The trypanosomatid Leptomonas collosoma 7SL RNA gene. Analysis of elements controlling its expression
The trypanosomatid Leptomonas collosoma 7SL RNA gene. Analysis of elements controlling its expressionHerzel Ben-Shlomo, Alexander Levitan, Oded Béjà and Shulamit Michaeli*
Department of Biological Chemistry, The Weizmann Institute of Science, Rehovot 76100, Israel
Received August 26, 1997Revised and Accepted November 3, 1997
We have previously reported the co-purification of a tRNA-like molecule with the Trypanosoma brucei SRP complex [Béjà et al. (1993) Mol. Biochem. Parasitol. 57, 223-230]. To examine whether the trypanosome SRP has a unique composition compared with that of other eukaryotes, we analyzed the 7SL RNA and the SRP complex of the monogenetic trypanosomatid Leptomonas collosoma. The 7SL RNA from L.collosoma was cloned, and its gene was sequenced. In contrast to T.brucei, two 7SL RNA transcripts were detected in L.collosoma that originate from a single-copy gene. Using stable cell lines expressing tagged 7SL RNA, we demonstrate that the tRNAArg gene located 98 bp upstream to the 7SL RNA serves as part of the 7SL RNA extragenic promoter. The steady-state level of 7SL RNA was found to be tightly regulated, since the presence of the gene on the multi-copy plasmid repressed the synthesis of the chromosomal gene. Cell lines carrying truncated 7SL RNA genes were established and their expression indicated that domain I is essential for expressing the 7SL RNA. No constructs carrying portions of the 7SL RNA were expressed, except for a construct which lacked 23 nt from the 3' end of the RNA. This suggests that 90% of the 7SL RNA molecule is important for its maintenance as a stable small RNA. We propose that the repression phenomenon may originate from a regulatory mechanism that coordinates the level of the 7SL RNA by its binding proteins.
The signal recognition particle (SRP) in eukaryotes is a cytoplasmic ribonucleoprotein that targets presecretory proteins and membrane proteins to the endoplasmic reticulum (ER) membrane (1 ). SRP was shown in vitro to bind the signal sequence emerging from ribosomes and to trigger a transient pause in elongation. This blockage is relieved when SRP interacts with the ER SRP receptor, and normal protein synthesis is resumed, resulting in co-translational translocation of the protein into the ER lumen (1 ). The eukaryotic SRP carries a single RNA molecule, the 7SL RNA, that is composed of four stem-loop structures and six SRP proteins (2 ). Different functions were assigned to the different proteins. It was shown that SRP54 binds the signal peptide as it emerges from the ribosome, SRP 9/14 bind to domain I and function in elongation arrest, SRP68/72 mediate translocation into the ER, whereas SRP19 facilitates the binding of SRP54 to the RNA (3 ).
Extensive phylogenetic studies were performed on SRP RNAs and indicate that the different RNAs vary in size and in composition but all carry an invariant domain IV (4 ,5 ). It was proposed that the ancestral SRP molecule was reduced in size during bacterial evolution (4 ). However, it is currently unknown whether, in organisms carrying truncated forms of the SRP RNA, the missing RNA domains reside in other yet unidentified small RNAs. Extensive changes in the size and shape of domain I are found among eukaryotic 7SL RNAs. For example, domain I of higher eukaryotic 7SL RNAs can be folded as a tRNA-like molecule, but those of Trypanosoma brucei (6 ) and Tetrahymena lack one of the arms of the tRNA-like structure (7 ) and yeast have a significantly truncated form that bears no resemblance to the tRNA structure (8 ).
Structure-function aspects of the SRP were addressed in vitro in the canine system and in vivo in the fission yeast Schizosaccharomyces pombe. The results obtained from both systems indicate that the four domains (including domain III, that is missing from the bacterial SRPs) are critical for SRP function (9 ,10 ). It is currently unknown whether domain I of yeast also contributes to protein arrest, since the phenotype observed for domain I mutants may result from a failure to transcribe the gene (10 ).
