To explore how Mier1 affiects high-fat diet-fed, aging and adipose tissue Lipe knockout mice liver regeneration, we specifically knocked out liver Mier1 in high-fat diet-fed, aging and adipose tissue Lipe knockout mice through adeno-associated virus (AAV). The mice we used were knocked in a Cre-induced Cas9 expression cassette. Through tail vein injection, we delivered the AAV expressing Cre-recombinase under TBG promoter, and sgRNA targeting Mier1 (AAV-Mier1 sgRNA), into the adult Cas9 knockin mice to knock out the Mier1 gene in liver. AAV vectors with no sgRNA inserted (AAV-Cre) were used in control animals. To assess the role of MIER1 in liver regeneration, we performed 70% partial hepatectomy on high-fat diet-fed, aging and adipose tissue Lipe knockout mice. We then performed RNA sequencing analysis on liver tissues collected from control and Mier1 ko groups at 0 h, 48h or 36 h after partial hepatectomy. RNA-seq analysis showed that after partial hepatectomy, MIER1 loss caused significant upregulation of genes enriched in cell proliferation relevant pathways. Overall design: For liver regeneration analysis in chronic high-fat diet-fed mice, 6-week-old Cas9 knockin mice were fed with normal chow diet or HFD for 8 weeks to induce hepatic steatosis. After 5 weeks on HFD, AAV8 vectors were administered by tail vein injection to deplete MIER1 in hepatocytes. For liver regeneration analysis in aging mice, about one-year-old Cas9 knockin mice were administered with AAV8 vectors expression Cre recombinase and Mier1 sgRNA to deplete MIER1 in liver cells. Lipe loxp/loxp mice were generated by a homologous recombination and classic embryonic stem cell targeting strategy. Loxp sequences were inserted flanking the second exon. Adipose tissue-specific Lipe knockout mice were generated by crossing Lipe loxp/loxp mice with Adiponectin-Cre mice. Lipe loxp/loxp mice were then bred with Lipe loxp/loxp mice: Cre/+ animals to generate Lipe loxp/loxp mice (used as control) and Lipeloxp/loxp: Cre/+ mice (Lipe-AKO) as adipose-specific knockout animals. To further deplete MIER1 in the Lipe-AKO animals, Cas9 knockin mice were bred with the Lipe-AKO mice to generate the Lipe-AKO; Cas9 animals, which were then subjected to MIER1 depletion via treatment with AAV-delivered Mier1 sgRNA. Animals were subjected to PHx surgery for liver regeneration analysis at around 8-11 weeks’ old. Each RNA-seq sample contained pooled RNA from three biological replicas that were mixed with an equal mass of RNA. Two samples from each group were included for sequencing.