Immunohistochemical detection of hepatocyte nuclear factor‐4α in vertebrates. Issue 12 (1st July 2021)

• Record Type: Journal Article
• Authors: Jiang, Shuying; Tanaka, Toshiya; Yagami, Ren; Hasegawa, Go; Umezu, Hajime; Fujiyoshi, Yukio; Kodama, Tatsuhiko; Naito, Makoto; Ajioka, Yoichi
• Is Part Of: Microscopy research and technique. Volume 84:Issue 12(2021)
• Page Start: 2906
• Page End: 2914
• Publication Date: 2021-07-01
• Subjects: hepatocyte nuclear factor 4α -- immunohistochemistry -- invertebrate -- vertebrate -- Western blot; Electron microscopy -- Technique -- Periodicals; Microscopy -- Periodicals; Microscopy -- Technique -- Periodicals; 502.825
• DOI: 10.1002/jemt.23848
• Languages: English
• ISSNs: 1059-910X

Abstract

Abstract: Hepatocyte nuclear factor‐4α (HNF4α) presents in multiple isoforms generated using alternative promoter (P1 and P2) and splicing. Neither conservation of tissue distribution of HNF4α isoforms, nor presence of alternative promoter usage is known. In this study, to detect the expression of HNF4α in some species of animals, we have applied monoclonal antibodies against P1 (K9218) and P2 (H6939) promoter‐driven and P1/P2 promoter‐driven H1415 HNF4α for immunohistochemistry and western blot analysis. Antibody K9218 was observed in the hepatocytes, proximal tubules of the kidney, and epithelial cells in the mucosa of the small intestine and colon of rats, chicken, and tortoise, whereas antibody H6939 signal were detected in the stomach, pancreas, bile duct, and pancreatic duct of human and rats. The signal for antibody K9218 was recognized in tissues of a wide range of mammals, bird, reptile, amphibian, and fish as well. Antibody H1415 displayed a positive reaction in hepatocytes and intestinal epithelial cells in chicken and tortoise, whereas the bile duct, mucosal epithelial cells in the stomach, or pancreas in these animals were negative. Western blotting showed the binding of the antibody with HNF4α protein from each animal. The sequence of human HNF4α was 100% identical to murine and rat HNF4α, 88.9% to chicken, 77.8% to Xenopus HNF4α, and 81.5% to medaka. However, the specific part of human and invertebrate Drosophila HNF4 shares only 14.8% sequence identity. This antibody is useful for detecting HNF4α isoforms in a wide range of vertebrates, and suggests many insights into animal evolution. Abstract : The N‐ and C‐terminal amino acid sequences in the DNA‐binding domains of the HNF4α protein are not well preserved among animal species. Functional identity and tissue‐specific expression of HNF4α isoforms in animal species is unknown. In most vertebrate animals except lamprey eel, the monoclonal antibody HNF4α K9218 was observed in the hepatocytes, proximal tubules of the kidney, and epithelial cells in the mucosa of the small intestine. Bile duct epithelial cells, gastric mucosal cells, acinar cells and Langerhans cells, and ductal epithelial cells of the pancreas, were negative. The positive cells and negative cells are indicated by red arrows and black arrows, respectively. The tissues of invertebrate animals did not show immunostaining for any of the antibodies.