Expression of constitutive androstane receptor splice variants in human tissues and their functional consequences.
The constitutive androstane receptor (CAR) NR1I3 is a transcription factor that upon activation by xenobiotics induces transcription of drug-metabolizing and drug transporter genes. Our goal was to identify whether alternative splicing of CAR makes an important contribution to the generation of novel CAR proteins. The wild-type CAR mRNA (CAR.1) and splice variants (SVs) were amplified from human liver cDNAs and in a panel of cDNAs from 36 human tissues, using exon 1- and 3'-untranslated region primers, cloned and sequenced. Twenty-two unique hCAR splice variants (CAR-SVs) containing different combinations of splicing (deletion of exons 2, 4, 5, 7, partial deletion of exon 9, or insertion of 12 or 15 base pairs from introns 6 or 7) were identified. CAR mRNAs were expressed in small intestine, kidney, testis, adrenal, and brain caudate nucleus. Intestine expressed only CAR.1 mRNA, whereas spleen, heart, and prostate expressed only CAR-SVs. In vitro transcription and translation of CAR-SVs lacking exon 2 (missing ATG start site) generated CAR proteins that differed in M(r) from CAR.1. These CAR-SVs used a translation initiation site in exon 1, generating CAR with a unique amino-terminal sequence. Transcripts lacking part of exon 9 altered the CAR reading frame generating CAR proteins with a unique carboxy-terminal end. CAR-SVs demonstrated compromised binding to CYP2B6 NR elements and transcriptional activation of a CYP2B6 luciferase reporter. The expression of CAR in additional human cell types increases the range of tissue specific transcriptional responses regulated by this receptor, suggesting additional biological roles for CAR and CAR-SV proteins in these tissues.
Lamba
Jatinder K
JK
Department of Pharmaceutical Sciences, St. Jude Children's Research Hospital, 332 North Lauderdale Street, Memphis, TN 38105, USA.
Lamba
Vishal
V
Yasuda
Kazuto
K
Lin
Yvonne S
YS
Assem
Mahfoud
M
Thompson
Emma
E
Strom
Stephen
S
Schuetz
Erin
E
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D001483:Base Sequence; D019295:Computational Biology; D000090702:Constitutive Androstane Receptor; D004247:DNA / Q000032:analysis / Q000378:metabolism; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D006801:Humans; D008969:Molecular Sequence Data; D020033:Protein Isoforms; D012333:RNA, Messenger / Q000378:metabolism; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics* / Q000502:physiology; D014157:Transcription Factors / Q000235:genetics* / Q000502:physiology; D015533:Transcriptional Activation
D000090702:Constitutive Androstane Receptor; C000716327:NR1I3 protein, human; D020033:Protein Isoforms; D012333:RNA, Messenger; D018160:Receptors, Cytoplasmic and Nuclear; D014157:Transcription Factors; D004247:DNA