Murine antigen H-2.7: In vitro phenotypic conversion of erythrocytes

The H-2.7 antigen in normal mouse serum can be passively adsorbed to H-2.7^– erythrocytes in 10 percent sucrose (low ionic strength) solution. This antigen can also be stripped off the H-2.7⁺ erythrocytes under the same conditions provided the H-2.7⁺normal serum is absent. The stripped red blood cells can regain the H-2.7 antigen upon reincubation with H-2.7⁺ normal serum. The attachment of the H-2.7 antigen to erythrocytes probably occurs via a specific receptor.Abbreviations used in this paper BSA bovine serum albumin - B10 C57BL/10Sn - HA hemagglutination - LIS low ionic strength solution - NMS normal mouse serum - PBS phosphate-buffered saline - PVP polyvinylpyrrolidone - RBCs red blood cells     

Murine antigen H-2.7: Phenotypic conversion of erythrocytes in radiation chimeras

By use of radiation chimeras produced between H-2.7⁺ and H-2.7^}- strains, A.SW and A.BY and B10.S(7R) and B10.S(9R), we demonstrate that the H-2.7 antigen can be passively attached to or detached from red blood cells. Thus, genetically H-2.7^}- red blood cells derived from H-2.7^}- bone marrow cells, gain H-2.7 antigen while maturing in the H-2.7⁺ host. Similarly, genetically H- 2.7⁺ red blood cells derived from H-2.7⁺ bone marrow cells become H-2.7^}- while maturing in H-2.7^– recipients. This behavior of the H-2.7 antigen is similar to that described for human Chido and Rodgers blood group antigens.Abbreviations used in this paper BMT bone marrow transfer - BSA bovine serum albumin - CT cytotoxicity test - HA hemagglutination - HBSS10 Hank's balanced salt solution containing 10% fetal calf serum - NMS normal mouse serum - PBS phosphate-buffered saline - PVP polyvinylpyrrolidone - RBCs red blood cells     

Murine antigen H-2.7: Localization of its antigenic determinant to the Ss (C4) molecule

Murine blood group antigen H-2.7 is encoded by a locus mapping in the vicinity of theS locus which codes for the Ss antigen carried by the fourth component of the complement pathway (C4). Normal mouse serum of H-2.7-positive strains contains a substance which inhibits anti-H-2.7 hemagglutination. This substance cannot be removed by passage of the serum through an anti-Ss immunoabsorbent column indicating that the Ss and H-2.7 antigens are present on separate molecules or molecular fragments in the serum. In contrast, fresh plasma either does not contain the H-2.7-bearing substance at all or it contains it at a far lower concentration than normal serum, although it has a normal level of the Ss-antigen-bearing substance. However, the H-2.7-positive substance appears when the plasma is allowed to stand for several hours, or when it is dialyzed and treated subsequently in a manner favoring spontaneous degradation of complement components. Removal of the Ss substance from the fresh plasma prevents the appearance of the H-2.7 antigen at any time thereafter. These findings indicate that the Ss and H-2.7 antigens are carried by the same molecule or molecular complex. The intact molecule expresses only the Ss antigen; the H-2.7 antigen is either hidden or masked so that it is inaccessible or poorly accessible to H-2.7 antibodies. Degradation of these molecules results in the generation of two fragments, a large fragment carrying the Ss antigen and a smaller H-2.7-positive fragment. The data are consistent with the interpretation that the H-2.7 antigen is encoded by the S locus, and that it is carried by that portion of the C4 molecule split off during complement activation.Abbreviations used in this paper NMS normal mouse serum - DNP dinitrophenyl - EDTA ethylene-diamine-tetraacetate - PVP polyvinylpyrrolidone     

Murine antigen H-2.7: localization of its antigenic determinant to the Ss (C4) molecule

