Search PubMed⌕ Search

Biomedical subjects

A Kuhn

Publications and source records attributed to A Kuhn.

At least 145 records · Page 8Linked to original sources
PubMed1990

Initial steps in protein membrane insertion. Bacteriophage M13 procoat protein binds to the membrane surface by electrostatic interaction.

Bacteriophage M13 procoat protein is synthesized on free polysomes prior to its assembly into the inner membrane of Escherichia coli. As an initial step of the membrane insertion pathway, the precursor protein interacts with the cytoplasmic face of the inner membrane. We have used oligonucleotide-directed mutagenesis to study the regions of the procoat protein involved in membrane binding. We find that there is an absolute requirement for positively charged amino acids at both ends of the protein. Replacing these with negatively charged residues resulted in an accumulation of the precursor in the cytoplasm. We propose that the positively charged amino acids are directly involved in membrane binding, possibly directly to the negatively charged phospholipid head groups. This was tested in vitro with artificial liposomes. Whereas wild-type procoat interacted with these liposomes, we found that procoat mutants with negatively charged amino acids at both ends did not bind. Therefore, we conclude that newly synthesized M13 procoat protein binds electrostatically to the negatively charged inner membrane of E. coli.

Amino Acid Sequence↗
PubMed1990

A 140-base-pair repetitive sequence element in the mouse rRNA gene spacer enhances transcription by RNA polymerase I in a cell-free system.

We show that the repetitive 140-base-pair (bp) elements present in the spacer of mouse rRNA genes function as enhancers for RNA polymerase I. Attachment of these elements to the rDNA promoter stimulates rRNA synthesis both in vivo and in vitro. The cis-activating effect of the spacer repeats is orientation-independent and increases with increasing numbers of the 140-bp elements. Competition experiments demonstrate that the spacer repeats bind one or more of the transcription factors interaction with the rDNA promoter. Both the 140-bp elements and the core promoter act cooperatively and thus are functionally linked. The 60/81-bp enhancer repeats from Xenopus laevis rDNA compete for a murine transcription factor(s) and stimulate transcription often fused to the mouse rDNA promoter. The results indicate that despite the marked species specificity of rDNA transcription initiation, common factors may interact with both the rDNA promoter and the enhancer.

Animals↗
PubMed1989

[Lymphomatoid papulosis resembling ecthyma. A case report].

A 54-year-old female patient recurrently developed disseminated papules, red-brown in color and partly ulcerous, which spontaneously disappeared after 3-6 weeks. Histological examination revealed dense infiltration of atypical lymphocytes with epidermotropism. Immunohistological analysis showed predominantly helper T-cells (CD 4). Accordingly, we established the diagnosis of lymphomatoid papulosis. There was no evidence of malignant lymphoma.

Biopsy↗
PubMed1989

Molecular basis for the regulation of cell fate by the lethal (2) giant larvae tumour suppressor gene of Drosophila melanogaster.

Tumour suppressor genes act as recessive determinants of cancer. Their function is required for normal cell growth and differentiation during development. When both alleles of these developmental genes are inactivated, cell growth becomes unrestricted. In Drosophila, a series of genes have been identified which when mutated produce tissue-specific tumours. Of these the lethal(2)giant larvae (l(2)gl) gene is the best studied. Homozygous l(2)gl mutations cause the development of malignant tumours in the brain and the imaginal discs. Genomic DNA from the l(2)gl locus has been cloned, introduced back into l(2)gl mutant animals by P-element-mediated transformation and shown to restore normal development. The nucleotide sequence of the l(2)gl gene (13.1 kb) has been determined, as well as the sequences of the two classes of transcripts. These transcripts encode two polypeptides of 127 kDa and 78 kDa, respectively. Both proteins have been immunologically identified. Analyses of the spatial distribution of both l(2)gl transcripts and proteins revealed that during early embryogenesis the l(2)gl gene is uniformly expressed in all cells and tissues. In late embryos, the l(2)gl expression becomes gradually restricted to tissues presenting no morphological or neoplastic alteration in the mutant animals. Further mosaic experiments pointed out that the critical period for the establishment of tumorigenesis is limited to early embryogenesis at a time when the l(2)gl expression is most intense in all cells.

Animals↗
PubMed1989

3'-end formation of mouse pre-rRNA involves both transcription termination and a specific processing reaction.

