Elsevier

Biochemical Pharmacology

Volume 72, Issue 4, 14 August 2006, Pages 512-522
Biochemical Pharmacology

Alterations in transporter expression in liver, kidney, and duodenum after targeted disruption of the transcription factor HNF1α

https://doi.org/10.1016/j.bcp.2006.03.016Get rights and content

Abstract

The transcription factor hepatocyte nuclear factor 1α (HNF1α) is involved in regulation of glucose metabolism and transport, and in the expression of several drug and bile acid metabolizing enzymes. Targeted disruption of the HNF1α gene results in decreased Cyp1a2, and Cyp2e1 expression, and increased Cyp4a1 and Cyp7a1 expression, suggesting these enzymes are HNF1α target genes. Since hepatic metabolism can be coordinately linked with drug and metabolite transport, this study aims to demonstrate whether HNF1α regulates expression of a variety of organic anion and cation transporters through utilization of an HNF1α-null mouse model. Expression of 32 transporters, including members of the Oat, Oatp, Oct, Mrp, Mdr, bile acid and sterolin families, was quantified in three different tissues: liver, kidney, and duodenum. The expression of 17 of 32 transporters was altered in liver, 21 of 32 in kidney, and 6 of 32 in duodenum of HNF1α-null mice. This includes many novel observations, including marked downregulation of Oats in kidney, as well as upregulation of many Mrp and Mdr family members in all three tissues. These data indicate that disruption of HNF1α causes a marked attenuation of several Oat and Oatp uptake transporters in liver and kidney, and increased expression of efflux transporters such as Mdrs and Mrps, thus suggesting that HNF1α is a central mediator in regulating hepatic, renal, and intestinal transporters.

Introduction

Transporters serve a multitude of functions, including import and export of critical endogenous substrates, as well as aiding in absorption, distribution, and elimination of many xenobiotics. Transporters characteristically move water-soluble compounds and hydrophilic metabolites into and out of cells because these molecules do not pass readily through membranes. Due to a wide range of compounds that need passage through membranes, a host of transporters have evolved to manage the cellular environment. In this regard, several transporter families exist to aid in disposition of chemicals, including the organic anion transporters (Oat), organic anion transporting polypeptide transporters (Oatp), organic cation transporters (Oct), multidrug resistance protein transporters (Mdr), multidrug resistance-associated protein transporters (Mrp), the bile acid and sterolin transporter families, along with Bcrp (breast cancer resistance protein; Abcg2). Whereas this is only a fraction of the total transporters that exist, these transporter families have been implicated as being important in the uptake and elimination of xenobiotics in liver, kidney, and intestine—all organs which play a vital role in drug disposition.

HNF1α is a transcription factor that is known to regulate the basal expression of many genes, and has been referred to as a “master” transcription factor due to its control over other known nuclear receptors and transcription factors [1]. HNF1α is expressed in liver, kidney, intestine, stomach, and pancreas [2], [3], and has been implicated in the regulation of bile acid, fatty acid, and drug metabolism in vivo [4], [5], [6]. Disruption of HNF1α results in decreased expression of several cytochrome P450s including Cyp1a2, Cyp2c29, Cyp2d9, Cyp2e1, and Cyp7b1, and increased expression of Cyp2b10, Cyp3a11, Cyp4a1, Cyp7a1, Cyp8b1, and Cyp39a1 [6]. Similarly, HNF1α is believed to be critical for regulation of insulin-like growth factor 1 (IGF-1), and the resulting loss of IGF-1 expression in the absence of HNF1α gives a phenotype resembling Laron dwarfism, and non-insulin-dependent diabetes mellitus (NIDDM) [7] suggesting the possibility of compensation by other transcription factors or altered interactions with co-activator and co-repressor proteins that are central to normal expression of HNF1α target genes [8], [9].

The development of HNF1α-null mice allows for an in vivo model for examining the physiological role of HNF1α. Mutations in the HNF1α gene exist in humans and lead to maturity-onset diabetes of the young type 3 (MODY-3), and the resulting physiological consequences seem to be modeled well by the null mouse [7].

