Evidence for Trans splicing in trypanosomes

Sutton, Richard E.; Boothroyd, John C.

doi:10.1016/0092-8674(86)90617-3

Cited by 456 publications

(307 citation statements)

References 47 publications

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“…Furthermore, the tubulin gene family organization displays noticeable variation among several T. cruzi stocks and cloned lines analysed [3]. Besides the peculiar organization of the tubulin genes in trypanosomatids, the mRNA synthesis in these organisms and other kinetoplastids has been shoran to present puzzling features like the addition of a common leader sequence (mini-exon) to the 5' termini of all mRNAs, carried out by a process of trans-splicing [5][6][7][8][9]. Equally intriguing is the phenomenon called RNA editing of mitochondrial transcripts.…”

Section: Introductionmentioning

confidence: 99%

α‐ and β‐tubulin mRNAs of Trypanosoma cruzi originate from a single multicistronic transcript

et al. 1989

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The cluster of alternated r,. and fl-tubulin genes in the genome of Trypanosoma cruzi was shown to be transcribed into a single RNA molecule which upon processing gives rise to the mature ~-and fl-tubulin mRNAs. This conclusion was based on: (i) nuclear RNA species with the same molecular mass hybridize to both a-and fl-tubulin eDNA probes; (ii) $1 nuclease assay of the clustered tubulin genes has shown protected DNA fragments of the same size and of greater molecular mass than that corresponding to the mRNAs, hybridizable to both ~t-and fl-tubulin eDNA probes; (iii) fl-tubulin hybrid selected RNA is still able to hybridize to ~t-tubulin probe.

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Section: Introductionmentioning

confidence: 99%

α‐ and β‐tubulin mRNAs of Trypanosoma cruzi originate from a single multicistronic transcript

et al. 1989

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“…cDNA was prepared from total RNA using random hexamer oligonucleotides and the Superscript reverse transcription kit (Life Technologies, Inc.). The sense primer, 5Ј-CCAACGCTATATAAGTATCAGTTTCTGTACTTTATTG-3Ј, in the PCR reaction was designed to the L. donovani mini-exon (23) that is trans-spliced onto the 5Ј end of all leishmanial mRNAs (24), whereas the antisense primer 5Ј-CCTTCTCCTCGTCAATCAG-3Ј was designed to amino acid residues 218 -223 of the LdPEX5 sequence. PCR was performed using a 3-min hot start at 95°C and 35 cycles of denaturation at 94°C for 1 min, 62°C for 1 min, and extension at 72°C for 2 min with Taq DNA polymerase (Promega, Madison, WI).…”

Section: Chemicals and Reagents-[mentioning

confidence: 99%

Peroxisomal Targeting Signal-1 Receptor Protein PEX5 fromLeishmania donovani

Jardim

Liu

Zheleznova

et al. 2000

Journal of Biological Chemistry

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The human pathogens of the Leishmania and Trypanosoma genera compartmentalize glycolytic and other key metabolic pathways in unique subcellular microbodies called glycosomes, organelles related to the peroxisomes of mammals and yeast. The molecular machinery that carries out the specific targeting of glycosomal proteins to the organelle has not been characterized, although the bulk of glycosomal proteins contain the COOH-terminal tripeptide glycosomal peroxisomal targeting signal-1 (PTS-1) similar to the mammalian and fungal peroxisomal targeting signal. To characterize the mechanisms of glycosomal targeting, the gene encoding PEX5, designated LdPEX5, has been isolated from Leishmania donovani. LdPEX5 encodes a 625-amino acid protein with a molecular mass of 69.7 kDa. Like its homologs in yeast and humans, LdPEX5 predicts a protein with seven copies of a tetratricopeptide repeat in its COOH-terminal half proposed to mediate PTS-1 binding and three copies of a WXXX(Y/F) motif in its NH 2 terminus conjectured to be essential for protein translocation into the organelle. LdPEX5 was overexpressed in Escherichia coli and purified to homogeneity for binding experiments and generation of antibodies. Recombinant LdPEX5 bound xanthine phosphoribosyltransferase (XPRT), a PTS-1 containing glycosomal protein with a K D of 4.2 nM, but did not bind an XPRT in which the PTS-1 had been deleted. Moreover, binding studies with the COOH-terminal half of the LdPEX5 confirmed that this portion of the PEX5 protein was capable of binding the XPRT PTS-1 with an affinity of 17.3 nM. Confocal microsocopy revealed that LdPEX5 was predominantly in the cytosolic milieu, and genetic analysis implied that LdPEX5 was an essential gene.