Little is known about the regulation of 7SL RNA transcription. The only well characterized 7SL RNA promoter is the human promoter which is comprised of both upstream and internal elements (11 ,12 ). Homologies to A and B boxes of tRNA promoters can be found within the coding region of 7SL RNA. However, a CG dinucleotide at position +15 and +16 located outside this box was found to be essential for transcription whereas most of the mutations made within the A box had little effect (13 ). Analysis of the T.brucei 7SL RNA gene demonstrated the existence of extragenic elements located 97 bp upstream to the start site that controls the synthesis of the 7SL RNA. This element resides within a tRNALys that is transcribed in the opposite direction (14 ).
Very little is known about protein translocation in trypanosomes. The finding that T.brucei has an SRP homologue was not surprising (6 ). However, the presence of a co-migrating tRNA-like molecule, that co-purifies with the T.brucei 7SL RNA (15 ), led us to hypothesize that the trypanosomatid SRP may differ from other SRPs, and is composed of two small RNP particles.
In this study, we have cloned, sequenced and expressed mutated and truncated versions of the monogenetic trypanosomatid Leptomonas collosoma 7SL RNA. This study is the first step towards understanding the structure-function relationship of the trypanosomatid SRP and the mechanism that tightly regulates its level of expression. The results indicate that L.collosoma 7SL RNA has a unique property as it is the only 7SL RNA described so far that is present in two stable RNA conformations. Studies performed with stable cell lines expressing truncated and mutated 7SL RNA demonstrate that the tRNAArg located upstream to the 7SL RNA gene is part of the extragenic promoter element and that an intragenic element controlling the expression of the gene exists in domain I. This study also indicates that the steady-state level of 7SL RNA in trypanosomes is tightly regulated since the presence of mutated 7SL RNA on a multi-copy plasmid repressed the synthesis of the wild-type RNA. The repression was observed only when all 7SL RNA domains (known as the SRP protein binding sites) were present, suggesting a mechanism that coordinates the level of the 7SL RNA by its binding proteins.
Total RNA was prepared with TRIzol reagent (GIBCO BRL). The RNA samples were fractionated on a 7 M urea/10% polyacrylamide gel and electroblotted onto a Nytran membrane. Hybridization with labelled oligonucleotides was performed at 37°C in 5× SSC (1× SSC consisted of 150 mM NaCl and 15 mM sodium citrate), 0.1% SDS, 5× Denhardt's solution and 100 µg/ml salmon sperm DNA. The L.collosoma 7SL RNA probe was obtained from excising the two 280 and 300 nt 7SL RNA molecules enriched in PRS from a preparative denaturing gel. The RNA was treated with alkaline phosphatase (16 ) to remove the phosphate termini and was 5' end labeled with polynucleotide kinase (16 ).
Approximately 20 000 plaques of an L.collosoma [lambda]EMBL3 were screened with 5' end-labeled 7SL RNA and four independent clones were isolated that also hybridized with antisense oligonucleotide complementary to nt 165-184 of the T.brucei 7SL RNA (6 ). The clones were digested with SalI and a 2.8 kb fragment was subcloned into the pGEM-3 vector, and the L.collosoma 7SL 4/78 construct was generated. Two fragments, a 400 bp Sau3A fragment and a 1 kb TaqI fragment, were subcloned and sequenced using SP6 and T7 primers and internal gene primers.
Leptomonas collosoma cells were grown as previously described (17 ). Cells (5 × 1010) were harvested, washed with PBS and resuspended in Buffer A containing 35 mM HEPES-KOH (pH 7.9), 10 mM MgCl2, 50 mM KCl, 5 µg/ml leupeptin and 5 mM [beta]-mercaptoethanol. The cell suspension was equilibrated in a nitrogen cavitation bomb (Parr Instruments Company) at 1000 psi for 10 min, and disrupted by release from the bomb. Post-ribosomal supernatant (PRS) preparation and DEAE-chromatography were essentially as described previously (18 ).