HNF1α has also been implicated in the downregulation of only a few transporters, including human organic-anion-uptake transporters OATP-C and OATP8, the human sodium-glucose transporter (SGLT1), as well as mouse Oatp4 and the sodium-phosphate transporters Npt1, 2, and 4 in mouse [10], [11], [12]. However, this probably represents only a handful of transporters that might be regulated by HNF1α, whether direct or indirect. Decreased binding of HNF1α to 5′ flanking regions of transporters such as Ntcp and Oatps may be a critical event in bile-acid homeostasis, as this may lead to impaired hepatic uptake during cholestasis [13], [14]. However, very little work has focused on whether HNF1α regulates other solute carrier transporter families (i.e. Oats, Octs, Oatps), as well as its effects on the efflux transporters (i.e. Mdrs, Mrps). Similarly, the tissue distribution of HNF1α suggests that this transcription factor may be critical in other organs besides liver, especially in kidney and in intestine. To identify HNF1α target genes in these three tissues the branched DNA assay (Genospectra; Fremont, CA) was utilized to examine expression of a large battery of transporters in both wildtype (WT) and HNF1α-null mice. This study aims to determine whether HNF1α is critical for endo- and xenobiotic transporter expression in a variety of tissues.

Section snippets

Mice

HNF1α-null mice were engineered using Cre-loxp recombination as described previously [7]. Because HNF1α-null mice are infertile, null mice were derived by cross-breeding heterozygote male and female mice. HNF1α(+/+) littermates were used as wildtype (WT) controls. Tissues from eight male HNF1α WT and eight HNF1α-null adult mice under 4 months of age were collected and snap frozen in liquid nitrogen. Tissues were stored at −80 °C until further use.

RNA isolation

Total RNA was isolated using RNAzol Bee reagent

Hnf1α disruption alters expression of the organic anion transporter (Oat) family in liver, kidney, and duodenum

Oats are solute uptake transporters known to play an important role in kidney. Oat2, which has the highest expression of Oats in liver, had markedly lower hepatic mRNA expression in Hnf1α-null mice as compared to WT mice (Fig. 1), whereas Oat1 and Oat3 remained unchanged (Table 2). Oat1, 2 and 3 expression in kidney was lower in null than WT mice; expression of Oat1 and 2 in HNF1α-null kidney was <20% than in WT mice. In duodenum, Oat3 expression was five-fold higher in Hnf1α-null mice than in

Discussion

The results of the present study demonstrate that Hnf1α controls a multitude of transporters involved in the disposition of a wide variety of substrates. These data support the concept that Hnf1α plays a role not only in the regulation of drug and bile acid metabolism, but also in the uptake and export of substrates for these enzymes.

Hnf1α serves as a central mediator for many other transcription factors, and while much of this regulation is not understood, speculation of possible methods of

Acknowledgements

The authors would like to thank Katy Allen and Drs. Matt Dieter, Hong Lu, Chuan Chen, and Susan Buist for input and discussion, as well as the Laboratory Animal Resource Center at KUMC for their excellent contributions in helping with the mice.

References (42)

  • F. Chen et al.

    Liver receptor homologue-1 mediates species- and cell line-specific bile acid-dependent negative feedback regulation of the apical sodium-dependent bile acid transporter

    J Biol Chem

    (2003)
  • C.J. Soroka et al.

    Cellular localization and up-regulation of multidrug resistance-associated protein 3 in hepatocytes and cholangiocytes during obstructive cholestasis in rat liver

    Hepatology

    (2001)
  • K. Ogura et al.

    Full-length cDNA cloning and genomic organization of the mouse liver-specific organic anion transporter-1 (lst-1)

    Biochem Biophys Res Commun

    (2000)
  • P. Borst et al.

    The multidrug resistance protein family

    Biochim Biophys Acta

    (1999)
  • S. Baumhueter et al.

    HNF-1 shares three sequence motifs with the POU domain proteins and is identical to LF-B1 and APF

    Genes Dev

    (1990)
  • Y.H. Lee et al.

