The Role of Leucine Carboxyl Methyltransferase-1 (LCMT-1) in Liver Lipid Homeostasis Restricted; Files Only

Abstract

Protein phosphatase 2A (PP2A) family phosphatases (PP2A, PP4 and PP6) are an important family of phosphatases that regulate a variety of functions during cell growth, division, and development. LCMT-1 is a protein methyltransferase that methylates the catalytic C subunit of PP2A, PP4, and PP6. Loss of LCMT-1 leads to a reduction in methylation of PP2A family phosphatases, resulting in disruption of normal development. Previous studies indicate that LCMT-1 plays a key role in fetal liver development and its reduction is also associated with insulin resistance, a symptom linked to non-alcoholic fatty liver disease (NAFLD). NAFLD affects around 25% of people in the world and is especially common among middle-aged Americans. The long-term goal of our research is to determine whether modulation of PP2A family phosphatase methylation might be therapeutically beneficial. Preliminary data from experiments done in the Pallas lab using homozygous albumin-cre-directed LCMT-1 conditional knockout (cKO) mice suggest that LCMT-1 may play a vital part in liver lipid homeostasis. This research project studies a cKO mouse model in which LCMT-1 is only knocked out in liver cells in order to further understand the role of LCMT-1 in liver lipid accumulation and liver cell viability. Two feeding studies were performed with similar designs, the only difference being that the first used homozygous Alb-cre mice while the second used heterozygous Alb-cre mice. In each feeding study, cKO and corresponding sex-and-age-matched control mice were placed on a high-fat diet and a normal diet. After a 17-week feeding period, their livers and serum were harvested, analyzed, and compared to see whether the loss of LCMT-1 affects liver lipid accumulation and cell viability, both hallmarks of NAFLD. Alanine transferase (ALT) levels in serum served as an indicator of liver cell death, while western blot analysis provided insights into LCMT-1’s mechanistic effects. Our data show that loss of LCMT-1 induced by both homozygous and heterozygous Alb-cre expression results in enlargement of livers and spleens and elevated alanine transferase (ALT) activity in blood in cKO mice placed on high-fat diet. In homozygous Alb-cre cKO mice, the loss of LCMT-1 also leads to faster weight gain. Overall, our data suggest that LCMT-1 plays a vital role in liver lipid homeostasis. 

Table of Contents

I. Abstract..................................................................................................4

II. Acknowledgements...................................................................................6

III. Introduction..........................................................................................9

IV. Methods.............................................................................................12

a. Albumin-cre-directed LCMT-1 cKO mouse model ....................................12

b. Feeding period and diets...................................................................13

c. Subsequent analysis........................................................................13

d. Colony maintenance........................................................................14

e. Data analysis.................................................................................14

V. Results.................................................................................................15

a. Conditional loss of LCMT-1 in liver results in greater liver to the body weight ratio with either normal diet or high-fat diet......................................................15

b. Conditional loss of LCMT-1 in liver results in greater systemic inflammation when on a high-fat diet ......................................................................17

c. Conditional loss of LCMT-1 in liver results in elevated ALT activity when on a high-fat diet......................................................................................18

d. Conditional loss of LCMT-1 in liver results in faster weight gain in homozygous Alb-cre mice on high-fat diet................................................................19

e. Conditional loss of LCMT-1 in livers results in reduced ribosomal S6 phosphorylation in homozygous Alb-cre mice when on a high-fat diet..................21

f. High-fat diet results in elevated relative p57 NFkB phosphorylation in homozygous Alb-cre cKO mice and control mice..........................................................22

VI. Conclusion & Future Work........................................................................23

VII. References...........................................................................................30

About this Honors Thesis

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School	Emory College
Department	Biology
Degree	B.S.
Submission	Honors Thesis
Language	English
Research Field	Biology, Molecular
Parola chiave	PP2A liver lipid homeostasis LCMT-1 PP4
Committee Chair / Thesis Advisor	Dr. David C. Pallas, Emory University
Committee Members	Dr. Alexander Escobar, Emory University Dr. Patricia A. Marsteller, Emory University

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