The L1Tc C-Terminal Domain from Trypanosoma cruzi Non-Long Terminal Repeat Retrotransposon Codes for a Protein That Bears Two C₂H₂ Zinc Finger Motifs and Is Endowed with Nucleic Acid Chaperone Activity

Abstract

L1Tc, a non-long terminal repeat retrotransposon from Trypanosoma cruzi, is a 4.9-kb actively transcribed element which contains a single open reading frame coding for the machinery necessary for its autonomous retrotransposition. In this paper, we analyze the protein encoded by the L1Tc 3 region, termed C2-L1Tc, which contains two zinc finger motifs similar to those present in the TFIIIA transcription factor family. C2-L1Tc binds nucleic acids with different affinities, such that RNA > tRNA > single-stranded DNA > double-stranded DNA, without any evidence for sequence specificity. C2-L1Tc also exhibits nucleic acid chaperone activity on different DNA templates that may participate in the mechanism of retrotransposition of the element. C2-L1Tc promotes annealing of complementary oligonucleotides, prevents melting of perfect DNA duplexes, and facilitates the strand exchange between DNAs to form the most stable duplex DNA in competitive displacement assays. Mapping of regions of C2-L1Tc using specific peptides showed that nucleic acid chaperone activity required a short basic sequence accompanied by a zinc finger motif or by another basic region such as RRR. Thus, a short basic polypeptide containing the two C 2 H 2 motifs promotes formation of the most stable duplex DNA at a concentration only three times higher than that required for C2-L1Tc.

The diversity of retrotransposons and the properties of their reverse transcriptases

“…L1Tc represents a distinct lineage of non-LTR retrotransposons that is somewhat unusual in that the element does not contain a first gag-like ORF typical of most non-LTR elements with APE domains. However the C-terminal region of L1Tc's single ORF has been shown to contain nucleic acid chaperone activity similar to the L1 ORF1 (Heras et al 2005). This chaperone activity has been suggested to be involved in the TPRT reaction because it can promote the annealing of complimentary oligonucleotides and can facility strand exchange between DNAs to form the most stable duplexes.…”

L1Tc non-LTR retrotransposons from Trypanosoma cruzi contain a functional viral-like self-cleaving 2A sequence in frame with the active proteins they encode

“…In addition, the recombinant RNaseHL1Tc exhibited RNaseH activity on different types of substrates and is active in a wide spectrum of pH and temperature [8]. Recently, it has been also reported that the protein encoded by the C-terminal end of L1Tc (C2-L1Tc) has in vitro nucleic acid chaperone activity and binds several types of nucleic acids with different affinity [9]. Most of these protein active domains are shared by L1Tc and LINE L1 from several mammals; however, significant divergences exist at nucleotide level between these elements, so that it has been conducted to include them in different clades [3].…”

“…The chemotherapy to treat the disease is highly toxic, and has low efficacy during chronic phase. L1Tc codes for proteins with endonuclease, RT and RNaseH activities, and for a protein with nucleic acid chaperone activity [6][7][8][9]. Some L1Tc elements comprise a single ORF, while others encode the different domains in more than one ORF [10].…”

The L1Tc non-LTR retrotransposon of Trypanosoma cruzi contains an internal RNA-pol II-dependent promoter that strongly activates gene transcription and generates unspliced transcripts

“…L1Tc has been reported as being associated with a gene coding for a transporter protein belonging to the ABC family (15), as being integrated into the coding sequence of the DNAj gene endowed with chaperone activity (16), as present in the expressed RHS multigene family located in a subtelomeric region, and as associated with SINE-like sequences (17–19). It codes for all the enzyme machinery involved in its retrotransposition, including AP endonuclease (20), 3′ phosphatase, 3′ phosphodiesterase (21), reverse transcriptase (22), RNAse H (23) and a nucleic acid chaperone (24). …”