Trypanosoma brucei spp. cause African human and animal trypanosomiasis, a burden on health and economy in Africa. These hemoflagellates are distinguished by a kinetoplast nucleoid containing mitochondrial DNAs of two kinds: maxicircles encoding ribosomal RNAs (rRNAs) and proteins and minicircles bearing guide RNAs (gRNAs) for mRNA editing. All RNAs are produced by a phage-type RNA polymerase as 3' extended precursors, which undergo exonucleolytic trimming. Most pre-mRNAs proceed through 3' adenylation, uridine insertion/deletion editing, and 3' A/U-tailing. The rRNAs and gRNAs are 3' uridylated. Historically, RNA editing has attracted major research effort, and recently essential pre- and postediting processing events have been discovered. Here, we classify the key players that transform primary transcripts into mature molecules and regulate their function and turnover.

• Klíčová slova
• RNA decay, RNA editing, Trypanosoma, kinetoplast, mitochondria, polyadenylation,
• MeSH
• editace RNA fyziologie   MeSH
• RNA mitochondriální genetika   metabolismus   MeSH
• RNA protozoální genetika   metabolismus   MeSH
• Trypanosoma brucei brucei genetika   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• RNA mitochondriální MeSH
• RNA protozoální MeSH

RNA-binding proteins (RBPs) are critical to posttranscriptional gene regulation. Therefore, characterization of the RNA molecules bound by RBPs in vivo represent a key step in elucidating their function. The recently developed iCLIP technique allows single nucleotide resolution of the RNA binding footprints of RBPs. We present the iCLIP technique modified for its application to Trypanosoma brucei and most likely other kinetoplastid flagellates. By using the immuno- or affinity purification approach, it was successfully applied to the analysis of several RBPs. Furthermore, we also provide a detailed description of the iCLIP/iCLAP protocol that shall be particularly suitable for the studies of trypanosome RBPs.

• Klíčová slova
• Posttranscriptional gene regulation, RNA-binding proteins (RBPs), iCLAP, iCLIP,
• MeSH
• imunoprecipitace metody   MeSH
• nukleotidy genetika   metabolismus   MeSH
• parazitologie metody   MeSH
• proteiny vázající RNA analýza   genetika   metabolismus   MeSH
• protozoální proteiny analýza   genetika   metabolismus   MeSH
• RNA protozoální genetika   metabolismus   MeSH
• RNA genetika   metabolismus   MeSH
• Trypanosoma brucei brucei genetika   MeSH
• ultrafialové záření MeSH
• vazba proteinů genetika   účinky záření   MeSH
• vazebná místa genetika   MeSH
• zobrazení jednotlivé molekuly metody   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• nukleotidy MeSH
• proteiny vázající RNA MeSH
• protozoální proteiny MeSH
• RNA protozoální MeSH
• RNA MeSH

MRP1/2 is a heteromeric protein complex that functions in the trypanosomatid mitochondrion as part of the RNA editing machinery, which facilitates multiple targeted insertions and deletions of uridines. MRP1/2 was shown to interact with MRB8170, which initiates RNA editing by marking pre-edited mRNAs, while TbRGG2 is required for its efficient progression on pan-edited mRNAs. Both MRP1/2 and TbRGG2 are capable of modulating RNA-RNA interactions in vitro. As determined by using iCLIP and RIP-qPCR, RNAs bound to MRP1/2 are characterized and compared with those associated with MRB8170 and TbRGG2. We provide evidence that MRP1 and MRB8170 have correlated binding and similar RNA crosslinking peak profiles over minimally and never-edited mRNAs. Our results suggest that MRP1 assists MRB8170 in RNA editing on minimally edited mRNAs.

• Klíčová slova
• RNA binding proteins, RNA editing, iCLIP, mitochondrion, ribonuclear protein, trypanosome,
• MeSH
• editace RNA MeSH
• messenger RNA genetika   metabolismus   MeSH
• mitochondrie genetika   metabolismus   MeSH
• proteiny vázající RNA metabolismus   MeSH
• protozoální proteiny genetika   metabolismus   MeSH
• RNA mitochondriální genetika   metabolismus   MeSH
• Trypanosoma genetika   metabolismus   MeSH
• vazba proteinů MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• gBP21 protein, Trypanosoma brucei MeSH Prohlížeč
• messenger RNA MeSH
• mitochondrial messenger RNA MeSH Prohlížeč
• proteiny vázající RNA MeSH
• protozoální proteiny MeSH
• RNA mitochondriální MeSH