Biomaterial Database

Uridylate-specific 3' 5'-exoribonucleases involved in uridylate-deletion RNA editing in trypanosomatid mitochondria. 2007

Kestrel Rogers, and Guanghan Gao, and Larry Simpson

Department of Microbiology, Immunology, and Molecular Genetics, David Geffen School of Medicine at UCLA, Los Angeles, California 90095, USA.

In kinetoplastid protists, maturation of mitochondrial pre-mRNAs involves the insertion and deletion of uridylates (Us) within coding regions, as specified by mitochondrial DNA-encoded guide RNAs. U-deletion editing involves endonucleolytic cleavage of the pre-mRNA at the editing site followed by U-specific 3'-5'-exonucleolytic removal of nonbase-paired Us prior to ligation of the two mRNA cleavage fragments. We showed previously that an exonuclease/endonuclease/phosphatase (EEP) motif protein from Leishmania major, designated RNA editing exonuclease 1 (REX1) (Kang, X., Rogers, K., Gao, G., Falick, A. M., Zhou, S.-L., and Simpson, L. (2005) Proc. Natl. Acad. Sci. U. S. A. 102, 1017-1022), exhibits 3'-5'-exonuclease activity. Two EEP motif proteins have also been identified in the Trypanosoma brucei editing complex. TbREX1 is a homologue of LmREX1, and TbREX2 shows homology to another editing protein in L. major, which lacks the EEP motif (LmREX2*). Here we have expressed the T. brucei EEP motif proteins in insect cells and purified them to homogeneity. We showed that these are U-specific 3'-5'-exonucleases that are inhibited by base pairing of 3' Us. The recombinant EEP motif alone also showed 3'-5' U-specific exonuclease activity, and mutations of the REX EEP motifs greatly reduced exonuclease activity. The absence of enzymatic activity in LmREX2* was confirmed with a purified recombinant protein. We showed that pre-cleaved U-deletion editing could be reconstituted with either TbREX1 or TbREX2 in combination with either RNA ligase, LmREL1, or LmREL2. Down-regulation of TbREX2 expression by conditional RNA interference had little effect on parasite viability or sedimentation of the L-complex, suggesting either that TbREX2 is inactive in vivo or that TbREX1 can compensate for the loss of TbREX2 function in down-regulated cells.

UI	MeSH Term	Description	Entries
D008928	Mitochondria	Semiautonomous, self-reproducing organelles that occur in the cytoplasm of all cells of most, but not all, eukaryotes. Each mitochondrion is surrounded by a double limiting membrane. The inner membrane is highly invaginated, and its projections are called cristae. Mitochondria are the sites of the reactions of oxidative phosphorylation, which result in the formation of ATP. They contain distinctive RIBOSOMES, transfer RNAs (RNA, TRANSFER); AMINO ACYL T RNA SYNTHETASES; and elongation and termination factors. Mitochondria depend upon genes within the nucleus of the cells in which they reside for many essential messenger RNAs (RNA, MESSENGER). Mitochondria are believed to have arisen from aerobic bacteria that established a symbiotic relationship with primitive protoeukaryotes. (King & Stansfield, A Dictionary of Genetics, 4th ed)	Mitochondrial Contraction,Mitochondrion,Contraction, Mitochondrial,Contractions, Mitochondrial,Mitochondrial Contractions
D008969	Molecular Sequence Data	Descriptions of specific amino acid, carbohydrate, or nucleotide sequences which have appeared in the published literature and/or are deposited in and maintained by databanks such as GENBANK, European Molecular Biology Laboratory (EMBL), National Biomedical Research Foundation (NBRF), or other sequence repositories.	Sequence Data, Molecular,Molecular Sequencing Data,Data, Molecular Sequence,Data, Molecular Sequencing,Sequencing Data, Molecular
D011994	Recombinant Proteins	Proteins prepared by recombinant DNA technology.	Biosynthetic Protein,Biosynthetic Proteins,DNA Recombinant Proteins,Recombinant Protein,Proteins, Biosynthetic,Proteins, Recombinant DNA,DNA Proteins, Recombinant,Protein, Biosynthetic,Protein, Recombinant,Proteins, DNA Recombinant,Proteins, Recombinant,Recombinant DNA Proteins,Recombinant Proteins, DNA
D005095	Exoribonucleases	A family of enzymes that catalyze the exonucleolytic cleavage of RNA. It includes EC 3.1.13.-, EC 3.1.14.-, EC 3.1.15.-, and EC 3.1.16.-. EC 3.1.-	Exoribonuclease
D005786	Gene Expression Regulation	Any of the processes by which nuclear, cytoplasmic, or intercellular factors influence the differential control (induction or repression) of gene action at the level of transcription or translation.	Gene Action Regulation,Regulation of Gene Expression,Expression Regulation, Gene,Regulation, Gene Action,Regulation, Gene Expression
D000595	Amino Acid Sequence	The order of amino acids as they occur in a polypeptide chain. This is referred to as the primary structure of proteins. It is of fundamental importance in determining PROTEIN CONFORMATION.	Protein Structure, Primary,Amino Acid Sequences,Sequence, Amino Acid,Sequences, Amino Acid,Primary Protein Structure,Primary Protein Structures,Protein Structures, Primary,Structure, Primary Protein,Structures, Primary Protein
D000818	Animals	Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA.	Animal,Metazoa,Animalia
D014346	Trypanosoma brucei brucei	A hemoflagellate subspecies of parasitic protozoa that causes nagana in domestic and game animals in Africa. It apparently does not infect humans. It is transmitted by bites of tsetse flies (Glossina).	Trypanosoma brucei,Trypanosoma brucei bruceus,Trypanosoma bruceus,brucei brucei, Trypanosoma,brucei, Trypanosoma brucei,bruceus, Trypanosoma,bruceus, Trypanosoma brucei
D014542	Uridine Monophosphate	5'-Uridylic acid. A uracil nucleotide containing one phosphate group esterified to the sugar moiety in the 2', 3' or 5' position.	UMP,Uridylic Acid,Uridine 5'-Monophosphate,Uridylic Acids,5'-Monophosphate, Uridine,Acid, Uridylic,Acids, Uridylic,Monophosphate, Uridine,Uridine 5' Monophosphate
D015536	Down-Regulation	A negative regulatory effect on physiological processes at the molecular, cellular, or systemic level. At the molecular level, the major regulatory sites include membrane receptors, genes (GENE EXPRESSION REGULATION), mRNAs (RNA, MESSENGER), and proteins.	Receptor Down-Regulation,Down-Regulation (Physiology),Downregulation,Down Regulation,Down-Regulation, Receptor