Oncogenic Met receptor induces ectopic structures in Xenopus embryos.

When aberrantly expressed or activated, the Met receptor tyrosine kinase is involved in tumor invasiveness and metastasis. In this study, we have used the Xenopus embryonic system to define the role of various Met proximal-binding partners and downstream signaling pathways in regulating an induced morphogenetic event. We show that expression of an oncogenic derivative of the Met receptor (Tpr-Met) induces ectopic morphogenetic structures during Xenopus embryogenesis. Using variant forms of Tpr-Met that are engineered to recruit a specific signaling molecule of choice, we demonstrate that the sole recruitment of either the Grb2 or the Shc adaptor protein is sufficient to induce ectopic structures and anterior reduction, while the recruitment of PI-3Kinase (PI-3K) is necessary but not sufficient for this effect. In contrast, the recruitment of PLCgamma can initiate the induction, but fails to maintain or elongate supernumerary structures. Finally, evidence indicates that the Ras/Raf/MAPK pathway is necessary, but not sufficient to induce these structures. This study also emphasizes the importance of examining signaling molecules in the regulatory context that is provided by receptor/effector interactions when assessing a role in cell growth and differentiation.

The human hepatitis B virus transactivator X gene product regulates Sp1 mediated transcription of an insulin-like growth factor II promoter 4.

Human hepatitis B virus (HBV) is one of the causative agents of hepatocellular carcinoma (HCC). The virus encodes a 17 kDa protein, X, which is known to be a causative agent in the formation of HCC. An insulin-like growth factor-II (IGF-II) is expressed during the formation of HCC. Among the four promoters of the IGF-II gene, promoters 2, 3 and 4 become activated during the formation of HCC. The high frequency of detection of hepatitis B virus X (HBV-X) antigen in liver cells from patients with chronic hepatitis, cirrhosis, and liver cancer suggested that the expressions of HBV-X and IGF-II are associated. Studies were carried out to test the relationship between the HBV-X gene product and the activation of IGF-II promoter 4. We demonstrated that the HBV-X protein increases the endogenous IGF-II expression from promoter 3 and 4 of IGF-II gene. Analysis of the fourth promoter of IGF-II gene showed that the HBV-X gene product positively regulates transcription. Two copies of a motif are responsible for conferring HBV-X regulation on the fourth promoter of IGF-II. These motifs have been identified as Sp1 binding sites. Sp1 binding to IGF-II P4 promoter was identified by gel mobility shift assay using purified Sp1. By using a GAL4-Sp1 fusion protein it was demonstrated that HBV-X positively regulates the Spl mediated transcriptional activity of IGF-II in vivo. A protein-affinity chromatography experiment showed that HBV-X protein does not bind directly to Sp1, but HBV-X does augment the DNA binding activity of the phosphorylated form of Sp1 in HepG2 cells. Sp1 was phosphorylated by HBV-X and its DNA-binding activity was up-regulated upon HBV-X transfections. Various HBV-X mutant expression vectors were used for the demonstration of specific interactions between Sp1 and HBV-X. These results indicate that HBV-X functions as a positive regulator of transcription, and that Sp1 is a direct target for the transcriptional regulation of IGF-II. Increasing the DNA binding ability of the phosphorylated form of Sp1 by HBV-X might be an important mechanism for regulating the IGF-II gene expression and possibly promoting cell division during hepatic carcinogenesis. Our experimental results suggest that expression of HBV-X might induce the expression of IGF-II and the IGF-II might play a role in hepatitis B virus pathogenesis during the formation of HCC.

Cloning and characterization of cDNAs coding for heavy and light chains of agglutinating monoclonal antibody (HAG12islrh) specific for human red blood cells.

Complementary DNAs encoding the heavy and light chains of the Fab fragment of mouse agglutinating monoclonal antibody against human red blood cells were cloned by polymerase chain reaction and their nucleotide sequences were determined. The sequence analysis showed that the variable regions of the heavy and light chains were the members of mouse heavy-chain subgroup IIa and kappa light-chain subgroup I, respectively. A few unusual amino acids in the constant regions of the heavy chain were also recognized.