About 4 × 107 cells were used for electroporation, conducted with two pulses of 500 µF and 2.5 kV/cm in a Bio-Rad gene pulser. Cells were transfected with 50-100 µg of the different DNA constructs. Transformants were selected in liquid medium in the presence of 20-50 µg/ml G418 (GIBCO). To elevate the copy number of the plasmid the transformants were grown in the presence of elevated G418 (500-1000 µg/ml).
To construct the 7SL RNA gene tagged in the SacII site a linker (oligo 10021) was inserted into the unique SacII site in L.collosoma 7SL RNA 4/78, located in position 239 within domain II of the 7SL RNA. The fragment carrying the mutation was subcloned into the pX expression vector (19 ). Two deletions of the upstream region were constructed using the 1460 bp AccI-SalI and the 1360 bp TaqI-SalI fragments. To generate a pX plasmid containing the 7SL RNA gene transcription termination signals, (pX-7SL-t), a 1.2 kb HindIII-SmaI fragment, derived from the 4/78 plasmid, was cloned to the pX vector. The constructs described below are illustrated in Figure 5 A. The construct (dI) carrying domain I was obtained by ligating to pX a SmaI-PvuII containing a 1.3 kb upstream sequence and 41 nt of the 7SL RNA. The construct [p(dI)t] was made by ligating the same fragment as for [p(dI)] but to the pX-7SL-t. Constructs p(H)t and p(B)t were made by ligating the SmaI-HincII fragment carrying 86 nt of the 7SL RNA or SmaI-BssHI carrying 147 nt of the 7SL RNA, respectively, to pX-7SL-t. Plasmid pdIX2 was constructed by cloning a PCR product in to the BamHI site obtained using oligos 8721 and 15137 into the pdI plasmid. Plasmid [p(-dII)t] was constructed by cloning a PCR product, obtained from oligos 18816 and 15366, to pX-7SL-t. The p(-3')t plasmid was constructed by ligating the PCR product generated with oligos 18662 and 15366 to the SmaI site. The [p(-dI)t] construct was made by three point-ligation. The PCR product obtained using oligos 15367 and 15366 was digested with XbaI and PvuII; the second PCR product, made with oligos 8721 and 15137, was digested with PvuII and BamHI. The two PCR products were ligated to pX digested with BamHI and XbaI. The loop IV mutation was obtained by the Kunkel method (20 ). To generate the site-directed mutation, the 2.8 kb SalI fragment was cloned into pBluescript KS(-). Oligonucleotide 12615 was used to generate the mutation, and the mutated fragment was cloned to the pX vector. The mutation was confirmed by sequencing. The sequence of the constructs generated by PCR and site-directed mutagenesis was verified. To synthesize an antisense 7SL RNA probe, a PCR product carrying the entire 7SL RNA gene (generated with oligos 8721 and 15137) was cloned into the BamHI site of pGEM-3.
Antisense 7SL RNA was synthesized using SP6 polymerase. Total RNA (~10 µg) was mixed with 100 000 c.p.m. of in vitro transcribed anti-7SL RNA probe in buffer containing 80% formamide, 40 mM PIPES (pH 6.4), 0.4 M sodium acetate and 1 mM EDTA. After incubation at 85°C for 5 min, the RNA was hybridized for 12 h at 45°C. The unprotected RNA was digested with RNase One (Promega), and after deproteinization, the RNA was separated on a 6% denaturing gel.
As a first step towards elucidating the SRP complex of L.collosoma we identified its 7SL RNA. Whole cell extracts were prepared from L.collosoma and the 7SL RNA content was examined in these extracts after depletion of ribosomes (PRS). The RNA profile and the hybridization results with an antisense oligonucleotide, complementary to the T.brucei 7SL RNA (oligo EU-53) probe, are presented in Figure 1 A, II. The results indicate the existence of two 7SL RNA molecules of 280 and 300 nt that hybridize to the 7SL RNA probe. These two molecules are present in a 1:1 ratio and are highly enriched in PRS. To examine whether the two molecules are distinct stable transcripts, the potential for interconversion of one species to the other was investigated. A fraction enriched with 7SL RNA was end labelled at the 5' end and the labeled RNA (Fig. 1 B, lane 1) was separated on a 10% denaturing gel. The separated 7SL I and 7SL II molecules were excised and analyzed. The results (Fig. 1 B) suggest that there is no conversion in vitro between these species. Further separation of the 7SL RNA in different gel systems suggests that the 7SL RNA migrates as a single species only in the presence of 75% formamide and 7 M urea (Fig. 1 C, panel 2), suggesting that only under severe denaturation conditions can the molecule be completely denatured. These results also suggest that the two distinct 7SL RNA conformations are stable and are generated in vivo.
