Identification of an hepatitis delta virus-like ribozyme at the mRNA 5′-end of the L1Tc retrotransposon from Trypanosoma cruzi

Abstract

L1Tc is a non-LTR LINE element from Trypanosoma cruzi that encodes its transposition machinery and bears an internal promoter. Herewith, we report the identification of an in vitro active hepatitis delta virus-like ribozyme located in the first 77 nt at the 5′-end of the L1Tc mRNA (L1TcRz). The data presented show that L1TcRz has a co-transcriptional function. Using gel-purified uncleaved RNA transcripts, the data presented indicate that the kinetics of the self-cleaving, in a magnesium-dependent reaction, fits to a two-phase decay curve. The cleavage point identified by primer extension takes place at +1 position of the element. The hydroxyl nature of the 5′-end of the 3′-fragment generated by the cleavage activity of L1TcRz was confirmed. Since we have previously described that the 77-nt long fragment located at the 5′-end of L1Tc has promoter activity, the existence of a ribozyme in L1Tc makes this element to be the first described non-LTR retroelement that has an internal promoter–ribozyme dual function. The L1Tc nucleotides located downstream of the ribozyme catalytic motif appear to inhibit its activity. This inhibition may be influenced by the existence of a specific L1Tc RNA conformation that is recognized by RNase P.

Evolution of the R2 Retrotransposon Ribozyme and Its Self-Cleavage Site

“…Cleavage at the R2 5' junction coincides with the cleavage site associated with the ribozymes from other non-LTR retrotransposon clades- R4 (R4 [12]; Dong , Figure 6A), L1 [13], RTE [12]. In this report, analyses of the self-cleavage activity of diverse R2 elements indicated that in most species cleavage is within the 28S rRNA gene from 9 to 36 nt upstream of the R2 5' junction.…”

“…Some elements have been shown to encode a promoter, which initiates RNA synthesis at the precise 5' end of each element, while other elements are suggested to rely on co-transcription with cellular genes. Strong support for the latter was obtained when catalytically active HDV-like ribozymes were found in many non-LTR elements [11,12,13]. …”

Processing and Translation Initiation of Non-long Terminal Repeat Retrotransposons by Hepatitis Delta Virus (HDV)-like Self-cleaving Ribozymes

“…Recently, the first suggestion that 5Ј processing is performed by a ribozyme encoded in the 5Ј UTR of the retrotransposon was proposed when an HDV-like ribozyme was discovered at the 5Ј end of an Anopheles gambiae RTE (5). Subsequent analysis of the Drosophila R2 and Trypanosoma cruzi L1 elements showed that they also harbor HDVlike ribozyme at their 5Ј termini (6,7).…”

The wide expansion of hepatitis delta virus-like ribozymes throughout trypanosomatid genomes is linked to the spreading of L1Tc/ingi clade mobile elements

“…In addition, two in-tandem Pr77-hallmarks were found in T. brucei SIDER2 ( Tb SIDER2(126)Rz, Figure 2 and Additional file 1: Table S1) showing a hybrid folding of ribozymes L1 Tc and R2 [23, 25]. No SIDER1 signature was found, either in T. brucei or T. congolense that fitted any folding compatible with an HDV-like ribozyme.…”