Eberhard Günther

Comparative analysis of the three major histocompatibility complex-linked heat shock protein 70 (Hsp70) genes of the rat

The organization of the three major histocompatibility complex (Mhc)-linked heat shock protein 70 (Hsp70) genesHsp70-1, Hsp70-2, andHsp70-3, and the nucleotide sequences of these genes, are presented for the rat.Hsp70-1 andHsp70-2 gene products are identical at the amino acid level. From the pattern of sequence similarity of the orthologous Mhc-linkedHsp70 genes of rat, human, and mouse, it is concluded that the gene duplications leading to the three-gene cluster occurred before the separation of the primate and rodent lines and that theHsp70-1 andHsp70-2 genes of rat and human might have undergone homogenization of their sequences.

The distribution of Tap2 alleles among laboratory rat RT1 haplotypes.

We are reporting the cDNA sequences of Tap2 from two cima and two cimb rat strains. Comparison of the cDNA sequences shows that these alleles fall into two groups, which we refer to as Tap2-A and Tap2-B. We found that alleles from the Tap2-B group are more closely related to the mouse homologue than are Tap2-A alleles, and among the 48 nucleotides which differ between the Tap2-A and Tap2-B cDNAs, three affect restriction sites. We defined pairs of oligonucleotides which allow amplification of the regions bearing these restriction sites from genomic DNA or cDNA, and this technique has been successful for the genotyping of all of the 56 laboratory strains of Rattus norvegicus tested and for five cell lines tested so far. All 14 known RT1 standard haplotypes were tested, and 7 found to belong to the Tap2-B group, and 7 to Tap2-A. We also found that intron sizes among the alleles of the Tap2-B group fall into two subgroups, providing further insight into the phylogency of these various haplotypes.

Biochemical markers in inbred strains of the rat (Rattus norvegicus).

Klaus Bender 1, M a r k Adams 2, Peter R. Baverstock 2, Maria den Bieman 3, Siegbert Bissbort 1, Rad im Brdi~ka 4, Geoffrey W. Butcher 5, Dona ld V. Cramer 6, Otto yon Deimling 7, Michael F .W. Festing 8, Eberhard Gtinther 9, Rona ld D. G u t t m a n n 1°, Hans J. Hedrich 11, Philip B. Kendall 12, Reinhard Kluge i t , Ren6 Moutier 13, Babette Simon 7, James E. W o m a c k ~4, Junzo Yamada ~5, and Bert van Zutphen 3

Heat Shock Protein 70 Is Able to Prevent Heat Shock-Induced Resistance of Target Cells to CTL

Heat shock or transfection with heat shock protein 70 (Hsp70) genes has been shown to protect tumor cell lines against immune mechanisms of cytotoxicity. We have reported previously that heat shock confers resistance to CTL in the rat myeloma cell line Y3 that is Hsp70 defective. Evidence is now presented that Hsp70 is able to prevent the induction of the resistant phenotype. In Con A-stimulated lymphocytes and in lymphocyte × Y3 somatic cell hybrid clones a severe, non-Hsp70-inducing heat shock elicits resistance to CTL in contrast to a heat shock that results in Hsp70 expression. Thus, Hsp70 expression appears to be negatively associated with the development of resistance. Furthermore, loading of Y3 cells with recombinant Hsp70 protein before heat shock is able to prevent resistance. Because apoptosis induced in Y3 cells by heat shock is not affected, Hsp70 appears to interfere selectively with the CTL-induced lethal pathway that is found to be calcium but not caspase dependent. It is suggested that after heat shock Hsp70 enhances the CTL-induced apoptotic pathway by chaperoning certain proteins in the target cell that are involved in the execution of cell death. Thus, although shown to confer protection against many cytotoxic mechanisms, Hsp70 does not appear to be generally cytoprotective. This observation could also be of relevance when interpreting the effectiveness of tumor immunity.

Factor XII (Hageman) deficiency in women with habitual abortion: new subpopulation of recurrent aborters?

Objective To examine the possible association between factor XII (FXII) deficiency and an elevated number of abortions. Design Factor XII activity, FXII antigen concentration, other blood clotting parameters, and phospholipid antibodies were examined in venous blood from 43 women with repeated (3 to 7) abortions before the 28th week of gestation but without gynecological and chromosomal abnormalities. The data were compared with those obtained from 49 age-matched women without fetal loss. Results Eight cases with moderately reduced FXII activity (35% to 68% of normal) could be identified in the abortion group, whereas among controls no abnormalities in FXII activity and antigen concentration were found. The relative occurrence of reduced FXII level was higher among patients with more than three abortions as compared with those with three abortions. Conclusion Repeated abortions may be associated with reduced level of FXII activity of unknown origin.

