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Biomedical subjects

B Koller

Publications and source records attributed to B Koller.

At least 37 records · Page 2Linked to original sources
PubMed1992

Major histocompatibility complex class I-restricted T cells are required for resistance to Mycobacterium tuberculosis infection.

Mice with a targeted disruption in the beta 2-microglobulin (beta 2m) gene, which lack major histocompatibility complex class I molecules and consequently fail to develop functional CD8 T cells, provided a useful model for assessing the role of class I-restricted T cells in resistance to infection with virulent Mycobacterium tuberculosis. Of mutant beta 2m-/-mice infected with virulent 10(6) M. tuberculosis, 70% were dead or moribund after 6 weeks, while all control mice expressing the beta 2m gene remained alive for > 20 weeks. Granuloma formation occurred in mutant and control mice, but far greater numbers of tubercle bacilli were present in the lungs of mutant mice than in controls, and caseating necrosis was seen only in beta 2m-/-lungs. In contrast, no differences were seen in the course of infection of mutant and control mice with an avirulent vaccine strain, bacille Calmette-Guérin (BCG). Immunization with BCG vaccine prolonged survival of beta 2m-/-mice after challenge with M. tuberculosis for 4 weeks but did not protect them from death. These data indicate that functional CD8 T cells, and possibly T cells bearing gamma delta antigen receptor, are a necessary component of a protective immune response to M. tuberculosis in mice.

Animals↗
PubMed1992

Major histocompatibility complex class I-specific and -restricted killing of beta 2-microglobulin-deficient cells by CD8+ cytotoxic T lymphocytes.

Cytotoxic T lymphocytes (CTLs) recognize major histocompatibility complex (MHC) class I molecules, normally composed of a heavy chain, a beta 2-microglobulin (beta 2m), and peptide antigens. beta 2m is considered essential for the assembly and intracellular transport of MHC class I molecules as well as their peptide presentation to CTLs. Contrary to this dogma, we now report the generation of allospecific and restricted CD8+ and TCR alpha beta+ CTLs (where TCR is T-cell receptor) capable of killing beta 2m-deficient cells. Such CTLs were obtained by priming mice with live allogeneic beta 2m- spleen cells or mutant lymphoma cells producing MHC class I protein but no detectable beta 2m. Although both beta 2m- and beta 2m-expressing lymphoma cells were rejected in allogeneic mice, only the former were efficient inducers of CTLs recognizing beta 2m- cells. These CTLs were MHC class I (H-2Kb or Db)-specific and CD8-dependent and did not require serum as a source of external beta 2m in the culture. They could be induced across major and minor histocompatibility barriers. The H-2-restricted CTLs generated in the latter case failed to kill the antigen-processing-deficient target RMA-S cells. The results show that MHC class I heavy chains in beta 2m- cells can be transported to the cell surface and act as antigens or antigen-presenting molecules to allospecific and MHC-restricted CTLs.

Animals↗
PubMed1991

Recognition of beta 2-microglobulin-negative (beta 2m-) T-cell blasts by natural killer cells from normal but not from beta 2m- mice: nonresponsiveness controlled by beta 2m- bone marrow in chimeric mice.

The role of major histocompatibility complex (MHC) class I expression in control of the sensitivity of normal cells to natural killer (NK) cells was studied by the use of mutant mice made deficient for expression of beta 2-microglobulin (beta 2m) through homologous recombination in embryonal stem cells. T-cell blasts from beta 2m-deficient (beta 2m -/-) mice were killed by NK cells from normal mice in vitro, while beta 2m +/- blasts were resistant. The beta 2m defect also affected the NK effector cell repertoire: NK cells from beta 2m -/- mice failed to kill beta 2m -/- blasts, while they retained the ability to kill the prototype NK cell target lymphoma YAC-1, although at reduced levels. The inability to recognize beta 2m -/- blasts could be transferred with beta 2m -/- bone marrow to irradiated beta 2m-expressing mice. In contrast, the development of CD8+ T cells (deficient in beta 2m -/- mice) was restored in such chimera. These results indicate that loss of MHC class I/beta 2m expression is sufficient to render normal cells sensitive to NK cells, and that the same defect in the hemopoietic system of a mouse renders its NK cells tolerant to beta 2m-deficient but otherwise normal cells. In the beta 2m -/- mice, NK cells may be selected or educated by other bone marrow cells to tolerate the MHC class I deficiency. Alternatively, the specificity may be controlled directly by the class I molecules on the NK cells themselves.

Animals↗
PubMed1990

[Can sudden cardiac death be prevented by treatment with anti-arrhythmia drugs?].

