The Lu Laboratory currently has five active National Institutes of Health (NIH)-funded research programs. The Lu Lab’s longest-running program focuses on the regulation of hepatic glutathione (GSH) synthesis. GSH is vital in defense against oxidative stress. Over the past 20 years, the Lu Laboratory has shown how the GSH synthetic enzymes are regulated transcriptionally and posttranscriptionally. More recently the Lu Lab has described dysregulation of GSH synthesis under several pathological conditions of the liver. Elucidating molecular mechanisms of GSH may lead to novel strategies to enhance hepatic GSH level and ameliorate liver injury, including fibrosis. One such example is chronic cholestasis (Figure 1).
Figure 1. Chronic cholestasis triggers c-Myc up-regulation, which induces miR-27a/b that in turn lowers the expression of its target mRNAs, prohibitin 1 (Phb1) and Nrf2. In normal liver the expression of glutamate-cysteine ligase (GCL, made up of catalytic or GCLC and modifier or GCLM subunits), the rate-limiting enzyme in GSH synthesis, is positively regulated by Nrf2-mediated trans-activation of the anti-oxidant response element (ARE). Nrf2 heterodimerize with other proteins such as small Mafs or Jun to trans-activate ARE. We found in addition to these, c-Myc and Phb1 also interact with Nrf2 at ARE. While c-Myc serves as a co-repressor, Phb1 is a co-activator. During cholestasis, induction of MafG and c-Maf also occur and they form complexes to inhibit ARE. These changes all work together to down-regulate GCLC and GCLM expression, GSH level, further exacerbating liver fibrosis. From Yang, et al, Antioxid Redox Signal. 2015;22:259-274.
The Lu Lab’s second research program examines the hepatic regulation of methionine adenosyltransferases (MATs). MAT is a critical cellular enzyme catalyzing S-adenosylmethionine (SAMe) formation, the principal biologic methyl donor, a precursor for polyamine synthesis and a major precursor for hepatic GSH through the transsulfuration pathway. Two genes encode for MAT: MAT1A is expressed in normal differentiated liver, and MAT2A is expressed in all extrahepatic tissues as well as in fetal liver. As the liver matures, MAT2A is replaced by MAT1A. The Lu Laboratory was the first to describe a switch from MAT1A to MAT2A expression in human hepatocellular carcinoma (HCC). This is pathogenetically important because MAT2A expression provides a growth advantage. Recently, the Lu Lab has found that a third MAT gene, MAT2B, encodes for splicing variants that not only regulate MAT2A-encoded isoenzyme, but also serve as scaffold complexes that regulate multiple steps in the Ras-Raf-MEK-ERK signaling pathway. This research is now focused on answering important questions these key observations pose: What are the molecular mechanisms responsible for the switch in MAT gene expression in liver cancer? How do MAT proteins control growth and death pathways?
A third research program in the Lu Laboratory examines the role of SAMe in liver function and injury. Hepatic MAT activity decreases in cirrhosis of all causes. This is due to inactivation of the enzyme as well as decreased MAT1A expression. In collaboration with José Mato, PhD, the Lu Lab developed the Mat1a knockout (KO) murine model. Not only do these KO animals develop spontaneous steatohepatitis, but over time they also manifest a high frequency of HCC (Figure 2). This model proves the importance of maintaining normal SAMe levels and MAT1A expression in the liver. Recently, we have also discovered that if hepatic SAMe is not properly metabolized, as in glycine N-methyltransferase KO mice, liver injury and cancer ensue. Based on these data, we have been elucidating the mechanisms by which SAMe modulates liver injury, including development of nonalcoholic fatty liver disease and cancer.
Copyright (2001) National Academy of Sciences, U.S.A.
Figure 2. Phenotype of the Mat1a knockout (KO) mouse model. Three-month old KO mice have larger livers but are otherwise normal. KO mice developed massive fatty liver after six days of choline deficient diet, steatohepatitis on a normal diet by eight months and HCC by 18 months. KO mice also have increased oxidative stress and abnormalities in multiple oncogenic signaling pathways. From PNAS 2001;98:5560-5565 and FASEB J 2002; doi: 10.1096/fj.02-0078fje. Reviewed in Physiol Rev. 2012;92:1515-1542.
A fourth research program, funded by the National Cancer Institute, is to elucidate the role of prohibitin 1 (Phb1) in liver injury and cancer. Mat1a KO mice have reduced Phb1 expression from birth to development of non-alcoholic steatohepatitis. To better understand the role of Phb1 in liver physiology, the Lu Laboratory generated the liver-specific Phb1 KO mouse model and showed these mice developed severe liver injury, fibrosis, increased oval cell population and preneoplastic changes as early as three weeks (Figure 3) and multifocal HCC by 35 weeks. This research program is elucidating the molecular mechanisms in order to gain a better understanding of Phb1’s functions in liver pathobiology.
Figure 3. Increased staining for oval cells (OV6, green) and glutathione-S-transferase Pi (GSTP), both pre-neoplastic markers, in the Phb1 KO hepatocytes at 3 weeks. From Hepatology 2010;52:2096-2108.
The newest research program funded by the National Institute of Diabetes and Digestive and Kidney Diseases was launched in July 2015 with new collaborations established after the Lu Laboratory moved to Cedars-Sinai. At Cedars-Sinai, the Lu Laboratory will investigate how altered SAMe level affects protein post-translational modifications, with the help of Jennifer Van Eyk, PhD, and examine whether SAMe may be useful to prevent HCC recurrence in the animal model.