Genetic control of susceptibility to mercury-induced immune nephritis in various strains of rat

Susceptibility to mercury-induced immune complex-type glomerulonephritis has been studied in various inbred rat strains. It has been shown that susceptibility is under genetic control and that genes linked to the major histocompatibility complex play a major role. Genes nonlinked to the major histocompatibility complex may be also involved in Wistar—Furth rats. This study also demonstrates that HgCl2 injections can induce different immunomorphological types of glomerulonephritides, depending on the rat strain tested. Circulating anti-single-stranded DNA (SS DNA) antibodies were detected in rat strains which developed an immune-type nephritis and data in PVGc rats suggest that these antibodies might be of pathogenic significance. Both the genetic background and the genes linked to the major histocompatibility complex are probably involved in the genetic control of the anti-SS DNA antibody response in rats injected with mercuric chloride. This study provides models to study the pathogenesis and the genetic control of susceptibility to the induction of autoimmune disorders induced by toxic agents.

Cim: an MHC class II-linked allelism affecting the antigenicity of a classical class I molecule for T lymphocytes

Two alleles at the major histocompatibility complex (MHC)-linked locus cim determine “gain and loss” changes in the rat RT1.Aa class I molecule which affect its structure both as an alloantigen and as a restriction element. Alleles at the cim locus also influence the post-translational modification of RT1.Aa. These effects may reflect the participation of the cim gene product in the processes of peptide loading or assembly of RT1.Aa. In this study we have used the discriminating RT1.Aa-specific monoclonal antibody JY3/84, as well as cytotoxic T cells raised in appropriate combinations, to determine the cim alleles of eight haplotypes in 15 independent inbred strains of rat. We have also employed the same techniques to analyse a panel of F1 hybrid animals derived from various MHC recombinant strains. These experiments map the cim locus to the class II region of RT1, probably between the DP-related genes (RT1.H) and the DQ-related RT1.Bα.

Evidence for a gonad-specific receptor for H-Y antigen: Binding of exogenous H-Y antigen to gonadal cells is independent of β2-microglobulin

This report addresses the question whether two different types of binding exist for the reaction of H-Y antigen with the cell surface. Anti-H-Y antiserum in the presence of complement was cytotoxic only for gonadal cells expressing their own H-Y antigen, but not to ovarian cells loaded with H-Y antigen. H-Y antigen was co-redistributed with beta 2--microglobulin on newborn testicular cells, but some residual H-Y activity was found on similarly treated testis cells from 15 day old rats. After beta 2--microglobulin redistribution, testis cells maintained their binding capacity for exogenous H-Y antigen prepared from epididymal fluid or Daudi cell culture supernatants. This result suggests that exogenous H-Y antigen is bound via a gonad-specific receptor which is independent of beta 2--microglobulin and that this type of binding for H-Y antigen is different from the beta 2--m-associated expression of H-Y antigen on the cell surface.

Cytotoxic T lymphocytes of the rat are predominantly restricted by RTL1.A and Not RT1.C-determined major histocompatibility class I antigens

Two types of biochemically defined class I major histocompatibility complex (MHC) antigens are found in the rat, RTLA antigens that are ubiquitously expressed and RTLC antigens which so far are detectable only on certain cell types, notably B and T lymphocytes. It is shown that the cytotoxic T lymphocyte response to minor H antigens of the LEW strain, including the H-Y antigen, and to TNP-modified syngeneic lymphoid cells is restricted by RTLA but not RTLC gene products. This conclusion is based on bulk culture assays including cold target inhibition tests and limiting dilution experiments using recombinants between the RT1a and RT1u haplotypes. The possibility that class I MHC antigens exist which have no major restriction function is discussed.

Physical Map and Expression Profile of Genes of the Telomeric Class I Gene Region of the Rat MHC

The rat is an important model for studying organ graft rejection and susceptibility to certain complex diseases. The MHC, the RT1 complex, plays a decisive role in controlling these traits. We have cloned the telomeric class I region of the RT1 complex, RT1-C/E/M, of the BN inbred rat strain in a contig of overlapping P1-derived artificial chromosome clones encompassing ∼2 Mb, and present a physical map of this MHC region. Forty-five class I exon 4-hybridizing BamHI fragments were detected, including the previously known rat class I genes RT1-E, RT-BM1, RT1-N, RT1-M2, RT1-M3, and RT1-M4. Twenty-six non-class I genes known to map to the corresponding part of the human and mouse MHC were tested and could be fine mapped in the RT1-C/E/M region at orthologous position. Four previously known microsatellite markers were fine mapped in the RT1-C/E/M region and found to occur in multiple copies. In addition, a new, single-copy polymorphic microsatellite has been defined. The expression profiles of several class I genes and the 26 non-class I genes were determined in 13 different tissues and exhibited restricted patterns in most cases. The data provide further molecular information on the MHC for analyzing disease susceptibility and underline the usefulness of the rat model.