Sudden cardiac death is defined as death due to a primary cardiac cause or mechanism, occurring within one hour of the onset of acute illness in a person thought to be free of, or with symptomatically mild, heart disease, or simply prehospital death. Of persons dying suddenly, 90% have coronary artery disease, less commonly, dilated cardiomyopathy or hypertrophic cardiomyopathy, preexcitation syndrome, long QT-syndrome, conduction disturbances, congenital or valvular heart disease as well as cardiac tamponade are responsible. In the USA, the incidence of sudden cardiac death is approximately 450,000 per year, in the Federal Republic of Germany the number lies at about 70,000 to 80,000. The most important risk factors for sudden cardiac death are impaired left ventricular ejection fraction, myocardial ischemia and arrhythmias. In general, sudden cardiac death is caused by ventricular fibrillation which arises mainly by degeneration of ventricular tachycardia (VT). The terminal arrhythmia, it is assumed, is precipitated by premature ventricular beats originating in an arrhythmogenic substrate. MEDICAL ANTIARRHYTHMIC TREATMENT IN PATIENTS WITH CORONARY ARTERY DISEASE AFTER MYOCARDIAL INFARCTION: STUDIES WITH CLASS I DRUGS: The results of nine large, randomized , controlled studies are available in which the mortality of patients on antiarrhythmic treatment has been studied (Table 1). Two studies each were carried out with aprindine, phenytoin, mexiletine and tocainide as well as one study with endainide, flecainide or morizicine. With the exception of the CAST study, no study showed a significant difference between treated patients and the control group with respect to mortality or incidence of sudden cardiac death. The CAST study was terminated after ten months because the administration of flecainide and encainide led to overall mortality of 7.7% vs. 3.0% in the control group and the rate of sudden cardiac death at 4.5% was significantly higher in the treatment group than the 1.2% incidence found in controls (Table 2). For nearly all of the studies described, the patient groups were not sufficiently large and subgrouping according to patient characteristics was not carried out such that possibly, inhomogeneity of the entire collective may not have been recognized precluding identification of some individuals who may have shown benefit from antiarrhythmic treatment. The necessity for treatment in many of those receiving drugs is questionable since generally the rhythm profile of the patients was not taken into consideration for the decision to treat. Proarrhythmic effects, accordingly, were also not assessed. Individual treatment and dosage adjustment by monitoring with effectiveness criteria was carried out in one study only in which, even here, criteria for effectiveness were arbitrarily capable of eliciting antiarrhythmic actions. Calculation of mortality rates was carried out on the basis of the total number of deaths in the respective groups without taking into consideration that by the end of the study, in the treatment group the medication had been discontinued in up to 40% of the patients. STUDIES WITH CLASS II DRUGS: For treatment with beta-receptor blockers there are 15 large, controlled, randomized, long-term studies available in which total mortality and the incidence of sudden cardiac death were studied.(ABSTRACT TRUNCATED AT 400 WORDS)

Anti-Arrhythmia Agents↗
PubMed1989

Entamoeba histolytica ribosomal RNA genes are carried on palindromic circular DNA molecules.

Highly abundant DNA fragments obtained after restriction enzyme digests of nuclear DNA of Entamoeba histolytica strain HM-1:IMSS have been cloned and characterized. Northern blot hybridization to E. histolytica rRNA and sequence analysis identified the abundant DNAs as ribosomal DNA containing species. Several overlapping clones containing these abundant DNAs were isolated from 4 different genomic libraries of E. histolytica. Alignment of the restriction maps was consistent with a circular molecule, about 24.6 kilobase pairs (kb) in size. Nuclease BA131 digestion provided additional evidence for the circular nature of this DNA. The ribosomal DNA molecule contains two large inverted repeat-regions, each at least 5.2 kb in length. Sequence analysis of clone R715 revealed homology to the large rRNA units of various eukaryotic organisms. This clone was located in both inverted repeats, suggesting two rRNA cistrons per molecule. The inverted repeats are flanked by stretches of DNA which contain tandemly reiterated sequences. Southern blot analysis of E. histolytica nuclear DNA revealed the presence of two populations of molecules. These molecules have identical arrangements of restriction sites, but differ in size (0.7 kb) in a fragment containing tandemly reiterated sequences. Analysis of E. histolytica nuclear DNA by electron microscopy also revealed circular molecules. These molecules are about 26.6 kb +/- 0.5 kb in size and contain structural features predicted by the restriction map of the extrachromosomal ribosomal DNA of E. histolytica.

