1/2/2023 0 Comments Sp flash tool v5 1352.01![]() brucei is comprised of ∼50 identical maxicircles and thousands of heterogeneous minicircles ( 12, 13). Kinetoplastids are named for their distinctive mitochondrial DNA network, known as the kinetoplast DNA (kDNA), within their single mitochondrion. ![]() Indeed, a number of these processes, such as trans splicing, polycistronic transcription, antigenic variation, glycosylphosphatidylinositol anchoring, and mitochondrial RNA editing, were first described in trypanosomes and provided novel paradigms for eukaryotic biology ( 1, – 11). brucei is a deeply divergent eukaryote, the study of which advanced the understanding of many fundamental biological processes and eukaryotic evolution. ![]() The kinetoplastid parasite Trypanosoma brucei is the etiologic agent of human African trypanosomiasis, which is transmitted by the tsetse fly and is a health threat to millions of people in sub-Saharan Africa. Hence, the degenerate RNase III domain and a newly identified domain are critical for KREPB5 function and have differential effects between the life cycle stages of T. Amino acid changes at a substantial number of positions, including up to 7 per allele, allowed complementation and thus did not block KREPB5 function. Eight of these mutations were lethal in bloodstream form parasites but not in procyclic-form parasites, showing that multiple domains function in a life cycle-dependent manner. Exclusive expression of these mutated KREPB5 alleles in the absence of wild-type allele expression resulted in growth inhibition, the loss of ∼20S editosomes, and inhibition of RNA editing in BF cells. Seven of these were within the RNase III domain, and two were in the C-terminal region that has no homology to known motifs. We identified nine single amino acid substitutions that could not complement the conditional loss of wild-type KREPB5. brucei bloodstream form cell lines that express randomly mutagenized KREPB5 alleles. To explore the function of this protein, we used a novel approach to make and screen numerous conditional null T. KREPB5 is an essential component of ∼20S editosomes in Trypanosoma brucei which contains a degenerate, noncatalytic RNase III domain.
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