Murine blood group antigen H-2.7 is encoded by a locus mapping in the vicinity of the S locus which codes for the Ss antigen carried by the fourth component of the complement pathway (C4). Normal mouse serum of H-2.7-positive strains contains a substance which inhibits anti-H-2.7 hemagglutination. This substance cannot be removed by passage of the serum through an anti-Ss immunoabsorbent column indicating that the Ss and H-2.7 antigens are present on separate molecules or molecular fragments in the serum. In contrast, fresh plasma either does not contain the H-2.7-bearing substance at all or it contains it at a far lower concentration than normal serum, although it has a normal level of the Ss-antigen-bearing substance. However, the H-2.7-positive substance appears when the plasma is allowed to stand for several hours, or when it is dialyzed and treated subsequently in a manner favoring spontaneous degradation of complement components. Removal of the Ss substance from the fresh plasma prevents the appearance of the H-2.7 antigen at any time thereafter. These findings indicate that the Ss and H-2.7 antigens are carried by the same molecule or molecular complex. The intact molecule expresses only the Ss antigen; the H-2.7 antigen is either hidden or masked so that it is inaccessible or poorly accessible to H-2.7 antibodies. Degradation of these molecules results in the generation of two fragments, a large fragment carrying the Ss antigen and a smaller H-2.7-positive fragment. The data are consistent with the interpretation that the H-2.7 antigen is encoded by the S locus, and that it is carried by that portion of the C4 molecule split off during complement activation.     

An erythrocyte and serum antigen with a specificity antithetical to the mouse C4 associated H-2.7 antigen

Repeated immunization of intra-H-2 recombinant strain A.TBR16 with lymphoid tissue from strain A.TBR13 produced an antiserum that agglutinated the erythrocytes from inbred strains of mice carrying the b, d, r, q, u, wr7, w13, w17, w19, and w21 haplotypes of the H-2 complex. The antiserum was negative with erythrocytes of strains carrying the haplotypes f, j, k, p, and s. This pattern of reactivity among fifteen H-2 haplotypes is unlike the strain distribution for any known H-2 erythrocyte antigen, and is exactly antithetical to the S region-controlled H-2.7 antigen. An examination of 12 intra-H-2 recombinant haplotypes mapped the genetic determinant controlling the new antigen to the IC or S regions of the H-2 complex. In addition, hemagglutination inhibition studies revealed the antigen was also expressed in serum and plasma. The serologic, genetic, and tissue distribution data suggest the gene controlling the newly defined antigen is allelic to the gene controlling the H-2.7 antigen.     

Identification of specificityH-2.7 as an erythrocyte antigen: Control by an independent locus,H-2G,between theS andD regions

SpecificityH-2.7 is expressed predominantly on erythrocytes and controlled by a gene that maps within theH-2 gene complex at a locus, designated asH-2G, which apparently lies between regionsS andD. Three phenotypes have been observed with respect to this antigen: a) positive by direct test and absorption (haplotypesH-2 ^f ,H-2 ^j ,H-2 ^p ,H-2 ^s ); b) positive only by absorption (H-2 ^k ); and c) negative (H-2 ^b ,H-2 ^d ,H-2 ^q ). New crossover positions have been established for severalH-2 recombinants based on classifications for theH-2G locus.     

Immunological demonstrations of PKC Murine tissue distribution, ontogeny, cellular localization and translocation

An antiserum raised against an PKC-specific peptide recognizes PKC with an apparent molecular weight of 97 kDa in cytosol of mouse brain. No cross-reaction with α,β,γPKC or the δPKC-like p76-kinase is observed. PKC is mainly present in brain. Just traces of this PKC isoenzyme can be detected in some other murine tissues. Ontogenetic studies indicate that the amount of PKC in murine brain increases constantly and reaches a maximal level at day 7 after birth. Upon TPA activation PKC is translocated from the cytosol to the particulate fraction in a brain homogenate.     

Biochemical genetics of rat esterases: Polymorphism,tissue expression,and linkage of four loci

Two alleles at each of four esterase loci in Rattus norvegicus are described with regard to tissue expression, electrophoretic characterization, and genetic linkage. A previously described dominant gene for prealbumin serum esterase is demonstrated to exist as two codominant alleles in the genetically determined absence of the characteristic albumin esterase. The allelic composition of 16 inbred strains for four esterase genes is provided, and the heretofore ambiguous nomenclature of rat esterase genetics is standardized. Linkage of Es-1, Es-2, and Es-3 is demonstrated. Es-2 and Es-3 are tightly linked in that no recombination has been observed in 55 offspring. The same offspring demonstrated 9% recombination between Es-1 and the other two loci.This work was supported by a grant from the Brown-Hazen Fund of Research Corporation.     

