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http://icahn.mssm.edu/departments-and-institutes/developmental-and-regenerative-biology
 

Sadler Lab - Research

Overview.

    The liver accomplishes an amazing array of physiological tasks, including energy homeostasis, xenobiotic metabolism, serum protein secretion and bile production. Despite the remarkable regenerative capacity of the liver, it is not always able to undergo efficient repair, and diseases of the liver are common. Our lab uses zebrafish to understand liver development and disease.

The role of uhrf1 in liver development.

    Zebrafish are a powerful system to identify genes that control development. The zebrafish liver is fully formed by 5 days post-fertilization (dpf) and can be imaged using a variety of techniques, including using transgenic fish expressing a fluorescent protein under the control of the hepatocyte-specific fabp10 promoter and in situ hybridization (Figure 1). We hypothesize that a common set of genes regulate liver cell proliferation in the embryo and in the adult during

Figure 1. The liver is well formed in 5 day embryos. A. Transgenic fish expressing GFP in hepatocytes. B. In situ hybridization with the fabp10 hepatocyte-specific probe.

regeneration, and that some of these genes may be co-opted by cells as they undergo tumorigenesis. To test this, we carried out a forward genetic screen to identify genes required for liver growth in embryos and identified uhrf1 as essential for embryonic development and for hepatic outgrowth. Both uhrf1 mutants and larvae with morpholino-mediated uhrf1 knock-down (Figure 2) have small livers. By developing a technique to
remove part of the adult liver and watch it regenerate, we have found

Figure 2. The liver is small in 4 day embryos that lack uhrf1. uhrf1 morphants (bottom 4 larvae) expressing GFP in hepatocytes have small livers and developmental defects compared to controls (top)

that haploinsufficiency of uhrf1 prevents regeneration (Sadler et al., PNAS; 2007). This demonstrates that uhrf1 is required for physiologic liver growth in both embryos and adults.

Is uhrf1 an oncogene?

    Our experiments demonstrate that uhrf1 is required for liver growth under physiologic conditions. Our next question is to determine whether high levels of uhrf1 contribute to unregulated hepatocyte proliferation, as is found in hepatocellular carcinoma. To address this question, we collaborate with the Mount Sinai Liver Cancer Research Program directed by Dr. Josep Llovet. We have found that UHRF1 is highly expressed in patients with advanced hepatocellular carcinoma, and studies are underway to establish the significance of this finding.

How does uhrf1 function to regulate hepatocyte proliferation?

    Recent, exciting data indicates that a primary role of Uhrf1 in mammals is to direct DNA methyl transferase (DNMT1) to hemi-methylated DNA during replication. UHRF1 is required for maintaining DNA methylation and loss of UHRF1 causes significant changes in methylation. Our working model is that zebrafish with depleted uhrf1 fail to undergo methylation-mediated silencing of genes which halt the cell cycle (i.e. tumor suppressors). Currently, work in the lab is focused on identifying potential important uhrf1 target genes.

How is Uhrf1 regulated?

    While the mechanism by which Uhrf1 functions is beginning to be elucidated, there is very little known about how this interesting protein is regulated. The nucleus is thought to be the primary site of Uhrf1 action, and we and others have found that mammalian UHRF is typically localized to the nucleus (Figure 3). We have confirmed this in zebrafish. Additionally, we

Figure 3. UHRF1 is localized to the nucleus in mammalian cells.

have identified point mutations in UHRF1 which cause the protein to be localized to the cytoplasm. In collaboration with the Ukomadu laboratory (Brigham and Women’s Hospital/Harvard Medical School; Boston). We are currently investigating how Uhrf1 nuclear localization affects its function as well as identifying the factors that regulate Uhrf1 localization.

The genetics of fatty liver disease

Fatty liver disease is most frequently associated with obesity or alcohol abuse, and a range from fat accumulating in hepatocytes (steatosis) to the severe steatohepatitis fall under the umbrella of this disease. We are using zebrafish genetics to understand the factors that contribute to steatosis.