Animals↗
PubMed1989

Biphenotypic leukemic lymphocyte precursors in CD2+CD19+ acute lymphoblastic leukemia and their putative normal counterparts in human fetal hematopoietic tissues.

During detailed immunophenotypic analyses of marrow blasts from 336 acute lymphoblastic leukemia (ALL) patients, a very small percentage of cases reactive with B-cell-directed as well as T-cell-directed monoclonal antibodies (MoAbs) were identified. Five ALL cases were biphenotypic since they coexpressed CD2 (Tp50) and CD19 (Bp95) antigens at the single-cell level. The composite immunophenotype of these biphenotypic ALL cases was [TdT+HLA-ABC+CD2+CD3-CD10+CD13-CD14-CD16-CD19+CD20+ ++-CD21-CD33-CD34+Bgp95-C mu- slg-]. Low-molecular-weight B-cell growth factor (LMW-BCGF), recombinant interleukin-2 (rIL-2), and rIL-3 stimulated the proliferative activity of biphenotypic leukemic lymphocyte precursors without inducing differentiation. In the presence of the phorbol ester TPA, leukemic blasts from two cases differentiated along the B precursor pathway to the [CD2-CD10+CD19+CD20+C mu+slg-] pre-B cell stage. Biphenotypic ALL cases did not share a common configuration and gene rearrangement pattern of the immunoglobulin heavy chain genes or T-cell receptor (TCR) genes. Three cases had rearranged C mu genes but germline TCR genes, one case showed rearrangement of both C mu and TCR genes, and the remaining case had rearranged TCR genes but germline C mu genes. All five patients attained prompt remission after standard induction chemotherapy. Three to four years after initial diagnosis, four patients are now off chemotherapy and remain alive in their first remission. One patient relapsed at 3 years, 7 months, but promptly achieved complete remission after reinduction chemotherapy and remains in second remission off chemotherapy greater than 3 years after her reinduction therapy. With two-color immunofluorescence staining techniques and multiparameter flow cytometric analyses, we identified a small population of CD2+CD19+ lymphoid cells in fetal livers (FLs) and fetal bone marrows (FBMs), which may represent the putative normal counterparts of biphenotypic ALL blasts. A CD2+CD19+ normal biphenotypic lymphoid precursor cell line, designated FL 8.2 CD2+, was established from an FL of 8-weeks of gestational age by Epstein-Barr virus (EBV)-induced blastoid transformation. The composite immunophenotype of FL 8.2 CD2+ cell line was [TdT+HLA-ABC+HLA-DR+ CD2+CD5-CD7-CD10+/-CD13-CD19+CD20-CD21+ CD22+CD33-CD34+/-Bgp95-CDw40+C mu-slgD-slgM-]. FL 8.2 CD2+ cells showed germline patterns of immunoglobulin heavy-chain joining region, heavy-chain constant region, kappa light-chain constant region genes, and TCR beta-chain genes. Cross-linking of CD2 as well as CD19 antigens on FL 8.2 CD2+ cells caused an increase of intracellular ionized calcium.(ABSTRACT TRUNCATED AT 400 WORDS)

Antibodies, Monoclonal↗
PubMed1987

A gene cluster in the spinach and pea chloroplast genomes encoding one CF1 and three CF0 subunits of the H+-ATP synthase complex and the ribosomal protein S2.

The regions of the spinach and pea chloroplast genomes containing the ATP synthase genes atpA, atpF and atpH have been sequenced. The encoded proteins, CF1 alpha, CF0I and CF0III, are well conserved between spinach and pea, and analogous to the alpha, b and c subunits of the Escherichia coli ATP synthase complex. The atpF gene is split by a single intron, and the exon/intron boundaries have been defined by isolating and sequencing a partial cDNA clone. Two other genes, designated atpI and rps2, located upstream from atpH, have also been sequenced. They encode a 27,000 Mr hydrophobic protein analogous to the F0a subunit of E. coli ATP synthase and a basic protein analogous to the S2 protein of the E. coli 30 S ribosomal subunit. Transcriptional analysis by electron microscopy of RNA-DNA hybrids, Northern blotting and primer extension experiments shows that these genes are transcribed and processed into a complex set of transcripts, with 5' ends mapping upstream from the rps2, atpI and atpH genes.

Amino Acid Sequence↗
PubMed1987

Evidence for in vivo trans splicing of pre-mRNAs in tobacco chloroplasts.

