HNF4β, a New Gene of the HNF4 Family with Distinct Activation and Expression Profiles in Oogenesis and Embryogenesis of Xenopus laevis

Abstract

The transcription factor hepatocyte nuclear factor 4 (HNF4) is an orphan member of the nuclear receptor superfamily expressed in mammals in liver, kidney, and the digestive tract. Recently, we isolated the Xenopus homolog of mammalian HNF4 and revealed that it is not only a tissue-specific transcription factor but also a maternal component of the Xenopus egg and distributed within an animal-to-vegetal gradient. We speculate that this gradient cooperates with the vegetally localized embryonic induction factor activin A to activate expression of HNF1␣, a tissue-specific transcription factor with an expression pattern overlapping that of HNF4. We have now identified a second Xenopus HNF4 gene, which is more distantly related to mammalian HNF4 than the previously isolated gene. This new gene was named HNF4␤ to distinguish it from the known HNF4 gene, which is now called HNF4␣. By reverse transcription-PCR, we detected within the 5 untranslated region of HNF4␤ two splice variants (HNF4␤2 and HNF4␤3) with additional exons, which seem to affect RNA stability. HNF4␤ is a functional transcription factor acting sequence specifically on HNF4 binding sites known for HNF4␣, but it seems to have a lower DNA binding activity and is a weaker transactivator than the ␣ isoform. Furthermore, the two factors differ with respect to tissue distribution in adult frogs: whereas HNF4␣ is expressed in liver and kidney, HNF4␤ is expressed in addition in stomach, intestine, lung, ovary, and testis. Both factors are maternal proteins and present at constant levels throughout embryogenesis. However, using reverse transcription-PCR, we found the RNA levels to change substantially: whereas HNF4␣ is expressed early during oogenesis and is absent in the egg, HNF4␤ is first detected in the latest stage of oogenesis, and transcripts are present in the egg and early cleavage stages. Furthermore, zygotic HNF4␣ transcripts appear in early gastrula and accumulate during further embryogenesis, whereas HNF4␤ mRNA transiently appears during gastrulation before it accumulates again at the tail bud stage. All of these distinct characteristics of the newly identified HNF4 protein imply that the ␣ and ␤ isoform have different functions in development and in adult tissues.A central event in early development is the establishment of different cell types from a single cell, the fertilized egg. This cell type is highly specialized, as it contains maternal components that define embryonic induction processes and the initiation of transcriptional cascades. This unique feature of the egg is especially evident in amphibians such as Xenopus laevis. In this species, no gene transcription occurs in early cleavage stages and all early events are regulated by preexisting transcripts and translational products made during oogenesis. Many of these maternal components are transcription factors that can most easily be analyzed in Xenopus (e.g., fos [13], B-myb [3], and Xrel [22]). Recently, we revealed that hepatocyte nuclear factor 4 (HNF4), previously known as a tissue-spe...

CREB-binding Protein Is a Transcriptional Coactivator for Hepatocyte Nuclear Factor-4 and Enhances Apolipoprotein Gene Expression

“…As seen in Fig. 3B, the transcriptional activity of (BA1) 5 CAT was increased approximately 2-fold by overexpression of CBP and HNF-4 as compared with HNF-4 alone, both in HeLa and HepG2 cells. In agreement with the results presented in Fig.…”

“…The reporter plasmids containing the apoCIII(Ϫ890/ϩ24) promoter mutated in the regulatory element B (BM5) and I 4 (I 4 M) have been described previously (14,43). Plasmid (BA1) 5 CAT contains five copies of an HNF-4 binding regulatory element from the apoB gene (element BA1) in front of the minimal apoB promoter (15). Plasmid pG 5 CAT contains five GAL4-binding sites upstream of the ␤-globin promoter and the CAT gene (19).…”

“…Plasmid (BA1) 5 CAT contains five copies of an HNF-4 binding regulatory element from the apoB gene (element BA1) in front of the minimal apoB promoter (15). Plasmid pG 5 CAT contains five GAL4-binding sites upstream of the ␤-globin promoter and the CAT gene (19). The construction of the various HNF-4 mutants cloned in the vector pcDNAI/Amp (Invitrogen), as well as the GAL-HNF-4 chimeras have been previously described (19).…”

“…To discern whether the synergism between HNF-4 and CBP observed is entirely due to HNF-4, two simpler promoter plasmids were used in transfection experiments, one containing five HNF-4-binding sites [(BA1) 5 CAT] and the other with a heterologous promoter [pG 5 CAT] (19). As seen in Fig.…”

“…Originally identified by its importance in the regulation of liver-specific genes, it is also expressed in the pancreas, kidney, stomach, skin, and intestine (1-3). There are currently three family members HNF-4␣, -␤, and -␥ (4,5), and seven splice variants of HNF-4␣ have been identified from human, rat, and mouse cDNAs (6 -11). HNF-4 is a crucial regulator of several metabolic pathways, including those for glucose and lipid homeostasis.…”