RNA editing sites in plant mitochondria and plastids are addressed by pentatricopeptide repeat (PPR) proteins with E or E and DYW domains, which recognize a specific nucleotide motif upstream of the edited nucleotide. In addition, some sites require MORF proteins for efficient RNA editing. Here, we assign the novel E domain-containing PPR protein, MEF13, as being required for editing at eight sites in Arabidopsis thaliana. A SNP in ecotype C24 altering the editing level at only one of the eight target sites was located by genomic mapping. An EMS mutant allele of the gene for MEF13 was identified in a SNaPshot screen of a mutated plant population. At all eight target sites of MEF13, editing levels are reduced in both morf3 and morf8 mutants, but at only one site in morf1 mutants, suggesting that specific MEF13-MORF interactions are required. Yeast two-hybrid analyses detect solid connections of MEF13 with MORF1 and weak contact with MORF3 proteins. Yeast three-hybrid (Y3H) analysis shows that the presence of MORF8 enhances the connection between MEF13 and MORF3, suggesting that a MORF3-MORF8 heteromer may form stably or transiently to establish interaction with MEF13.
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http://dx.doi.org/10.1016/j.molp.2015.05.008 | DOI Listing |
Kardiol Pol
January 2025
1st Department of Cardiology, Poznan University of Medical Sciences, Poznań, Poland.
PLoS One
December 2024
Department of Microbiology and Immunology, McGill University, Montreal, Quebec, Canada.
The ability to determine the essentiality of a gene in the protozoan parasite Leishmania is important to identify potential targets for intervention and understanding the parasite biology. CRISPR gene editing technology has significantly improved gene targeting efficiency in Leishmania. There are two commonly used CRISPR gene targeting methods in Leishmania; the stable expression of the gRNA and Cas9 using a plasmid containing a Leishmania ribosomal RNA gene promoter (rRNA-P stable protocol) and the T7 RNA polymerase based transient gRNA expression system in promastigotes stably expressing Cas9 (T7 transient protocol).
View Article and Find Full Text PDFNat Genet
January 2025
Institute of Molecular Oncology, Philipps-University, Marburg, Germany.
The mutational landscape of TP53, a tumor suppressor mutated in about half of all cancers, includes over 2,000 known missense mutations. To fully leverage TP53 mutation status for personalized medicine, a thorough understanding of the functional diversity of these mutations is essential. We conducted a deep mutational scan using saturation genome editing with CRISPR-mediated homology-directed repair to engineer 9,225 TP53 variants in cancer cells.
View Article and Find Full Text PDFNat Commun
January 2025
Interdisciplinary Life Sciences Graduate Programs, University of Texas at Austin, Austin, TX, 78712, USA.
Type II CRISPR endonucleases are widely used programmable genome editing tools. Recently, CRISPR-Cas systems with highly compact nucleases have been discovered, including Cas9d (a type II-D nuclease). Here, we report the cryo-EM structures of a Cas9d nuclease (747 amino acids in length) in multiple functional states, revealing a stepwise process of DNA targeting involving a conformational switch in a REC2 domain insertion.
View Article and Find Full Text PDFSci Data
January 2025
Gakushuin University, Faculty of Science, Department of Life Science, Mejiro 1-5-1, Toshima-ku, Tokyo, 171-8588, Japan.
The wild silk moth, Bombyx mandarina, is the closest relative of the domesticated silk moth, Bombyx mori. National BioResource Project of Japan (NBRP) maintains a B. mandarina strain derived from individuals captured at Sakado (Saitama, Japan) in 1982.
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