Synthesis of human gamma-glutamyl transpeptidase (GGT) during the fetal development of liver

We have determined expression of human GGT gene encoding gamma-glutamyl transpeptidase (GGT) during fetal development of liver using the Northern-blot analysis with a cloned human GGT cDNA and immunohistochemistry with a monoclonal antibody. GGT mRNA could be detected as early as the 12th week of gestation. It then increased gradually to a peak of approx. threefold the amount at week 12, at week 40, just before birth. The size of the mRNA in the fetal liver was 2.7 kb and mRNA of the same size was detected both in the human fetal kidney and human hepatocellular carcinoma as well as normal adult liver. Immunohistochemical analyses show that GGT increased as the fetal liver developed in parallel with the increase in mRNA. Histochemically, GGT was shown to be located in the wall of bile canaliculi when synthesis was low in early development, but to be distributed, in addition, all over the cell membrane of the fetal hepatocytes when synthesis was high at the later stage of development.     

Three-dimensional study of the cytoskeleton in macrophages and multinucleate giant cells by quick-freezing and deep-etching method.

The three-dimensional ultrastructure of multinucleate giant cells in subcutaneous granulomas was compared with those of peritoneal macrophages using a quick-freezing and deep-etching method. Subcutaneous granulomas were induced by implanting plastic coverslips in the dorsal subcutaneous tissue of rats. The quick-freezing and deep-etching replicas were prepared from the cells attached to the coverslips. Dense networks of actin filaments were distributed along all peripheral aspects (beneath the plasma membrane, and on free and coverslip-attached surfaces) of the multinucleate giant cells. On the coverslip-attached surface, numerous clathrin-coated pits and vesicles occurred between the actin filaments. In these cells, intermediate filaments, but not actin filaments, were the predominant cytoskeletal components in perinuclear regions and were attached to the cell nucleus, mitochondria and other vesicular cell organelles. A similar distribution of cytoskeletal components was observed in the mononuclear macrophages of the granulomas and the peritoneal macrophages. These results show that the cytoskeletal organization varies in different regions of the cytoplasm of multinucleate giant cells, while the characteristic cytoskeletal arrangement, resembling that of mononuclear macrophages, is maintained.     

A strong,preferential response of mice to polymorphic antigenic determinants of the chicken MHC,analyzed with mouse hybridoma (monoclonal) antibodies

CBA/J mice were immunized with B²/B² chicken RBC's (theB locus is the major histocompatibility complex (MHC) of the chicken). Spleen cells from these mice gave a much higher plaque-forming cell response when tested with RBC's bearing B² alloantigens, than with RBC's bearing any other MHC alloantigens. Similarly, immunization with chicken RBC's of other genotypes also produced responses that were highest when tested on the genotype used for immunization. Spleen cells from mice immunized 3 days previously with B²/B² RBC's were fused with mouse myeloma cells and hybrid clones which secreted anti-B² antibodies were selected. These monoclonal antibodies could be divided into three groups: (1) those which react with all genotypes of RBC's (panreactive); (2) those which react with only certain genotypes of RBC's and detect “public” MHC antigens; and (3) those which react with only B²/B² RBC's and detect “private” MHC antigens. Monoclonal antibodies which detect a private B² alloantigen were shown to be excellent typing reagents, as all birds of an outbred population which possessed the B² allele were easily detected using a simple one-step direct agglutination assay. No “false positives” were seen. The high and preferential response of the mouse to chicken MHC alloantigens suggested that mice might possess preexisting immunity to these antigens. In agreement with this hypothesis, normal mouse serum was found to have high titres of “natural” antibody against chicken MHC antigens.     

The rpoD gene functions as a multicopy suppressor for mutations in the chaperones, CbpA, DnaJ and DnaK, in Escherichia coli

Abstract The CbpA protein is an analog of the DnaJ molecular chaperone of Escherichia coli . The dnaJ ⁻ cbpA ⁻ double-null mutant exhibits severe defects in cell growth, namely, a very narrow temperature range for growth. To gain insight into the functions of CbpA as well as DnaJ, we isolated a multicopy suppressor gene that permits this dnaJ ⁻ cbpA ⁻~ mutant to grow normally at low temperatures. The suppressor gene was identified as rpoD , the gene that encodes the major σ ⁷⁰. The biological implications of this finding are examined and discussed.