Alcoholic liver disease

    Liver disease is among the most severe consequences of chronic alcohol abuse, and can lead to fibrosis, cirrhosis and liver failure. Acute exposure to high concentrations of alcohol (i.e. binge drinking) also affects the liver, typically resulting in steatosis. While steatosis can resolve, it may predispose

Figure 4. Acute exposure to ethanol causes steatosis in 4 day zebrafish larvae.

to more serious liver disease and liver damage following a binge can be exacerbated when combined with other factors, such as obesity or xenobiotic-mediated hepatotoxicity. We have developed a system for identifying the genes that contribute to steatosis in response to acute alcohol exposure in zebrafish larvae (Figure 4). Using this system we found that the SREBP transcription factors are essential for steatosis in response to alcohol (Passeri et al. Hepatology, 2008). Future work is aimed at uncovering other pathways that give rise to steatosis and liver damage in this model.

Metabolic liver disease

    We have discovered that mutation of the novel gene, which we called foie gras (fgr) results in hepatomegaly (large liver; Figure 5, top) and steatosis (Sadler and Hopkins Development, 2005). Additionally, we have found that hepatocytes in fgr mutants are enlarged and cannot carry out basic functions such as glycogen

Figure 5. fgr mutants have hepatomegaly (top) and defective hepatocytes which do not store glycogen (bottom).

storage (Figure 5; bottom) and apoptosis in mutant livers is nearly 4 times what is observed in wild-type larvae.  This and the other hepatic defects in fgr mutants are reminiscent of features found in patients with fatty liver disease. We believe that understanding the mechanism by which fgr mutants develop steatosis will illuminate pathophysiologic mechanisms of fatty liver disease in humans.

The fgr gene is highly conserved in animals, and the Fgr protein does not have any known function. Efforts are underway to determine the cellular function of Fgr and to elucidate the mechanism by which mutation of this novel gene gives rise to steatosis. 

    We find that fgr mutants have defects in protein secretion, activation of the unfolded protein response and hepatic endoplasmic reticulum (ER) stress. Current work is focused on determining how disruption of the secretory pathway and ER stress causes steatosis.

 
lab info



Kirsten Sadler-Edepli
ASSOCIATE PROFESSOR
kirsten.edepli@mssm.edu

212-241-7152 (office)
212-241-0227 (lab)
212-860-9279 (fax)

lab members:
Bambino, Kathryn
Bradley, Patrick
Chu, Jaime
Kent, Brandon
Nayar, Shika
Pistore, Christian
Rafiq, Ruhina
Vacaru, Ana

see photos and more here.

mailto:kirsten.edepli@mssm.edu?subject=sadlerlab.htmlsadlerlab.htmlsadlerlab.htmlsadlerlab.htmlsadlerlab.htmlsadlerlab.htmlsadlerlab.htmlsadlerlab.htmlsadlerlab.htmlshapeimage_15_link_0shapeimage_15_link_1shapeimage_15_link_2shapeimage_15_link_3shapeimage_15_link_4shapeimage_15_link_5shapeimage_15_link_6shapeimage_15_link_7shapeimage_15_link_8shapeimage_15_link_9
key publications

Metabolism & Fatty Liver Disease

Vacaru AM, Di Narzo AF, Howarth DL, Tsedensodnom O, Imrie D, Cinaroglu A, Amin S, Hao K, Sadler KC. (2014)
Molecularly defined unfolded protein response subclasses have distinct correlations with fatty liver disease in zebrafish.  
Dis Model Mech. 7(7):823-35.

Howarth DL, Lindtner C, Vacaru AM, Sachidanandam R, Tsedensodnom O, Vasilkova T, Buettner C, Sadler KC. (2014)
Activating transcription factor 6 is necessary and sufficient for alcoholic fatty liver disease in zebrafish. 
PLoS Genet. 10(5):e1004335.

