Transcription factor GATA-4 is abundantly expressed in childhood but not in adult liver tumors.

OBJECTIVES: Transcription factor GATA-4 is expressed in early fetal liver and essential for organogenesis. It is also implicated in carcinogenesis in several endoderm-derived organs. Hepatoblastoma (HB), the most common malignant pediatric liver tumor, has features of fetal liver including extramedullary hematopoiesis. We investigated the expression of GATA-4 and its purported target gene erythropoietin (Epo) in liver tumors and the role of GATA-4 in HB pathogenesis. PATIENTS AND METHODS: Immunohistochemistry, Western blotting, and reverse transcription-polymerase chain reaction were used for liver samples from patients with HB or hepatocellular carcinoma. To further investigate the role of GATA-4 in pediatric liver tumors, we used adenoviral transfections of wild-type or dominant negative GATA-4 constructs in the human HB cell line, HUH6. RESULTS: We found abundant GATA-4 expression in both types of liver tumors in children, whereas it was absent in adult hepatocellular carcinoma. A close family member GATA-6 was expressed in a minority of childhood but not adult liver tumors. Epo, present in the fetal liver, was also expressed in childhood liver tumors. Moreover, cell line HUH6 was GATA-4 positive and produced Epo. We found that altering the amount of functional GATA-4 in HUH6 cells did not significantly affect either proliferation or apoptosis. CONCLUSIONS: GATA-4 is abundant in pediatric liver tumors, but unraveling its exact role in these neoplasms requires further investigation.

Enhanced expression of transcription factor GATA-4 in inflammatory bowel disease and its possible regulation by TGF-beta1.

BACKGROUND: Transforming growth factor beta 1 (TGF-beta1) promotes epithelial healing in inflammatory bowel disease. We hypothesized that GATA-4, a transcription factor cooperating with TGF-beta signaling pathway, is upregulated by TGF-beta1 in the inflamed intestinal epithelium. METHODS: Normal and inflamed intestinal samples were subjected to immunohistochemistry for GATA-4/6 and the TGF-beta signaling pathway components Smad2/3/4. Proliferation and apoptosis were analyzed using Ki-67 and in situ DNA 3'-end labeling assays and Bax and Bcl-2 immunohistochemistry. Furthermore, GATA-4 was assessed in intestinal Caco-2 cells stimulated with TGF-beta1, or interleukin-6 and tumor necrosis factor alpha. RESULTS: GATA-4 was detected in only 20% of normal intestinal samples, but was upregulated in corresponding inflamed regions. GATA-6 expression remained unchanged during inflammation. TGF-beta1 and Smad3/4, but not Smad2, were expressed concomitantly with GATA-4 in inflamed bowel mucosa. In intestinal Caco-2 cells, TGF-beta1 upregulated GATA-4 and Smad2/3/4, whereas treatment with control cytokines had no effect. Inflammation was associated with increased epithelial cell apoptosis and the enhancement of Bcl-2, but not Bax. CONCLUSIONS: We surmise GATA-4 expression is upregulated in inflamed intestine correlating with the activation of TGF-beta signaling pathway. We speculate that TGF-beta1 drives GATA-4 expression during intestinal inflammation, these two components cooperating to promote epithelial healing.

Transcription factors GATA-4 and GATA-6 in normal and neoplastic human gastrointestinal mucosa.

BACKGROUND: Human gastrointestinal mucosa regenerates vigorously throughout life, but the factors controlling cell fate in mature mucosa are poorly understood. GATA transcription factors direct cell proliferation and differentiation in many organs, and are implicated in tumorigenesis. GATA-4 and GATA-6 are considered crucial for the formation of murine gastrointestinal mucosa, but their role in human gastrointestinal tract remains unexplored. We studied in detail the expression patterns of these two GATA factors and a GATA-6 down-stream target, Indian hedgehog (Ihh), in normal human gastrointestinal mucosa. Since these factors are considered important for proliferation and differentiation, we also explored the possible alterations in their expression in gastrointestinal neoplasias. The expression of the carcinogenesis-related protein Indian hedgehog was also investigated in comparison to GATA factors. METHODS: Samples of normal and neoplastic gastrointestinal tract from children and adults were subjected to RNA in situ hybridization with 33P labelled probes and immunohistochemistry, using an avidin-biotin immunoperoxidase system. The pathological tissues examined included samples of chronic and atrophic gastritis as well as adenomas and adenocarcinomas of the colon and rectum. RESULTS: GATA-4 was abundant in the differentiated epithelial cells of the proximal parts of the gastrointestinal tract but was absent from the distal parts. In contrast, GATA-6 was expressed throughout the gastrointestinal epithelium, and in the distal gut its expression was most intense at the bottom of the crypts, i.e. cells with proliferative capacity. Both factors were also present in Barrett's esophagus and metaplasia of the stomach. GATA-6 expression was reduced in colon carcinoma. Ihh expression overlapped with that of GATA-6 especially in benign gastrointestinal neoplasias. CONCLUSION: The results suggest differential but overlapping functions for GATA-4 and GATA-6 in the normal gastrointestinal mucosa. Furthermore, GATA-4, GATA-6 and Ihh expression is altered in premalignant dysplastic lesions and reduced in overt cancer.