To study the structure-function of the 7SL RNA, we sought to overexpress mutated and truncated 7SL RNA genes. The 7SL RNA gene was marked by inserting a linker in the unique SacII site (construct I in Fig. 5 A) located at position 239 of the 7SL RNA (domain II), indicated in Figure 2 B. A stable cell line expressing the mutated gene was obtained. The expression of the tagged RNA was examined by northern analysis, and the results indicate that the tagged 7SL RNA, which is larger than the wild-type RNA is efficiently expressed (Fig. 4 B, lane 2). The expression of the tagged 7SL RNA repressed the synthesis of the wild-type RNA, since RNA transcripts, corresponding in size to wild-type 7SL RNA, were absent in cell lines carrying the tagged molecule (compare lanes 1 and 2). Since only a single RNA species was observed in lane 2, it may suggest that the tagged 7SL RNA does not undergo the conformational change proposed for the wild-type RNA.
To characterize the sequences that regulate the expression of the 7SL RNA and to elucidate the mechanism that elicits the repression of the wild-type 7SL RNA by the gene carried on the multi-copy plasmid, several constructs were generated and are schematically presented in Figure 5 A.
One possibility to explain the repression phenomenon is that the multi-copy plasmid titrates the 7SL RNA transcription factors. To examine this possibility the level of 7SL RNA was examined in cell lines carrying the upstream regulatory region and domain I, since this domain was shown in humans to carry intragenic promoter elements (11 ). The results presented in Figure 5 B (lanes 1, 2 and 3) indicate that the level of the cellular 7SL RNA was not changed in cells carrying constructs p(dI) and p(dI)t. A small transcript of 40 nt that could have been generated from domain I was not detected in total RNA preparation, even when the proper termination signals from the 7SL RNA gene were present [construct p(dI)t]. This data may suggest that the repression is not due to competition for transcription factors.
Hybridization of the same northern with the tRNAArg probe (Fig. 5 B) indicates that the level of tRNAArg was elevated due to its presence on the multi-copy plasmid, suggesting that the tRNAArg carried on these plasmids is highly expressed, irrespective of the expression of 7SL RNA.
To examine the factors that regulate the expression of the 7SL RNA, two constructs that differ only in domain I, i.e., deletion of domain I [p(-dI)t] or duplication of this domain (pdIx2) were constructed, and the expression of the 7SL RNA was examined. The results indicate that in the absence of domain I the 7SL RNA is not expressed, (Fig. 5 B lane 9), suggesting that domain I may carry elements which are necessary for transcribing the gene, as in humans. Duplication in domain I did not interfere with the expression of the gene (lane 8), but this construct failed to repress the synthesis of the wild-type RNA.
To further explore the sequences essential for expressing the 7SL RNA, constructs harboring different lengths of the 7SL RNA were generated. Four constructs carrying split 7SL RNA genes harboring coding information for 86, 147, 216 and 257 nt were generated, and are schematically presented in Figure 5 A. All these constructs carried the 7SL RNA transcription termination signals. The constructs were used to establish stable cell lines and the level of 7SL RNA was examined. The results, presented in Figure 5 B, lanes 4-6, indicate that, despite the high level of expression of the accompanying tRNAArg, none of these truncated 7SL RNA were detectable in steady-state RNA preparation, suggesting that all four domains are essential for expressing stable 7SL RNA molecules.
