M.V. Lomonosov Moscow State University, Leninskie Gory 119991, Russia. Electronic address: V.Kotelianski@Skoltech.RU.
Abstract
BACKGROUND & AIMS: The Hippo pathway controls organ size through a negative regulation of the transcription co-activator Yap1. The overexpression of hyperactive mutant Yap1 or deletion of key components in the Hippo pathway leads to increased organ size in different species. Analysis of interactions of this pathway with other cellular signals corroborating organ size control is limited in part due to the difficulties associated with development of rodent models. METHODS: Here, we develop a new model of reversible induction of the liver size in mice using siRNA-nanoparticles targeting two kinases of the Hippo pathway, namely, mammalian Ste20 family kinases 1 and 2 (Mst1 and Mst2), and an upstream regulator, neurofibromatosis type II (Nf2). RESULTS: The triple siRNAs nanoparticle-induced hepatomegaly in mice phenocopies one observed with Mst1(-/-)Mst2(-/-) liver-specific depletion, as shown by extensive proliferation of hepatocytes and activation of Yap1. The simultaneous co-treatment with a fourth siRNA nanoparticle against Yap1 fully blocked the liver growth. Hippo pathway-induced liver enlargement is associated with p53 activation, evidenced by its accumulation in the nuclei and upregulation of its target genes. Moreover, injections of the triple siRNAs nanoparticle in p53(LSL/LSL) mice shows that livers lacking p53 expression grow faster and exceed the size of livers in p53 wild-type animals, indicating a role of p53 in controlling Yap1-induced liver growth. CONCLUSION: Our data show that siRNA-nanoparticulate manipulation of gene expression can provide the reversible control of organ size in adult animals, which presents a new avenue for the investigation of complex regulatory networks in liver.
BACKGROUND & AIMS: The Hippo pathway controls organ size through a negative regulation of the transcription co-activator Yap1. The overexpression of hyperactive mutant Yap1 or deletion of key components in the Hippo pathway leads to increased organ size in different species. Analysis of interactions of this pathway with other cellular signals corroborating organ size control is limited in part due to the difficulties associated with development of rodent models. METHODS: Here, we develop a new model of reversible induction of the liver size in mice using siRNA-nanoparticles targeting two kinases of the Hippo pathway, namely, mammalianSte20 family kinases 1 and 2 (Mst1 and Mst2), and an upstream regulator, neurofibromatosis type II (Nf2). RESULTS: The triple siRNAs nanoparticle-induced hepatomegaly in mice phenocopies one observed with Mst1(-/-)Mst2(-/-) liver-specific depletion, as shown by extensive proliferation of hepatocytes and activation of Yap1. The simultaneous co-treatment with a fourth siRNA nanoparticle against Yap1 fully blocked the liver growth. Hippo pathway-induced liver enlargement is associated with p53 activation, evidenced by its accumulation in the nuclei and upregulation of its target genes. Moreover, injections of the triple siRNAs nanoparticle in p53(LSL/LSL) mice shows that livers lacking p53 expression grow faster and exceed the size of livers in p53 wild-type animals, indicating a role of p53 in controlling Yap1-induced liver growth. CONCLUSION: Our data show that siRNA-nanoparticulate manipulation of gene expression can provide the reversible control of organ size in adult animals, which presents a new avenue for the investigation of complex regulatory networks in liver.
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