We have studied the sequence requirements for 3'-end formation of rDNA transcripts in a cell-free system and show that the generation of correct ends of mouse pre-rRNA is brought about by a two-step process that involves a bona fide termination reaction, followed by a specific trimming of the primary transcript by 10 nucleotides. We show that termination of mouse ribosomal gene transcription by RNA polymerase I (pol I) takes place in front of an 18-bp DNA sequence element (the 'Sal box'), which was previously shown to function as termination signal. Termination of pol I transcription occurs at a fixed distance (11 bp) upstream of the Sal box, independent of the sequence of adjacent gene regions. The processing reaction, however, is strongly influenced by sequences flanking the termination signal at the 5' site. Substitution of a cluster of T residues by guanines within the region of 3'-end formation abolishes the 3'-terminal trimming of the primary transcript. Interestingly, this 3'-terminal processing event, which can be uncoupled from the termination reaction, requires both a correct 3' end and specific sequences in the 3'-terminal region of the primary transcript. Read-through transcripts generated in the extract system or by SP6 RNA polymerase are no substrate for the processing nuclease(s). Because the termination and processing activity can be separated chromatographically, the nucleolytic activity does not reside in TTF-I, the factor that binds to the Sal box and directs transcription termination.

Animals↗
PubMed1989

Immunostaining of keratin and vimentin in epidermis: comparison of different post-embedding immunogold techniques for electron microscopy.

The present study compares different post-embedding staining methods, including conventional and low-temperature embedding techniques, for demonstration of the keratin and vimentin cytoskeleton of epidermal cells, applying commercially available polyclonal and monoclonal antibodies. Immunogold staining (5-nm particles) was performed on the following material: (a) osmium-fixed and Durcupan-embedded material, etched with various solutions; (b) aldehyde-fixed material embedded in Lowicryl K4M at 4 degrees C and -35 degrees C; (c) aldehyde-fixed material embedded in Lowicryl K11M at -60 degrees C with and without cryoprotection with glycerol. In conventionally embedded material we failed to demonstrate intermediate filaments, whereas they were stained after low-temperature embedding with Lowicryl, i.e., K4M 4 degrees C, K4M -35 degrees C, and K11M -60 degrees C. The keratin and vimentin cytoskeleton reacted exclusively with polyclonal antibodies. The best results for antigenicity as well as structural preservation were achieved by post-embedding staining of K4M -35 degrees C-embedded material. Antibodies to keratin stained the cytoskeleton in keratinocytes of all epidermal layers. Filaments were decorated in a linear alignment. Antibodies to vimentin stained the cytoskeleton of Langerhans cells and melanocytes. In these cells a linear distribution pattern of the reaction product along the filaments and an extrafilamentous cluster formation were observed, indicating staining of vimentin and a vimentin-associated protein.

Antibodies, Monoclonal↗
PubMed1988

Alterations in the extracellular domain of M13 procoat protein make its membrane insertion dependent on secA and secY.

The products of genes secA and secY (SecA and SecY) are putative components of a bacterial protein export machinery and are required for the export of many periplasmic and membrane proteins. Only a few proteins, among them the M13 procoat protein, insert independently of SecA and SecY. To investigate the reason why the procoat protein inserts independently of sec functions, various hybrid proteins were constructed. By in-frame gene fusions the central procoat region, which translocates across the membrane, was extended in size. Fragments of the ompA gene ranging from 522-294 bp were ligated with the procoat gene. The hybrid proteins were inserted into the membrane and processed normally, but only in the presence of functional SecA and SecY.

Amino Acid Sequence↗
PubMed1988

[Elastofibroma dorsi].

We report on an 80-years-old woman suffering from subacute prurigo simplex, who had developed a painless subcutaneous tumor on her right upper back, 8 cm in diameter. In palpation, the tumor was firm and did not adhere to the overlying skin. The movability of her arms and back was not restricted. The tumor could not be distinguished from its surrounding tissue by means of ultrasound. Surgery revealed that the tumor was firmly attached to the thoracic fasciae, periosteum, and ligaments, which prevented its complete removal. On histological examination, the tumor was diagnosed as elastofibroma dorsi. We discuss our microscopical findings of this very rare neoplasm in detail.

Aged↗
PubMed1988

[Vater-Pacini neurofibroma].

We report on a 49-year-old female patient suffering from Pacinian neurofibroma on her right middle finger. The microscopical findings of this rare neoplasm are discussed in detail.

Female↗
PubMed1988

The mouse ribosomal gene terminator consists of three functionally separable sequence elements.

The structural requirements for 3' end formation of mouse pre-rRNA have been studied. Three sequence elements are shown to be required for accurate and efficient transcription termination by RNA polymerase I (pol I) assayed both in a cell-free transcription system and in vivo after transfection of rDNA minigene constructs into 3T6 cells. The essential termination signal is the previously identified 18-bp conserved element (AGGTCGACCAGATTANTCCG) that contains a SalI restriction site. This sequence motif (the 'Sal box') interacts with a specific nuclear protein that directs transcription termination. Here we demonstrate that the 'Sal box' sequence motif is sufficient for termination of pol I transcripts and the release of the nascent RNA chains from the template. However, in addition to this termination signal, pyrimidine-rich sequences flanking the box at the 5' and 3' side play a role in the efficient and correct formation of authentic pre-rRNA termini. Downstream sequences contribute to the efficiency of the termination reaction, whereas the position of 3' end formation (i.e. 21 bp upstream of the 'Sal box') is affected by 5' flanking regions. These flanking regions are recognized by at least two different nuclear factors which specifically bind to DNA sequences located upstream and downstream of the 'Sal box'.