    Laron dwarfism and non-insulin-dependent diabetes mellitus in the Hnf-1alpha knockout mouse

    Mol Cell Biol

    (1998)
  • J. Eeckhoute et al.

    Hepatocyte nuclear factor 4 alpha isoforms originated from the P1 promoter are expressed in human pancreatic beta-cells and exhibit stronger transcriptional potentials than P2 promoter-driven isoforms

    Endocrinology

    (2003)
  • M.G. Martin et al.

    Regulation of the human Na(+)-glucose cotransporter gene, SGLT1, by HNF-1 and Sp1

    Am J Physiol Gastrointest Liver Physiol

    (2000)
  • M. Trauner et al.

    Endotoxin downregulates rat hepatic Ntcp gene expression via decreased activity of critical transcription factors

    J Clin Invest

    (1998)
  • X. Cheng et al.

    Tissue distribution and ontogeny of mouse organic anion transporting polypeptides (Oatps)

    Drug Metab Dispos

    (2005)
  • D.Q. Shih et al.

    Loss of HNF-1alpha function in mice leads to abnormal expression of genes involved in pancreatic islet development and metabolism

    Diabetes

    (2001)
  • Cited by (94)

    • Drug Transport—Uptake

      2022, Comprehensive Pharmacology
    • Upregulation of histone acetylation reverses organic anion transporter 2 repression and enhances 5-fluorouracil sensitivity in hepatocellular carcinoma

      2021, Biochemical Pharmacology
      Citation Excerpt :

      The transcription of OAT2 in liver is inhibited by prototypical chemicals (e.g., dioxin, rifampicin and phenobarbital) through activating multiple nuclear receptors such as farnesoid X receptor (FXR), pregnane X receptor (PXR), aryl hydrocarbon receptor (AhR), constitutive androstane receptor (CAR) and retinoid X receptor (RXR) [28]. Also, targeted disrupting hepatocyte nuclear factors 1α (HNF-1α) [29] or repressing hepatocyte nuclear factors 4α (HNF-4α) by bile acids [30] can decrease OAT2 expression. However, this is the first study explaining how histone acetylation regulates OAT2 to our knowledge.

    • Regulation of organic anion transporters: Role in physiology, pathophysiology, and drug elimination

      2021, Pharmacology and Therapeutics
      Citation Excerpt :

      Several transcription factors have been identified to be involved in the regulations of OATs. For example, in hepatocyte nuclear factor 1α (HNF1α)-null mice, the levels of renal Oat1, Oat3, and Urat1 mRNA were markedly reduced as compared to those in wild-type mice, and HNF1α overexpression enhanced OAT1, OAT3, and URAT1 promoter activity in vitro (Kikuchi et al., 2006; Kikuchi et al., 2007; Maher et al., 2006; Saji et al., 2008). In ex vivo experiments with kidney organ culture, HNF4α antagonist attenuated the expression of Oat1 and Oat3 mRNA (Martovetsky, Tee, & Nigam, 2013).

    • Diurnal expression of ABC and SLC transporters in jejunum is modulated by adrenalectomy

      2019, Comparative Biochemistry and Physiology Part - C: Toxicology and Pharmacology
      Citation Excerpt :

      So, it is plausible that Hnf4α mediates the effect of glucocorticoids on some multi-specific SLC transporters. In contrast, the arrhythmic expression of Hnf1α encoding the transcriptional regulator of intestinal Sglt1 (Rhoads et al., 1998; Vayro et al., 2001) and several Oat and Oatp transporters (Maher et al., 2006), excludes the possibility that this transcription factor might be a mediator of circadian- and glucocorticoid-regulated transcription of Sglt1 and Oatp1. Our study illustrates that the regulation of genes encoding SLC and ABC transporters can be interpreted in the framework of several regulatory pathways – circadian control (Mohawk et al., 2012; Richards and Gumz, 2012), neuroendocrine control of biogenesis and secretion of corticosteroids (Kalsbeek et al., 2012) and remote sensing and signaling model (Nigam et al., 2015; Wu et al., 2011).

    View all citing articles on Scopus

    National Institute of Health grants ES-09716 and ES-07079.

    View full text