Medizin - Open Access LMU - Teil 07/22 Archiv

The chicken lysozyme gene is constitutively expressed in macrophages, in oviduct cells its expression is controlled by steroid hormones, and in fibroblasts the gene is not expressed. A fusion gene consisting of promoter sequences of the lysozyme gene from –208 to +15 in front of the chloramphenicol acetyltransferase (CAT) coding region was more than 50 times less active in non-expressing cells as compared to expressing cells. In order to identify the element(s) responsible for this cell-type specificity 31 different linker scanning mutations were generated within this promoter fragment and analyzed by transient transfections in the three types of chicken cells mentioned above. Three mutation sensitive regions located around position –25, –100 and between –158 and –208 were detected in each cell type, however, several LS mutations displayed clear cell-type specific differences in their phenotypic effects. Interestingly, a few LS mutations led to an increase in promoter activity in fibroblasts suggesting that the corresponding wildtype sequences represent binding sites for negatively acting transcription factors.

Zwei deutsche Tropenreisende erkrankten nach einer zweitägigen Safari im Akagera-Nationalpark in Ruanda an ostafrikanischer Trypanosomiasis (Schlafkrankheit). Leitsymptome waren Fieber, Lymphadenopathie und eine typische Primärläsion (Trypanosomenschanker). Die Diagnose konnte durch den Nachweis von Trypanosomen im peripheren Blut gesichert werden. Das Zentralnervensystem war in beiden Fällen nicht beteiligt. Unter dem Einfluß einer Therapie mit Suramin, 1 g pro Woche intravenös über 6 Wochen, bildeten sich die Symptome und die Parasitämie rasch zurück. Nach der Zahl der seit 1970 berichteten Fälle ergibt sich für deutsche Tropenreisende ein Infektionsrisiko von 0,3 pro 100 000. Aufgrund der teilweise erheblichen Wiederzunahme der Schlafkrankheit in einigen afrikanischen Ländern kann mit einer Zunahme des Infektionsrisikos auch für Touristen gerechnet werden.

Passage of precursor proteins through translocation contact sites of mitochondria was investigated by studying the import of a fusion protein consisting of the NH2-terminal 167 amino acids of yeast cytochrome b2 precursor and the complete mouse dihydrofolate reductase. Isolated mitochondria of Neurospora crassa readily imported the fusion protein. In the presence of methotrexate import was halted and a stable intermediate spanning both mitochondrial membranes at translocation contact sites accumulated. The complete dihydrofolate reductase moiety in this intermediate was external to the outer membrane, and the 136 amino acid residues of the cytochrome b2 moiety remaining after cleavage by the matrix processing peptidase spanned both outer and inner membranes. Removal of methotrexate led to import of the intermediate retained at the contact site into the matrix. Thus unfolding at the surface of the outer mitochondrial membrane is a prerequisite for passage through translocation contact sites. The membrane-spanning intermediate was used to estimate the number of translocation sites. Saturation was reached at 70 pmol intermediate per milligram of mitochondrial protein. This amount of translocation intermediates was calculated to occupy approximately 1% of the total surface of the outer membrane. The morphometrically determined area of close contact between outer and inner membranes corresponded to approximately 7% of the total outer membrane surface. Accumulation of the intermediate inhibited the import of other precursor proteins suggesting that different precursor proteins are using common translocation contact sites. We conclude that the machinery for protein translocation into mitochondria is present at contact sites in limited number.

The covalent attachment of heme to apocytochrome c, and therefore the import of cytochrome c into mitochondria, is dependent on both NADH plus a cytosolic cofactor that has been identified to be FMN or FAD. NADH in concert with flavin nucleotides mediates the reduction of heme. Heme in the reduced state is a prerequisite for its covalent attachment to apocytochrome c by the enzyme cytochrome c heme lyase and thus for subsequent translocation of cytochrome c across the outer mitochondrial membrane during import.