Animals↗
PubMed1988

[Vesicular pityriasis rosea].

Pityriasis rosea is a frequently occurring skin disease of unknown aetiology. Ten clinical forms of this disease are known, with predominance of the macular type. We observed a healthy 24-year-old patient with the very rare vesicular variant of pityriasis rosea, which has to be differentiated from vesicular virus and drug-induced eruptions.

Adult↗
PubMed1988

Malignant blue nevus with metastases to the lung.

A malignant blue nevus of the right upper arm with hematogeneous lung metastases is presented. Histological examination showed that the tumor was composed of spindle, dendritic, and globular cells with hyperchromatic and polymorphic nuclei, atypical mitoses, and tumor cell necrosis. There was no proliferation of atypical melanocytes at the dermoepidermal junction. Histologically, malignant blue nevus should be distinguished from benign cellular blue nevus, primary cutaneous malignant melanoma, cutaneous metastases of malignant melanoma, and clear cell sarcoma.

Adult↗
PubMed1988

[Immunohistological studies of immune dermatoses on reconstituted paraffin sections. Value and comparison of different methods].

In a study on 26 patients with autoimmune diseases the immunostaining for immunoglobulins and complement in frozen sections was compared with that in formalin-fixed and paraffin-embedded sections. The formalin-fixed material of pemphigus vulgaris, lupus erythematosus, bullous pemphigoid, and dermatitis herpetiformis (Duhring's disease) was reconstituted and stained by means of the standard two-step technique (TST), the peroxidase-antiperoxidase technique (PAP), and the streptavidin-biotin method (SAB). In comparison with frozen sections, immunoglobulins and complement could also be demonstrated in formalin-fixed sections in all cases of pemphigus vulgaris and in 85% of cases of discoid lupus erythematosus, but in only 60% of cases of bullous pemphigoid or Duhring's disease. PAP and SAB proved to be about equally reliable, but TST was significantly less dependable.

Dermatitis Herpetiformis↗
PubMed1987

Bacteriophage M13 procoat protein inserts into the plasma membrane as a loop structure.

The major coat protein of bacteriophage M13 is synthesized as a precursor, the procoat, with a typical leader (signal) sequence of 23 residues at its NH2-terminus. A fusion protein that contains the NH2-terminal 141 residues of cytoplasmic ribulokinase and all but the first ten residues of M13 procoat was made. The fusion protein inserts into the plasma membrane of Escherichia coli and is processed by leader peptidase to give rise to a leader peptide of 155 residues and the mature coat protein of 50 residues. The NH2-terminus of the leader peptide remains in the cytoplasm and is protected from protease added to the medium outside of the cell. This indicates that M13 procoat inserts into the membrane as a loop structure and that the NH2-terminus of a leader peptide remains within the cytoplasm during membrane insertion.

Amino Acid Sequence↗
PubMed1987

The internal signal sequence of Escherichia coli leader peptidase is necessary, but not sufficient, for its rapid membrane assembly.

Leader peptidase of Escherichia coli, a protein of 323 residues, has three hydrophobic domains. The first, residues 1-22, is the most apolar and is followed by a polar region (23-61) which faces the cytoplasm. The second hydrophobic domain (residues 62-76) spans the membrane. The third hydrophobic domain, which has a minimal apolar character, and the polar, carboxyl-terminal two-thirds of the protein are exposed to the periplasm. Deletion of either the amino terminus (residues 4-50) or the third hydrophobic region (residues 83-98) has almost no effect on the rate of leader peptidase membrane assembly, while the second hydrophobic domain is essential for insertion (Dalbey, R., and Wickner, W. (1987) Science 235, 783-787). To further define the roles of these domains, we have replaced the normal, cleaved leader sequence of pro-OmpA and M13 procoat with regions containing either the first or second apolar domain of leader peptidase. The second apolar domain supports the translocation of OmpA or coat protein across the plasma membrane, establishing its identity as an internal, uncleaved signal sequence. In addition to this sequence, we now find that leader peptidase needs either the amino-terminal domain or the third hydrophobic domain to permit its rapid membrane assembly. These results show that, although a signal sequence is necessary for rapid membrane assembly of leader peptidase, it is not sufficient.

Base Sequence↗