To examine the level of 7SL RNA that may be present in minute amounts, a sensitive RNase protection assay was used. The results presented in Figure 5 C indicate that, except for a construct lacking 23 nt from the 3' end (lane 7), none of the truncated 7SL RNAs were expressed. Longer exposure of the gel did not reveal any additional bands that were not found in the wild-type control. However, the construct lacking only 23 nt from the 3' end was efficiently expressed and repressed the synthesis of the wild-type RNA, as no fragment corresponding to protection with wild-type RNA was observed. The presence of two transcripts in Figure 5 B, lane 7, suggests that this truncated 7SL RNA is capable of undergoing the conformational change, like the wild-type RNA.
The tight regulation on the 7SL RNA was also observed when the wild-type 7SL RNA locus was cloned into the pX plasmid. Despite the elevation in the copy number of the gene no increase in the cellular level of 7SL RNA was observed (Fig. 5 B, lane 10).
In this study we have cloned and sequenced the 7SL RNA gene of the monogenetic trypanosomatid L.collosoma and examined the elements that regulate its expression. The results indicate that the level of 7SL RNA is tightly regulated, since synthesis of mutated 7SL RNA repressed the wild-type 7SL RNA. The repression took place only when the 7SL RNA, carried on the plasmid, was expressed. Apart from the 7SL RNA gene lacking only 23 nt from the 3' end, none of the other split 7SL RNAs were expressed, suggesting that only when four 7SL RNA domains are present can the RNA be maintained as a stable small RNA. This study also demonstrates that the tRNAArg is part of the extragenic elements that control the transcription of 7SL RNA and that domain I, like in humans, is essential for expressing the gene.
One of the most intriguing findings regarding the trypanosome 7SL RNA is its high degree of similarity to the human RNA (60%) compared with the yeast S.pombe RNA (45%) (6 ). This is also true for the L.collosoma RNA, that shares 52% identity with the human RNA and only 38% with the S.pombe RNA. This result is even more surprising considering the earlier divergence of trypanosomes compared with yeast from the eukaryotic lineage (24 ). Genetic transfer from the mammalian host to the parasite, is a mechanism that could have generated such relatedness. However, since L.collosoma lacks a mammalian host, the genetic transfer hypothesis should be ruled out.
Despite the sequence diversity between the yeast, human and trypanosome RNAs, yeast and trypanosome RNAs possess a truncated domain I compared with the tRNA-like domain of human RNA. The trypanosome domain I is an intermediate between the highly-truncated yeast domain and the tRNA-like structure of the human domain. In the mammalian domain I, a potential for base-pairing between the two hairpin loops exists. This potential does not exist in the trypanosome or Tetrahymena RNAs, since these RNAs carry a truncated second hairpin loop. Interestingly, however, these three domains carry the sequence homologous to the consensus sequence 5'-GCG-N3-5-CCUGUAAYCY-3' that has been involved in binding the SRP9 and 14 (25 ). Based on the truncation of domain I in lower eukaryotes and the presence of a tRNA-like molecule that co-purifies with the trypanosome 7SL RNP (15 ) we propose that in those organisms which lack domain I, (like bacteria or eukaryotes that have a truncated domain I), a functional domain I may be carried by a separate RNP complex. We have previously reported the co-purification of a tRNA-like molecule with the T.brucei 7SL RNP (15 ) and recently identified the tRNA-like molecule that co-purifies with the L.collosoma 7SL RNP (ourunpublished results).
The results suggest that the L.collosoma 7SL RNA is a single copy gene and that no size variation exists between the two 7SL RNA alleles. The data also indicate that the two 7SL RNA transcripts represent two stable conformers of the RNA. We observed that the ratio between 7SL I and II changes during growth, i.e., actively growing cells contain more 7SL I, and the ratio between the molecules reaches 1:1 when the culture ages. In addition, 7SL I is preferentially associated with ribosomes, and the in vivo conversion of 7SL I to II is inhibited in the presence of cycloheximide. These data suggest that the conformational change takes place during protein translocation (Ben-Shlomo et al., unpublished data). The finding that 7SL RNA mutants located in domain II and IV exist in a single conformation may indicate that these mutated RNAs are not active in protein translocation and therefore do not undergo the conformational change.
