Transferrin is an early marker of hepatic differentiation, and its expression correlates with the postnatal development of oligodendrocytes in mice.

Transferrin (Tf), the iron transport protein, is essential for the growth and differentiation of cells. Therefore, it provides an excellent model to analyze the regulatory mechanisms controlling the expression of a eukaryotic gene in different cell types and during fetal and adult life. In this study, the tissue-specific and developmental regulation of the Tf gene in vivo were analyzed. Human Tf mRNA was detected mainly in fetal and adult liver. A weaker expression was observed in adult and fetal brain and in fetal spleen. By in situ hybridization the presence of mouse Tf mRNA was detected in the hepatic primordia. This is the first observation pointing out Tf as an early marker of hepatic differentiation, prior to the formation of the liver. Thus, TF may be an important tool to follow the hepatic specification of the gut endoderm. Mouse Tf mRNA was also detected in the liver bud and subsequently in the liver throughout fetal life, and in newborn and adult animals. No expression of the Tf gene was observed in the mouse fetal central nervous system (CNS). In contrast, Tf mRNA was detected from the 5th day after birth in the derivatives of the caudal part of the neural tube and subsequently in the derivatives of the rhomboencephalon and that of the prosencephalon. These results indicate that Tf gene expression correlates with the postnatal development of oligodendrocytes in the mouse CNS. To test whether the control elements of the human gene previously found in ex vivo experiments were also active in vivo during fetal and adult life, we fused the -4000/+395' flanking region of the human gene to the coding region of the lacZ gene and generated transgenic mice. The expression of the reporter gene during development was analyzed.

Cell type-specific expression of the human transferrin gene. Role of promoter, negative, and enhancer elements.

Transferrin (Tf), the iron transport protein of vertebrate serum, is mainly synthesized in the liver. cis-Acting DNA elements required for liver-specific expression of the human Tf gene were identified by transient and stable expression assays in human hepatoma (HepG2 and Hep3B) and epithelial carcinoma (HeLa) cell lines. Deletion analysis of the 5' DNA sequences of the gene have defined four functionally different regions: (a) A cell type-specific promoter located between positions -125 and -45 which interacts with two nuclear factors and is sufficient for liver-specific expression. (b) A distal promoter region from -620 to -125 base pairs containing positive and negative cis-acting elements which regulate the promoter activity. (c) A negative-acting region between -1.0 and -0.6 kilobase pairs which down-regulates transcription from the Tf promoter. (d) An enhancer located between -4.0 and -3.3 kilobase pairs which is more active in hepatoma than in HeLa cells. Thus, Tf gene expression is modulated by a combination of multiple positive and negative cis-acting elements. The expression results are discussed with respect to our previous description of the trans-acting factors interacting with the proximal and distal promoter regions.

Complete structure of the human transferrin gene. Comparison with analogous chicken gene and human pseudogene.

The complete structure of the human transferrin gene is presented. This gene has a total size of about 33.5 kb and is organized in 17 exons separated by 16 introns. The chicken ovotransferrin gene has a size of 10.5 kb and is also organized in 17 exons and 16 introns. The analysis of the structure of the two genes confirm, at the gene level, that transferrins originated by a gene duplication phenomenon. Finally, the existence of a new member of the transferrin family, a human transferrin non-processed pseudogene is demonstrated.

Structure of the metJBLF cluster in Escherichia coli K12. Sequence of the metB structural gene and of the 5'- and 3'-flanking regions of the metBL operon.

The total nucleotide sequence (1,158 nucleotides) of the metB gene of Escherichia coli coding for cystathionine gamma-synthase (386 amino acid residues, Mr = 41,503/chain) is presented. The nucleotide sequences of the flanking regions of the metB and metL genes are also presented. Analysis of these sequences and identification of a promoter region upstream from the metB gene confirms that metB and metL form an operon. The transcription direction is from metB to metL; the start site of the gene transcription has been determined. There is no structural evidence of a classical attenuation mechanism in the regulation of this operon coding for enzymes implicated in an amino acid biosynthetic pathway. Finally, the overall organization of the metJBLF gene cluster is discussed.

Construction and physical mapping of plasmids containing the metJBLF gene cluster of E. coli K12.

In vitro recombination techniques were used to clone the E. coli metJBLF gene cluster in a plasmid vector. Several chimeric plasmids were obtained, analyzed by restriction mapping and characterized genetically. The combined results establish that the met gene cluster is contained on an approximately 5.6 kilobase segment of bacterial DNA with metL between metB and metF. The origin of metL was localized precisely by its DNA sequence and its transcription direction was established.

The immune response of CBA mice to OSRA, a peripheral protein from the sheep erythrocyte membrane.

The capacity of a membrane protein from sheep erythrocytes (osmotic shock released antigen: OSRA) to elicit an immune response in CBA mice was investigated. While all OSRA preparations tested are antigenically identical en indistinguishable with respect to physicochemical characteristics, they are not equally efficient in stimulating immunocompetent lymphocytes for antibody production upon a primary immunization (immunogenic versus antigenic OSRA). Both antibody-forming and (T and B) memory cells are generated in mice primed with immunogenic OSRA. Evidence is presented that the failure of antigenic OSRA to induce a primary response to SRBC determinants is related to its inability to stimulate unprimed T-cells. OSRA appears to be a major antigenic determinant of the sheep erythrocyte membrane since it specifically inhibits up to 60% of an anti-SRBC response.

Purification and characterization of a peripheral protein from the sheep erythrocyte membrane.

A peripheral protein of the sheep red blood cell membrane has been purified to homogeneity. This protein is completely released from the membrane by osmotic shock. Its amino acid composition, molecular weight and subunit structure have been determined. Antibodies against this protein have been obtained in rabbits.