Murine glycosyltransferases responsible for the expression of globo-series glycolipids: cDNA structures, mRNA expression, and distribution of their products

Biological functions of globo-series glycosphingolipids are not well understood. In this study, murine cDNAs of two glycosyltransferases responsible for the synthesis of globo-series glycolipids and mRNA expression of those genes were analyzed. Distribution of their products was also analyzed. Murine cDNAs for Gb3/CD77 synthase and Gb4 synthase predicted that both of them are type II membrane proteins with 348 and 331 amino acids, respectively. In northern blotting, Gb3/CD77 synthase gene was mainly expressed in kidney and lung but also detected in many other tissues. Gb4 synthase was expressed in brain, heart, kidney, liver, skin, and testis. In the immunohistological analysis, Gb3/CD77 was mainly expressed in the proximal tubules as revealed with coincidental expression with angiotensin-converting enzyme (ACE). In spleen, it was detected in pre-B cells in the peripheral region of the white pulp, as suggested with coincidental expression with CD10. It was also expressed on the endothelia of the alveolar capillaries in lung and on the sebaceous ducts aside of the hair follicles. Gb4 was also detected mainly on the proximal tubules in kidney and on the endothelia of the alveolar capillaries in lung as Gb3/CD77. But it was also detected on the epithelium of the bronchus, seminiferous tubules and tails of spermatozoa in testis, blood vessels of choroids plexus and endothelial cells in brain, and central and hepatoportal veins in liver. The expression patterns of two genes and their products almost corresponded with some exception. The results would provide essential information for the functional studies of globo-series glycolipids.     

Immunological demonstration of epsilon PKC. Murine tissue distribution, ontogeny, cellular localization and translocation

An antiserum raised against an epsilon PKC-specific peptide recognizes epsilon PKC with an apparent molecular weight of 97 kDa in cytosol of mouse brain. No cross-reaction with alpha, beta, gamma PKC or the delta PKC-like p76-kinase is observed. Epsilon PKC is mainly present in brain. Just traces of this PKC isoenzyme can be detected in some other murine tissues. Ontogenetic studies indicate that the amount of epsilon PKC in murine brain increases constantly and reaches a maximal level at day 7 after birth. Upon TPA activation epsilon PKC is translocated from the cytosol to the particulate fraction in a brain homogenate.     

Seabream antiquitin: molecular cloning, tissue distribution, subcellular localization and functional expression

Subsequent to our earlier report on the first purification of antiquitin protein from seabream liver and demonstration of its enzymatic activity [FEBS Letters 516 (2002) 183-186], we report herein the cloning of its full-length cDNA sequence. The open reading frame encodes a protein of 511 amino acids. Results of RT-PCR indicate that antiquitin is highly expressed in both the seabream liver and kidney. Transfection studies in cultured eukaryotic cells provided further evidence that it is a cytosolic protein. Bacterial expression of the enzyme was also performed. The purified recombinant protein was demonstrated to exhibit similar kinetic properties as the native enzyme.     

Saposins: structure, function, distribution, and molecular genetics.

Saposins A, B, C, and D are small heat-stable glycoproteins derived from a common precursor protein, prosaposin. These mature saposins, as well as prosaposin, activate several lysosomal hydrolases involved in the metabolism of various sphingolipids. All four saposins are structurally similar to one another including placement of six cysteines, a glycosylation site, and conserved prolines in identical positions. In spite of the structural similarities, the specificity and mode of activation of sphingolipid hydrolases differs among individual saposins. Saposins appear to be lysosomal proteins, exerting their action upon lysosomal hydrolases. Prosaposin is a 70 kDa glycoprotein containing four domains, one for each saposin, placed in tandem. Prosaposin is proteolytically processed to saposins A, B, C and D, apparently within lysosomes. However, prosaposin also exists as an integral membrane protein not destined for lysosomal entry and exists uncleaved in many biological fluids such as seminal plasma, human milk, and cerebrospinal fluid, where it appears to have a different function. The physiological significance of saposins is underlined by their accumulation in tissues of lysosomal storage disease patients and the occurrence of sphingolipidosis due to mutations in the prosaposin gene. This review presents an overview of the occurrence, structure and function of these saposin proteins.     