The rps12 gene codes for chloroplast ribosomal protein S12. In the tobacco chloroplast genome (156 kbp circular DNA), exons II and III of this gene are separated by an intron of 536 bases and are present in two copies in the inverted repeat region, while exon I is located in the large single copy region at a distance of 90 kb and 126 kb from the two copies of exons II and III. These three exons were artificially combined in cloned DNA fragments and hybridized with tobacco chloroplast RNA. Electron microscopic analysis of RNA-DNA hybrids showed that exon I is transcribed as part of a polycistronic RNA containing upstream and downstream sequences; the same is true for exons II and III. Exon I is shown to be transcribed separately from exons II and III. In the most abundant class of the hybridized RNA molecules, exon I was covalently linked to exon II. In these molecules the sequences downstream of exon I and upstream of exon II are not present. These data indicate that maturation of rps12 pre-mRNAs in chloroplasts of tobacco involves trans splicing.

Chloroplasts↗
PubMed1986

The enigma of the gene coding for ribosomal protein S12 in the chloroplasts of Nicotiana.

A 2.9 kbp region from within the inverted repeat of Nicotiana chloroplast DNA hybridized with a chloroplast DNA fragment from Euglena containing the complete rps12 gene coding for ribosomal protein S12. Nucleotide sequencing within this region revealed the existance of two rps12 coding stretches interrupted by 540 bp having class II intron structure. Joining and decoding the exon regions produced a sequence of 85 amino acids colinear and 81% homologous to the S12 protein of Euglena chloroplasts and E. coli, starting from amino acid residue 38 to the stop codon. Immediately upstream of codon 38, conserved intron sequences were located. However, the 5' 37 codon of Nicotiana chloroplast rps12 could not be identified by electron microscopy of RNA-DNA hybrids within a DNA region extending 4000 bp upstream of codon 38, nor by computer search of a completely sequenced region extending for more than 9000 bp upstream of this codon. In E. coli, alteration in rps12 codons 42 or 87 causes streptomycin resistance. However, the nucleotide sequence of the identified rps12 exons in two Nicotiana chloroplast mutants resistant to streptomycin were found to be identical to that of wild type.

Amino Acid Sequence↗
PubMed1986

Transfer of cloned human class I major histocompatibility complex genes into HLA mutant human lymphoblastoid cells.

Three new kinds of recombinant DNA constructs were used to transfer cloned human class I HLA genes (A2 and B8) into unique HLA mutant lymphoblastoid cells: pHeBo(x): a class I gene, "x," in plasmid vector pHeBo, which contains a hygromycin resistance gene and Epstein-Barr virus oriP element that sustains extrachromosomal replication; pHPT(x): gene x in a vector with a hypoxanthine-guanine phosphoribosyltransferase (HPRT) gene; pHPTe(x): gene x in a vector with the HPRT gene and oriP element. Cell surface class I antigen expression was strong in transferents made with class I-deficient lymphoblastoid cell line mutants .144 (A-null), .53 (B-null), and .184 (A-null, B-null). Transferents expressing HLA-A2 were recognized specifically by HLA-A2-specific cytotoxic T lymphocytes. When introduced on either of the vectors with the Epstein-Barr virus oriP element, the class I gene replicated extrachromosomally and was lost at rates of 0.2 to 0.3 per cell division. When introduced with vector pHPT (lacking Epstein-Barr virus oriP), the B8 gene was inserted at different chromosomal locations. Introduction of the HLA-B8 gene failed to restore antigen expression by HLA-B-null mutant .174, providing evidence that, unlike mutants exemplified by .53, .144, and .184, some HLA antigen loss mutants are deficient in a trans-acting function needed for class I antigen expression. Of more general interest, the results obtained with HLA class I genes in vectors that replicate extrachromosomally suggest ways of relating genic expression to chromatin structure and function and of attempting to clone functional human centromeres.

Cell Line↗
PubMed1985

Separation of complementary strands of plasmid DNA using the biotin-avidin system and its application to heteroduplex formation and RNA/DNA hybridizations in electron microscopy.

A method for the separation of complementary strands with the help of the biotin-avidin system is described. Restriction fragments were terminally labeled at both ends with biotinylated nucleotides. The DNA was cut by a second restriction enzyme, and the fragments were bound to an avidin agarose column. The non-biotinylated strands were eluted with 0.1 M NaOH, and the biotin-labeled strands were subsequently released from the column by elution with 50% guanidine isothiocyanate/formamide. Contamination of the separated strands by complementary single strands was less than 4%.-Separated linear single strands of the vector pEMBL were prepared. On annealing with recombinant circular DNA a substitution loop is formed which provides position and orientation markers for the unambiguous electron microscopic analysis of heteroduplexes or hybrids formed with the inserted sequences. -The terminal biotin label was visualized by complex formation with a streptavidin-ferritin conjugate.