Cooperative interactions among intestinal GATA factors in activating the rat liver fatty acid binding protein gene.

GATA-4, GATA-5, and GATA-6 are endodermal zinc-finger transcription factors that activate numerous enterocytic genes. GATA-4 and GATA-6 but not GATA-5 are present in adult murine small intestinal enterocytes, and we now report the simultaneous presence of all three GATA factors in murine small intestinal enterocytes before weaning age. An immunohistochemical survey detected enterocytic GATA-4 and GATA-6 at birth and 1 wk of age and GATA-5 at 1 wk but not birth. Interactions among GATA factors were explored utilizing a transgene constructed from the proximal promoter of the rat liver fatty acid binding protein gene (Fabp1). GATA-4 and GATA-5 but not GATA-6 activate the Fabp1 transgene through a cognate binding site at -128. A dose-response assay revealed a maximum in transgene activation by both factors, where additional factor did not further increase transgene activity. However, at saturated levels of GATA-4, additional transgene activation was achieved by adding GATA-5 expression construct, and vice versa. Similar cooperativity occurred with GATA-5 and GATA-6. Identical interactions were observed with a target transgene consisting of a single GATA site upstream of a minimal promoter. Furthermore, GATA-4 and GATA-5 or GATA-5 and GATA-6 bound to each other in solution. These results are consistent with tethering of one GATA factor to the Fabp1 promoter through interaction with a second GATA factor to produce increased target gene activation. Cooperative target gene activation was specific to an intestinal cell line and may represent a mechanism by which genes are activated in the small intestinal epithelium during the period before weaning.

GATA-4, GATA-5, and GATA-6 activate the rat liver fatty acid binding protein gene in concert with HNF-1alpha.

Transcriptional regulation by GATA-4, GATA-5, and GATA-6 in intestine and liver was explored using a transgene constructed from the proximal promoter of the rat liver fatty acid binding protein gene (Fabpl). An immunohistochemical survey detected GATA-4 and GATA-6 in enterocytes, GATA-6 in hepatocytes, and GATA-5 in neither cell type in adult animals. In cell transfection assays, GATA-4 or GATA-5 but not GATA-6 activated the Fabpl transgene solely through the most proximal of three GATA binding sites in the Fabpl promoter. However, all three factors activated transgenes constructed from each Fabpl site upstream of a minimal viral promoter. GATA factors interact with hepatic nuclear factor (HNF)-1alpha, and the proximal Fabpl GATA site adjoins an HNF-1 site. GATA-4, GATA-5, or GATA-6 bounded to HNF-1alpha in solution, and all cooperated with HNF-1alpha to activate the Fabpl transgene. Mutagenizing all Fabpl GATA sites abrogated transgene activation by GATA factors, but GATA-4 activated the mutagenized transgene in the presence of HNF-1alpha. These in vitro results suggested GATA/HNF-1alpha interactions function in Fabpl regulation, and in vivo relevance was determined with subsequent experiments. In mice, the Fabpl transgene was active in enterocytes and hepatocytes, a transgene with mutagenized HNF-1 site was silent, and a transgene with mutagenized GATA sites had identical expression as the native transgene. Mice mosaic for biallelic Gata4 inactivation lost intestinal but not hepatic Fabpl expression in Gata4-deficient cells but not wild-type cells. These results demonstrate GATA-4 is critical for intestinal gene expression in vivo and suggest a specific GATA-4/HNF-1alpha physical and functional interaction in Fabpl activation.