Abstract
Hepatocyte Nuclear Factor 4a (HNF4a), a master regulator of liver-specific gene expression, is regulated by
two promoters (P1 and P2) which drive expression of two groups of HNF4a isoforms referred to as HNF4a1
and HNF4a7. HNF4a is a known regulator of gluconeogenesis and mutated in maturity onset diabetes of the
young one (MODY1). Conventionally, it was thought that HNF4a1, but not HNF4a7, is expressed in the normal
adult liver, while HNF4a1 is downregulated and HNF4a7 is upregulated in liver cancer. Now, research in our lab
reveals a previously undescribed role for HNF4a7 in the normal adult mouse liver – one involved in the diurnal
variations of lipid and carbohydrate metabolism. More specifically, HNF4a1 appears to be a major driver of
gluconeogenesis while HNF4a7 is a driver of ketogenesis: we propose that alterations in the levels of the HNF4a
isoforms during the day flip the molecular switch between the two. Our preliminary data also show that HNF4a7
is required for increased levels of circulating ketone bodies in female mice. AMP-Activated Protein Kinase
(AMPK), an energy-sensing enzyme, has been shown to phosphorylate HNF4a1 in vitro, but effects in vivo and
on HNF4a7 are not known. SIRT1 is a deacetylase that works with AMPK to regulate glucose and lipid
metabolism. HNF4a1 is known to be acetylated and our preliminary data suggest that HNF4a7 but not HNF4a1
interacts with SIRT1. Here, we propose to use HNF4a1-expressing (a1HMZ) and HNF4a7-expressing exon
swap mice (a7HMZ) to determine the physiological function of the HNF4a isoforms in the switch between
gluconeogenesis and ketogenesis, and to characterize the impact of sex on those functions. In Aim 1, we will
determine whether intermittent fasting and a ketogenic diet increase the levels of HNF4a7 in the liver, and
whether the increase occurs in all hepatocytes, or just a subset. We will determine the consequences of HNF4a7
on gene expression. Kidney and intestines will also be explored. In Aim 2, we will determine whether the AMPK
pathway acts in a differential fashion on the HNF4a isoforms to help flip the metabolic switch. Phosphorylation
by AMPK and deacetylation by SIRT1 will be explored. Finally, in Aim 3, we will determine whether the estrogen
pathway impacts the HNF4a isoforms in female mice and determine the consequences for the metabolic switch.
Our compelling preliminary data that the HNF4a isoforms are involved in the switch between
gluconeogenesis and ketogenesis shed new light on this basic metabolic process that occurs on a daily basis
and under conditions of feeding and fasting. The results from this proposal will illuminate not only the molecular
mechanism underlying the switch but also how that mechanism is impacted by sex. The proposed studies have
the potential to impact our understanding of numerous metabolic diseases, including diabetes, obesity, fatty liver
disease and cancer. Finally, given the fact that ketone bodies serve as a source of fuel for the brain, our results
could have a broader impact, including on neurological diseases, such as dementia.