Aspartate beta-hydroxylase and DNA damage in chronic liver diseases - Project summary
Chronic alcohol drinking has been linked to the development and progression of alcoholic liver disease (ALD) by
means of damaging cellular DNA, which results in cell death of hepatocytes, contributing to alcoholic
steatohepatitis, liver fibrosis, and potentially liver cancers. However, it remains unclear how alcohol promotes
DNA damage responses in ALD development and understanding the underlying mechanisms may yield insight
into therapeutic approaches toward treating ALD. In the pilot studies, it was demonstrated that aspartate beta-
hydroxylase (ASPH) is down-regulated in the rodent livers derived from the ethanol-fed group. In addition, DNA
damage markers, including γ-H2aX (a phosphorylated form of H2aX) and GADD45α, were found elevated when
compared to the control-diet fed group. The liver is predominantly composed of hepatocytes whose function is
critical in the maintenance of homeostasis thereof. Thus, we investigated whether there is a correlation between
ASPH and DNA damage in human hepatocytes. We ectopically expressed ASPH and challenged hepatocytes
with DNA damage inducers. Intriguingly, the expression of ASPH almost eliminated γ-H2aX expression; however,
knockdown of ASPH substantially promoted DNA damage. These observations suggest a protective role of
ASPH in the DNA damage response. Based on these findings, we hypothesize that chronic alcohol
consumption leads to ASPH down-regulation in the liver contributing to DNA damage responses and
hepatocyte cell death in chronic liver disease progression. Our long-term goal is to understand how ASPH
is involved in ethanol-induced hepatocyte DNA damage and the consequent impacts on ALD progression. To
test our hypothesis and achieve this goal, two specific aims are proposed. In aim 1, we will investigate how
the expression of ASPH alters ALD progression in vivo. We will determine if targeting hepatic ASPH using
an AAV strategy will exacerbate DNA damage and cell death in the hepatocytes of ethanol-fed mice. Furthermore,
we investigate if overexpression of ASPH protects hepatocytes in an ALD model. In aim 2, we will decipher
the mechanism by which ASPH protects hepatocytes from DNA damage induced cell death. We will
determine how ASPH is involved in ethanol-mediated DNA damage by using CRISPR/Cas9 to knock out ASPH
or ectopically expressing ASPH via a lentiviral system. To delineate the mechanisms involved in this regulation,
we will determine how the enzymatic functions of ASPH are associated with the ASPH-mediated DNA damage
responses and cell death. Using the proteomic approach, we will also illustrate innovative ASPH downstream
targets upon ethanol treatment. The results will significantly advance our understanding of the molecular
mechanisms in hepatocyte cell death and pathophysiological mechanisms underlying ALD progression. We also
anticipate that it will broaden our knowledge of ASPH expression and its relationship to hepatocyte function in
general.