Tsedensodnom, O., Vacaru, A.M., Howarth, D.L., Yin, C., and Sadler, K.C. (2013)
Ethanol metabolism and oxidative stress are required for unfolded protein response activation and steatosis in alcoholic liver disease.  
Dis Model Mech. 6:1213-1226.

Chu, J., Mir, A., Gao, N., Rosa, S., Monson, C., Sharma, V., Steet, R., Freeze, H.H., Lehrman, M.A. & Sadler, K.C.  (2012)
A zebrafish model of congenital disorders of glycosylation with phosphomannose isomerase deficiency reveals an early opportunity for corrective mannose supplementation.  
Dis Model Mech 6, 95-105. 

Cinaroglu, A., Gao, C., Imrie, D., and Sadler, K.C. (2011)
Activating transcription factor 6 plays protective and pathological roles in steatosis due to endoplasmic reticulum stress in zebrafish. 
Hepatology, 54, 495-508.

Passeri, M.J., Cinaroglu, A., Gao, C. & Sadler, K.C.  (2009)
Hepatic steatosis in response to acute alcohol exposure in zebrafish requires sterol regulatory element binding protein activation.
Hepatology, 49, 443-452.

Liver development, Cancer & UHRF1
Mudbhary, R., Hoshida, Y., Chernyavskaya, Y., Villanova, N., Jacob, V., Fiel, I., Chen, X., Kojima, K., Thung, S., Bronson, R.T., Lachenmayer, A., Alsinet, C., Desai, A., Senbanerjee, S., Ukomadu, C., Llovet, J.M., and Sadler, K.C. (2014)
Overexpression of UHRF1 drives DNA hypomethylation and hepatocellular carcinoma.  
Cancer Cell, 25, 1-14.

Chu, J., Loughlin, E.A., Gaur, N.A., Senbanerjee, S., Jacob, V., Monson, C., Kent, B., Oranu, A., Ding, Y., Ukomadu, C., and Sadler, K.C. (2012)
UHRF1 phosphorylation by cyclin A2/cyclin-dependent kinase 2 is required for zebrafish embryogenesis. 
Mol Biol Cell, 23, 59-70.

Tien, A.L., Senbanerjee, S., Kulkarni, A., Mudbhary, R., Goudreau, B., Ganesan, S., Sadler, K.C. & Ukomadu, C.  (2011)
UHRF1 depletion causes a G2/M arrest, activation of DNA damage response and apoptosis. 
Biochem, J 435, 175-185.
Zhao X, Monson C, Gao C, Gouon-Evans V, Matsumoto N, Sadler, KC*‡ and Friedman SL* (2010)
Klf6/copeb is required for hepatic outgrowth in zebrafish and for hepatocyte specification in mouse ES cells.
Developmental Biology, 344:79-93.
* equal contribution
‡ corresponding author

Sadler, K.C., Krahn, K.N., Gaur, N.A. & Ukomadu, C.  (2007)
Liver growth in the embryo and during liver regeneration in zebrafish requires the cell cycle regulator, uhrf1. 
Proc Natl Acad Sci USA, 104, 1570-1575.


Other Projects
Monson C and Sadler KC. (2010)
Inbreeding depression and outbreeding depression are evident in wildtype zebrafish lines. 
Zebrafish, 7(2):189-97.

see more publications here.sadlerpub.htmlshapeimage_17_link_0
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press release

Doctor Tipster - June 28
 
Key Protein in Alcoholic Fatty Liver Disease
There is a new causative link found between a key cell stress response pathway and alcoholic liver disease. This link was found by researchers from the Icahn School of Medicine at Mount Sinai. The researchers have discovered that by blocking a protein called ATF6 (or activating transcription factor) helped prevent alcohol-induced fatty liver disease while over expression of the protein caused the disease to develop in zebrafish. The results of this study were published in the journal PLoS Genetics.
Icahn School of Medicine at Mount Sinai

for more info click here.http://www.doctortipster.com/21650-possible-etiology-of-alcoholic-fatty-liver-explained.htmlshapeimage_20_link_0