H-2 class I antigen expression on mouse teratocarcinoma cell lines

Immunity against PCC3 teratocarcinoma cells (129, H-2 ^b) was induced in allogeneic (C3H, H-2 ^k) mice by preimmunization with L cells (C3H, H-2 ^k) expressing cosmid-introduced K ^b or D ^b genes, but not with nontransfected L cells. In addition, the growth of PCC3 cells in sublethally irradiated (C3H × B6-H-2 ^bm1)F₁ and (C3H × B6-H-2 ^bm13 )F₁ mice bearing the K ^bm1 and D ^bm13 mutations, respectively, was either prevented, stopped, or delayed in comparison with the (C3H × B6)F₁ (k × b) mice, which failed to reject the PCC3 cells. The teratocarcinoma line OC15S was exceptional because it reacted specifically with K^b- and D^b-specific (but not I^b-specific) alloantisera, and because K^b- and D^b-specific antibodies could be absorbed by OC15S cells. The subpopulation of OC15S cells bearing the ECMA-7 antigen characteristic for embryonic carcinoma (EC) cells was isolated by the fluorescence-activated cell sorter and was shown to react specifically with K^b- and D^b-specific antisera. These experiments show that teratocarcinoma cells express antigens similar or identical to the K-and D-region products of differentiated cells. The lack of expression of class I antigens is thus neither a condition nor a consequence of the pluripotentiality of the EC cells. The exact nature of the major histocompatibility complex antigens on EC cells has yet to be established using the methods of molecular biology and biochemistry.     

A novel guinea pig macrophage-specific polymorphic molecule. I. Biochemistry, genetics, and tissue distribution

In the course of studying Ia molecules from strain 2 and strain 13 guinea pig macrophages, with the intent of comparing them to B cell Ia molecules, it was observed that guinea pig alloserum prepared by cross-immunization of guinea pig lymphocyte Ag non-identical inbred guinea pigs immunoprecipitated not only conventional class I and class II molecules, but also a 98,000-Da molecule, termed gp98. Two different forms of the molecule were detected, indicating it is polymorphic. The genes encoding gp98 were shown not to be linked to the guinea pig lymphocyte Ag complex. The molecule gp98 was found on macrophages within populations of peritoneal exudate cells, resident peritoneal cells, bone marrow cells, and spleen. All gp98-bearing macrophages were also Ia-positive. However, only a subpopulation of macrophages bore gp98. The gp98 was not found on Ly-1 or Ig-bearing cells, indicating that B and T cells do not bear Ia. Thus, gp98 appears to be a highly immunogenic polymorphic macrophage-specific molecule that allows the characterization of guinea pig macrophage subsets.     

Biochemical evidence for expression of a semi-allogeneic, H-2 antigen by a murine adenocarcinoma

LT-85 is an alveolegenic adenocarcinoma induced in mutant C3HfB/HeN (C3Hf) mice. This tumor, however, grows preferentially in allogeneic, wild-type C3H/HeN (C3H) mice. The tumor-associated transplantation antigen has been mapped to the K end of the major histocompatibility complex. H-2K antigens were isolated from detergent extracts of LT-85 cells by immunoprecipitation with monoclonal antibody. The tryptic peptides of these antigens were compared, by using high-pressure liquid chromatography, with the tryptic peptides of H-2K antigens isolated from syngeneic mutant C3Hf and ancestral wild-type C3H spleen cells. We found that the H-2K antigens of the LT-85 tumor cells were very similar to, but distinct from, those present on syngeneic C3Hf lymphoid cells. We also found, however, that the H-2K antigens of LT-85 tumor cells were clearly different from the H-2K antigens of allogeneic C3H spleen cells. The H-2K antigens of LT-85 cells are therefore foreign to syngeneic C3Hf cells, but do not represent expression by the tumor cells of the allogeneic H-2K antigens expressed by normal C3H cells. Furthermore, the nature of the differences observed between the H-2K antigens of LT-85 cells and C3Hf and C3H spleen cells strongly suggests that the structure of the H-2K molecule of LT-85 cells is identical in some regions to the H-2K molecule of C3Hf cells, and in other regions to the H-2K molecule of C3H cells.