Avidin↗
PubMed1985

The wheat chloroplast gene for CF(0) subunit I of ATP synthase contains a large intron.

The gene for CF(0) subunit I of ATP synthase has been located in wheat chloroplast DNA, between the genes for CF(0) subunit III and alpha subunit of CF(1). Nucleotide sequencing and analysis of RNA-DNA hybrids indicated that the gene is interrupted by an 823-bp intron which has boundaries similar to those previously described for the introns in protein-coding chloroplast genes of Euglena gracilis. The deduced amino acid sequence of CF(0) subunit I indicates a polypeptide of 183 amino acid residues. However, N-terminal amino acid sequencing of the mature spinach CF(0) subunit I suggests that the protein is synthesised with a N-terminal extension of 17 amino acid residues and is processed to give a protein of mol. wt. 19 001 of 166 amino acids residues. The mature CF(0) subunit I shows similarities in primary and predicted secondary structure to F(0) subunit b of Escherichia coli ATP synthase. A major transcript of 3.3 kb containing sequences from the genes for CF(0) subunit III, subunit I and CF(1) subunit alpha has been observed by RNA-DNA hybridisation.

Journal Article↗
PubMed1985

The structure of precursor mRNAs and of excised intron RNAs in chloroplasts of Euglena gracilis.

Partially spliced precursor mRNAs (pre-mRNAs) in the steady-state population of RNA from chloroplasts of Euglena gracilis were found by electron microscopy. The structure and the frequency of the pre-mRNAs of the psbA gene (the gene for the 32-kd protein of photosystem II), which is split by four introns in Euglena chloroplasts was analysed by electron microscopy. A chloroplast DNA (cpDNA) fragment containing the psbA gene from Euglena, was cloned into a pEMBL vector. The single-stranded recombinant phage DNA of the coding strand was prepared and hybridized with cpRNA. The majority of hybrids were formed with mature mRNA, but approximately 8% of the hybrids were formed with pre-mRNAs. The pre-mRNAs were either unspliced or incompletely spliced. A detailed analysis of the structure and the frequency of the pre-mRNAs of the psbA gene showed that the four introns are neither spliced out in a strictly random way, nor in a 5'-3' or 3'-5' direction. Introns 2 and 3 are preferentially spliced out first, intron 1 intermediately and intron 4 is generally spliced out last. However, this sequence is not a strict rule. We conclude that the introns can be spliced independently, each one at a different rate. The coding strand from a fragment of the psbA gene was separated and annealed with low mol. wt. cpRNA, which was isolated from an agarose gel. Small circular hybrids were found at the positions of the four introns, demonstrating for the first time covalently closed circular excised intron RNAs (iRNAs) in chloroplasts.

Animals↗
PubMed1984

Nine introns with conserved boundary sequences in the Euglena gracilis chloroplast ribulose-1,5-bisphosphate carboxylase gene.

The single, chloroplast encoded gene for the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase (rbcL) from Euglena gracilis is found to contain nine intervening sequences. The intervening sequences were identified by heteroduplex analysis between Euglena rbcL and the non-intron-containing rbcL from Spinacea oleracea, by electron microscopy of Euglena rbcL DNA-mRNA hybrids, and by cloning, restriction endonuclease analysis, and partial DNA sequencing. The identification, locus, and coding properties for six of ten exons was confirmed by partial DNA sequence analysis. Each of the nine introns in the approximately 6.5 kb rbcL locus is approximately 0.5 kb in length. The DNA sequence of five 3'-intron/5'-exon and four 3'-exon/5'-intron boundaries are highly conserved. A proposed consensus sequence is (formula; see text) These conserved sequences could play a role in an mRNA splicing mechanism in chloroplasts analogous to that in eucaryotic nuclei.

Base Sequence↗
PubMed1984

Intervening sequences in chloroplast genomes.

Chloroplast DNAs from the green algae Euglena gracilis and from broad beans (Vicia faba) were hybridized with homologous chloroplast RNA. The hybrids were analyzed by electron microscopy. Almost all transcribed regions of the Euglena genome, except for the rRNA genes, contain intervening sequences. A minimum of 50 introns was observed amounting to about 32 kb of the 145 kb genome. Only four spliced transcripts with a total number of six introns (about 0.8 kb each) were detected in the bean chloroplast RNA. A complete transcription map of the Euglena chloroplast genome was established. Transcribed regions occur on the strand that codes for the rRNAs (plus strand) as well as on the complementary strand (minus strand). The polarity of transcription reverses at least six times.

Base Sequence↗