Differential Expression and Function of Alternative Splicing
Variants of Human Liver X Receptor
Kaori Endo-Umeda, Shigeyuki Uno, Ko Fujimori, Yoshikazu Naito, Koichi Saito,
Kenji Yamagishi,
1
Yangsik Jeong, Hiroyuki Miyachi, Hiroaki Tokiwa, Sachiko Yamada,
and Makoto Makishima
Division of Biochemistry, Department of Biomedical Sciences, Nihon University School of Medicine, Tokyo, Japan (K.E.-U.,
S.U., S.Y., M.M.); Laboratory of Biodefense and Regulation, Osaka University of Pharmaceutical Sciences, Osaka, Japan (K.F.);
Environmental Health Science Laboratory, Sumitomo Chemical Co., Ltd., Osaka, Japan (Y.N., K.S.); Research Information
Center for Extremophile (K.Y.) and Department of Chemistry (H.T.), Faculty of Science, Rikkyo University, Tokyo, Japan;
Department of Biochemistry, Institute of Lifestyle Medicine, and Nuclear Receptor Research Consortium, Yonsei University
Wonju College of Medicine, Gangwon-do, Republic of Korea (Y.J.); and Graduate School of Medicine, Dentistry, and
Pharmaceutical Sciences, Okayama University, Okayama, Japan (H.M.)
Received December 14, 2011; accepted March 7, 2012
ABSTRACT
The liver X receptor (LXR) is a nuclear receptor that is involved
in regulation of lipid metabolism, cellular proliferation and apopto-
sis, and immunity. In this report, we characterize three human
LXR isoforms with variation in the ligand-binding domain (LBD).
While examining the expression of LXR3, which lacks 60 amino
acids within the LBD, we identified two novel transcripts that
encode LXR-LBD variants (LXR4 and LXR5). LXR4 has an
insertion of 64 amino acids in helix 4/5, and LXR5 lacks the
C-terminal helices 7 to 12 due to a termination codon in an
additional exon that encodes an intron in the LXR1 mRNA.
LXR3, LXR4, and LXR5 were expressed at lower levels com-
pared with LXR1 in many human tissues and cell lines. We also
observed weak expression of LXR3 and LXR4 in several tissues
of mice. LXR ligand treatment induced differential regulation of
LXR isoform mRNA expression in a cell type-dependent manner.
Whereas LXR3 had no effect, LXR4 has weak transactivation,
retinoid X receptor (RXR) heterodimerization, and coactivator re-
cruitment activities. LXR5 interacted with a corepressor in a
ligand-independent manner and inhibited LXR1 transactivation
and target gene expression when overexpressed. Combination of
LXR5 cotransfection and LXR antagonist treatment produced
additive effects on the inhibition of ligand-dependent LXR1 ac-
tivation. We constructed structural models of the LXR4-LBD and
its complexes with ligand, RXR-LBD, and coactivator peptide. The
models showed that the insertion in the LBD can be predicted to
disrupt RXR heterodimerization. Regulation of LXR pre-mRNA
splicing may be involved in the pathogenesis of LXR-related
diseases.
Introduction
Liver X receptor (LXR; NR1H3) and LXR (NR1H2) are
transcription factors of the nuclear receptor superfamily (Ton-
tonoz and Mangelsdorf, 2003; Makishima, 2005). Whereas
LXR is ubiquitously expressed, LXR is localized to the liver,
adipose tissue, small intestine, and macrophages. Both recep-
tors are activated by oxysterols and have been linked to path-
ways involved in fatty acid and cholesterol homeostasis. LXRs
bind preferentially to LXR-responsive elements (LXREs) that
consist of a two-hexanucleotide (AGGTCA or a related se-
quence) direct repeat motif separated by four nucleotides (direct
repeat 4) as a heterodimer with retinoid X receptor (RXR;
NR2B). LXRs regulate intestinal absorption and biliary excre-
tion of cholesterol by inducing the expression of target genes
This work was supported in part by the Ministry of Education, Culture,
Sports, Science, and Technology of Japan [Grant-in-Aid for Scientific Research
on Priority Areas 18077005] (to M.M.).
1
Current affiliation: Department of Chemical Biology and Applied Chem-
istry, College of Engineering, Nihon University, Fukushima, Japan.
Article, publication date, and citation information can be found at
http://molpharm.aspetjournals.org.
http://dx.doi.org/10.1124/mol.111.077206.
ABBREVIATIONS: LXR, liver X receptor; LXRE, LXR-responsive element; RXR, retinoid X receptor; ABC, ATP-binding cassette; CYP7A,
cholesterol 7-hydroxylase; SREBP, sterol regulatory element-binding protein; LBD, ligand-binding domain; T0901317, N-(2,2,2-trifluoro-ethyl)-
N-[4 –(2,2,2-trifluoro-1-hydroxy-1-trifluoromethyl-ethyl)-phenyl]-benzenesulfonamide; GW3965, 3-[3-[N-(2-chloro-3-trifluoromethylbenzyl)-(2,2-di-
phenylethyl)amino]propyloxy]phenylacetic acid hydrochloride; HEK, human embryonic kidney; FBS, fetal bovine serum; PCR, polymerase chain
reaction; siRNA, small interfering RNA; CMV, cytomegalovirus; EMSA, electrophoretic mobility shift assay; DBD, DNA-binding domain; AF2,
activation function 2; SMRT, silencing mediator of retinoic acid and thyroid hormone receptor; N-CoR, nuclear receptor corepressor; SRC, steroid
receptor coactivator; DRIP205, vitamin D receptor-interacting protein 205; PPAR, peroxisome proliferator-activated receptor.
1521-0111/12/8106-800–810$25.00
MOLECULAR PHARMACOLOGY Vol. 81, No. 6
Copyright © 2012 The American Society for Pharmacology and Experimental Therapeutics 77206/3770094
Mol Pharmacol 81:800–810, 2012
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