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“…Finally, the trans-splice site is at the 59 end of the mRNA, not the pre-mRNA, and thus, the promoter is often not directly adjacent to the gene, but rather upstream of the outron or the entire operon (Blumenthal and Spieth 1996). SL trans-splicing has been reported in many phyla, including trypanosomes, nematodes, and even chordates (Sutton and Boothroyd 1986;Krause and Hirsh 1987;Vandenberghe et al 2001). In 1994, it was estimated that 70% of C. elegans genes were trans-spliced, based on the limited genomic and cDNA sequence data available (Zorio et al 1994).…”

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confidence: 99%

“…SL trans-splicing has been reported in many phyla, including trypanosomes, nematodes, and even chordates (Sutton and Boothroyd 1986;Krause and Hirsh 1987;Vandenberghe et al 2001). In 1994, it was estimated that 70% of C. elegans genes were trans-spliced, based on the limited genomic and cDNA sequence data available (Zorio et al 1994).…”

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A global analysis of C. elegans trans-splicing

Allen¹,

Hillier²,

Waterston³

et al. 2010

Genome Res.

158

256

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Trans-splicing of one of two short leader RNAs, SL1 or SL2, occurs at the 59 ends of pre-mRNAs of many C. elegans genes. We have exploited RNA-sequencing data from the modENCODE project to analyze the transcriptome of C. elegans for patterns of trans-splicing. Transcripts of~70% of genes are trans-spliced, similar to earlier estimates based on analysis of far fewer genes. The mRNAs of most trans-spliced genes are spliced to either SL1 or SL2, but most genes are not transspliced to both, indicating that SL1 and SL2 trans-splicing use different underlying mechanisms. SL2 trans-splicing occurs in order to separate the products of genes in operons genome wide. Shorter intercistronic distance is associated with greater use of SL2. Finally, increased use of SL1 trans-splicing to downstream operon genes can indicate the presence of an extra promoter in the intercistronic region, creating what has been termed a ''hybrid'' operon. Within hybrid operons the presence of the two promoters results in the use of the two SL classes: Transcription that originates at the promoter upstream of another gene creates a polycistronic pre-mRNA that receives SL2, whereas transcription that originates at the internal promoter creates transcripts that receive SL1. Overall, our data demonstrate that >17% of all C. elegans genes are in operons.[Supplemental material is available for this article. The sequence data from this study have been submitted to the NCBI Sequence Read Archive (http://www.ncbi.nlm.nih.gov/Traces/sra/sra.cgi) under accession nos. SRA008646 and SRA003622.] C. elegans uses two RNA processing features that distinguish it from other model organisms. First, the transcripts of many genes are trans-spliced to a spliced leader (SL). Trans-splicing is a process in which an SL replaces the 59 end of a transcript by spliceosomal splicing. The 22-nucleotide (nt) SL is donated by an ;100-nt SL snRNP (small nuclear ribonucleoprotein) to a pre-mRNA with an intron-like region, the outron, containing an unpaired 39 splice site located near the 59 end. The second distinguishing feature is that many genes are transcribed in polycistronic units, known as operons, where a single promoter serves several genes. The operons can be up to eight genes long, and the polycistronic pre-mRNAs are separated into individual cistrons by 39 end formation accompanied by SL trans-splicing.These two features have important implications for C. elegans research. For instance, deletions/insertions within an operon may affect not only the expression of the gene containing the mutation, but also the genes downstream from it in the operon (Cui et al. 2008). Similarly, in a strain with enhanced RNAi sensitivity, RNAi of an operon gene can also affect expression of genes downstream (Guang et al. 2010). Finally, the trans-splice site is at the 59 end of the mRNA, not the pre-mRNA, and thus, the promoter is often not directly adjacent to the gene, but rather upstream of the outron or the entire operon (Blumenthal and Spieth 1996). SL trans-splicing has been repor...

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Evidence for Trans splicing in trypanosomes

Cited by 456 publications

References 47 publications

α‐ and β‐tubulin mRNAs of Trypanosoma cruzi originate from a single multicistronic transcript

α‐ and β‐tubulin mRNAs of Trypanosoma cruzi originate from a single multicistronic transcript

Peroxisomal Targeting Signal-1 Receptor Protein PEX5 fromLeishmania donovani

A global analysis of C. elegans trans-splicing

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