The repression phenomenon observed in this study is similar to the observation made when human U1 RNA gene was expressed in mouse cells (26 ). It was found that despite the efficient expression of the human gene, the total amount of U1 RNA (both mouse and human) did not change, suggesting that multi-gene families encoding mammalian U1 RNA are subjected to dosage compensation. The finding of such a regulatory phenomenon shared by two different small RNPs, may suggest that there is a common regulatory mechanism that co-regulates the level of the RNA with its RNP binding proteins. The repression was observed only when the 7SL RNA was efficiently transcribed and maintained as a stable RNA. We, therefore, favor the hypothesis that the SRP proteins regulate the level of 7SL RNA. Although we cannot rule out the possibility that the repression is due to the titration of a factor that binds to the coding region of the 7SL RNA, we consider this possibility most unlikely. From all the truncated 7SL RNA only the construct missing 23 nt from the 3' end was expressed. This may suggest that the correct folding of the 7SL RNA into its protein binding domains is what dictates whether a truncated 7SL RNA will be maintained as a stable 7SL RNA. Support for a cellular mechanism that links and coordinates between the 7SL RNA and the SRP protein was obtained from the work in yeast (27 ) which indicated that cells impaired in the synthesis of SRP proteins showed a large reduction in the level of the SRP RNA.
Using transient transfection assays in T.brucei, it was previously demonstrated that the extragenic regulatory elements that control the synthesis of 7SL, U6 and U3 RNAs are found in the A and B boxes of the respective companion tRNA (14 ). The data presented in this study are consistent with this conclusion, since truncating the tRNAArg abolished synthesis of the tagged 7SL RNA. The 10- to 20-fold amplification of the tRNAArg compared to its level in wild-type cells supports the notion that no competition between factors that regulate the 7SL and tRNA gene occurs, as the same level of tRNA amplification was observed, regardless of how efficient the 7SL RNA gene was expressed. It is currently unknown how the companion tRNA elicits its regulatory effect on the 7SL RNA.
Several models were suggested to explain the transcriptional linkage between the small RNA and its companion tRNA gene. One model suggests that the tRNA locus might modify the chromatin structure at the 7SL RNA gene and render the region accessible for binding transcription factors. The second possibility is that the transcription factor TFIIB, that normally binds to the region located 45-50 nt upstream to the tRNA gene, may interact with a 7SL RNA transcription factor that is placed over the transcription start site of the 7SL RNA gene. Alternatively, the A and B boxes of the companion tRNA may have a dual function, they may serve as promoter elements for the tRNA itself while controlling the expression of 7SL RNA. However, it is difficult to rationalize this possibility, because the binding of transcription factors to tRNA genes occurs in an orientation-dependent manner, whereas the tRNA and the 7SL RNA are transcribed divergently (28 ). Our finding that only the level of the tRNAArg is greatly amplified in cell lines carrying the two genes on a multi-copy plasmid may suggest that the transcription rate in the two directions is different, making the latter possibility unlikely. However, we cannot rule out the possibility that the cellular level of 7SL RNA is mostly regulated post-transcriptionally.
The repression phenomenon observed in this study can be further utilized to examine the structure-function relationship of the trypanosomatid 7SL RNA. Since the level of wild-type 7SL RNA was undetectable in cells carrying 7SL RNA mutated in domains II and IV, it may suggest that trypanosomes, like yeast (29 ), utilize an alternative protein translocation pathway.
This work was supported by research grant from the Cemach and Anna Oiserman Research fund and from the Leo and Julia Forchheimer Center for Molecular Genetics of the Weizmann Institute. We wish to thank Ora Asher for excellent technical assistance.
15 Béjà,O., Ullu,E. and Michaeli,S. (1993) Mol. Biochem. Parasitol.,57, 223-230.
16 Maniatis,T., Fritsch,E.F. and Sambrook,J. (1982) Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY.
17 Goldring,A., Karchi,M. and Michaeli,S. (1995) Exp. Parasitol., 80, 333-338.
18 Michaeli,S., Roberts,T.G., Watkins,K.P. and Agabian,N. (1990) J. Biol. Chem., 265, 10582-10588.MEDLINE Abstract
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