Transport of Proteins into Mitochondria

The mitochondrial ADP/ATP carrier is an integral transmembrane protein of the inner membrane. It is synthesized on cytoplasmic ribosomes. Kinetic data suggested that this protein is transferred into mitochondria in a posttranslational manner. The following results provide further evidence for such a mechanism and provide information on its details. 1. In homologous and heterologous translation systems the newly synthesized ADP/ATP carrier protein is present in the postribosomal supernatant. 2. Analysis by density gradient centrifugation and gel filtration shows, that the ADP/ATP carrier molecules in the postribosomal fraction are present as soluble complexes with apparent molecular weights of about 120000 and 500000 or larger. The carrier binds detergents such as Triton X-100 and deoxycholate forming mixed micelles with molecular weights of about 200000–400000. 3. Incubation of a postribosomal supernatant of a reticulocyte lysate containing newly synthesized ADP/ATP carrier with mitochondria isolated from Neurospora spheroplasts results in efficient transfer of the carrier into mitochondria. About 20–30% of the transferred carrier are resistant to proteinase in whole mitochondria. The authentic mature protein is also largely resistant to proteinase in whole mitochondria and sensitive after lysis of mitochondria with detergent. Integrity of mitochondria is a prerequisite for translocation into proteinase resistant position. 4. The transfer in vitro into a proteinase-resistant form is inhibited by the uncoupler carbonyl-cyanide m-chlorophenylhydrazone but not the proteinase-sensitive binding. These observations suggest that the posttranslational transfer of ADP/ATP carrier occurs via the cytosolic space through a soluble oligomeric precursor form. This precursor is taken up by intact mitochondria into an integral position in the membrane. These findings are considered to be of general importance for the intracellular transfer of insoluble membrane proteins. They support the view that such proteins can exist in a water-soluble form its precursors and upon integration into the membrane undergo a conformational change. Uptake into the membrane may involve the cleavage of an additional sequence in some proteins, but this appears not to be a prerequisite as demonstrated by the ADP/ATP carrier protein

Neem-tree (Azadirachta indica Juss.) Extract as a Feed Additive Against the American Dog Tick (Dermacentor variabilis) in Sheep (Ovis aries)

Acaricides can be conveyed to ticks via the blood of their hosts. As fruit and kernel extracts from the Meliaceae family, and, in particular the tetranortriterpenoid azadirachtin (AZA) inhibits tick egg production and embryogenesis in the Ixodidae ticks, we investigated the effects of Neem Azal®, an extract containing 43% AZA, given as a feed additive to lambs artificially infested with engorging adult Dermacentor vairiabilis ticks. After tick attachment, the lambs were allotted to three dietary treatments: AZA0 (control, n = 10), AZA0.3 (n = 5), and AZA0.6 (n = 5), with feed containing 0%, 0.3%, and 0.6% AZA on DM basis, respectively. In half of the AZA0 lambs, ticks were sprayed on day 4 after attachment with an ethanol:water:soap emulsion containing 0.6% AZA (AZA0S). In spite of its very pungent odor, the neem extract was well accepted by all but one lamb. No differences were found between treatment groups in liver enzymes in blood, and there was no indication of toxicity. The plasma AZA concentrations after 7 and 14 days of feeding AZA were (4.81 and 4.35 μg/mL) for the AZA0.6 and (3.32 and 1.88 μg/mL) for the AZA0.3 treatments, respectively (P \u3c 0.0001). Treatments were not lethal to ticks, but tick weights at detachment were 0.64, 0.56, 0.48, and 0.37 g for ticks from the AZA0, AZA0.3, AZA0S, and AZA0.6 treatments (P \u3c 0.04), respectively, suggesting that blood AZA impaired blood-feeding. The highest mortality rate after detachment was for AZA0.6 (P \u3c 0.09). As AZA affects embryo development and ticks at the molting stages, we expect that following treatments of hosts for longer periods, one-host ticks will be more affected than the three-host tick D. variabilis

Neem-Tree (\u3ci\u3eAzadirachta indica\u3c/i\u3e Juss.) Extract as a Feed Additive Against the American Dog Tick (\u3ci\u3eDermacentor variabilis\u3c/i\u3e) in Sheep (\u3ci\u3eOvis aries\u3c/i\u3e)

Acaricides can be conveyed to ticks via the blood of their hosts. As fruit and kernel extracts from the Meliaceae family, and, in particular the tetranortriterpenoid azadirachtin (AZA) inhibits tick egg production and embryogenesis in the Ixodidae ticks, we investigated the effects of Neem Azal®, an extract containing 43% AZA, given as a feed additive to lambs artificially infested with engorging adult Dermacentor vairiabilis ticks. After tick attachment, the lambs were allotted to three dietary treatments: AZA0 (control, n=10), AZA0.3 (n=5), and AZA0.6 (n=5), with feed containing 0%, 0.3%, and 0.6% AZA on DM basis, respectively. In half of the AZA0 lambs, ticks were sprayed on day 4 after attachment with an ethanol:water:soap emulsion containing 0.6% AZA (AZA0S). In spite of its very pungent odor, the neem extract was well accepted by all but one lamb. No differences were found between treatment groups in liver enzymes in blood, and there was no indication of toxicity. The plasma AZA concentrations after 7 and 14 days of feeding AZA were (4.81 and 4.35 µg/mL) for the AZA0.6 and (3.32 and 1.88 µg/mL) for the AZA0.3 treatments, respectively (P\u3c0.0001). Treatments were not lethal to ticks, but tick weights at detachment were 0.64, 0.56, 0.48, and 0.37 g for ticks from the AZA0, AZA0.3, AZA0S, and AZA0.6 treatments (P\u3c0.04), respectively, suggesting that blood AZA impaired blood-feeding. The highest mortality rate after detachment was for AZA0.6 (P\u3c0.09). As AZA affects embryo development and ticks at the molting stages, we expect that following treatments of hosts for longer periods, one-host ticks will be more affected than the three-host tick D. variabilis