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1.
Over 50 introns have been reported in archaeal rRNA genes (rDNAs), a subset of which nests putative homing endonuclease (HEase) genes. Here, we report the identification and characterization of a novel archaeal LAGLIDADG-type HEase, I-ApeKI [corrected], encoded by the ApeK1.S908 intron within the 16S rDNA of Aeropyrum pernix K1. I-ApeKI [corrected] consists of 222 amino acids and harbors two LAGLIDADG-like sequences. It recognizes the 20 bp non-palindromic sequence 5'-GCAAGGCTGAAAC downward arrowTTAAAGG and cleaves target DNA to produce protruding tetranucleotide 3' ends. Either Mn2+ or Co2+ can be substituted for Mg2+ as a cofactor in the cleavage reaction. Of the 20 bases within the minimal recognition site, 7 are essential for cleavage and are located at positions proximal to the cleavage sites.  相似文献   

2.
Group I introns are autonomous genetic elements that can catalyze their own excision from pre-RNA. Understanding how group I introns move in nuclear ribosomal (r)DNA remains an important question in evolutionary biology. Two models are invoked to explain group I intron movement. The first is termed homing and results from the action of an intron-encoded homing endonuclease that recognizes and cleaves an intronless allele at or near the intron insertion site. Alternatively, introns can be inserted into RNA through reverse splicing. Here, we present the sequences of two large group I introns from fungal nuclear rDNA, which both encode putative full-length homing endonuclease genes (HEGs). Five remnant HEGs in different fungal species are also reported. This brings the total number of known nuclear HEGs from 15 to 22. We determined the phylogeny of all known nuclear HEGs and their associated introns. We found evidence for intron-independent HEG invasion into both homologous and heterologous introns in often distantly related lineages, as well as the "switching" of HEGs between different intron peripheral loops and between sense and antisense strands of intron DNA. These results suggest that nuclear HEGs are frequently mobilized. HEG invasion appears, however, to be limited to existing introns in the same or neighboring sites. To study the intron-HEG relationship in more detail, the S943 group I intron in fungal small-subunit rDNA was used as a model system. The S943 HEG is shown to be widely distributed as functional, inactivated, or remnant ORFs in S943 introns.  相似文献   

3.
Busse I  Preisfeld A 《Protist》2003,154(1):57-69
The gene coding for the small ribosomal subunit RNA of Ploeotia costata contains an actively splicing group I intron (Pco.S516) which is unique among euglenozoans. Secondary structure predictions indicate that paired segments P1-P10 as well as several conserved elements typical of group I introns and of subclass IC1 in particular are present. Phylogenetic analyses of SSU rDNA sequences demonstrate a well-supported placement of Ploeotia costata within the Euglenozoa; whereas, analyses of intron data sets uncover a close phylogenetic relation of Pco.S516 to S-516 introns from Acanthamoeba, Aureoumbra lagunensis (Stramenopila) and red algae of the order Bangiales. Discrepancies between SSU rDNA and intron phylogenies suggest horizontal spread of the group I intron. Monophyly of IC1 516 introns from Ploeotia costata, A. lagunensis and rhodophytes is supported by a unique secondary structure element: helix P5b possesses an insertion of 19 nt length with a highly conserved tetraloop which is supposed to take part in tertiary interactions. Neither functional nor degenerated ORFs coding for homing endonucleases can be identified in Pco.S516. Nevertheless, degenerated ORFs with His-Cys box motifs in closely related intron sequences indicate that homing may have occurred during evolution of the investigated intron group.  相似文献   

4.
Group I introns were discovered inserted at the same position in the nuclear small-subunit ribosomal DNA (nuc-ssu-rDNA) in several species of homobasidiomycetes (mushroom-forming fungi). Based on conserved intron sequences, a pair of intron-specific primers was designed for PCR amplification and sequencing of intron-containing rDNA repeats. Using the intron-specific primers together with flanking rDNA primers, a PCR assay was conducted to determine presence or absence of introns in 39 species of homobasidiomycetes. Introns were confined to the genera Panellus, Clavicorona, and Lentinellus. Phylogenetic analyses of nuc-ssu-rDNA and mitochondrial ssu-rDNA sequences suggest that Clavicorona and Lentinellus are closely related, but that Panellus is not closely related to these. The simplest explanation for the distribution of the introns is that they have been twice independently gained via horizontal transmission, once on the lineage leading to Panellus, and once on the lineage leading to Lentinellus and Clavicorona. BLAST searches using the introns from Panellus and Lentinellus as query sequences retrieved 16 other similar group I introns of nuc-ssu-rDNA and nuclear large-subunit rDNA (nuc-lsu-rDNA) from fungal and green algal hosts. Phylogenetic analyses of intron sequences suggest that the mushroom introns are monophyletic, and are nested within a clade that contains four other introns that insert at the same position as the mushroom introns, two from different groups of fungi and two from green algae. The distribution of host lineages and insertion sites among the introns suggests that horizontal and vertical transmission, homing, and transposition have been factors in intron evolution. As distinctive, heritable features of nuclear rDNAs in certain lineages, group I introns have promise as phylogenetic markers. Nevertheless, the possibility of horizontal transmission and homing also suggest that their use poses certain pitfalls.   相似文献   

5.
Although the examination of large subunit ribosomal RNA genes (LSU rDNA) is advanced in phylogenetic studies, no corresponding sequence data from trebouxiophytes have been published, with the exception of ‘Chlorellaellipsoidea Gerneck. We determined the LSU rDNA sequence of Chlorella vulgaris Beijerinck and of the symbiotic alga of green paramecium, Chlorella sp. NC64A. A total of 59 nucleotide substitutions were found in the LSU rDNA of the two species, which are disproportionately distributed. Primarily, 65% of the substitutions were encountered in the first 800 bp of the alignment. This segment apparently has evolved eight times faster than the complete SSU rDNA sequence, making it a good candidate for a phylogenetic marker and giving a resolution level intermediate between small subunit (SSU) rDNA and internal transcribed spacers. Green algae are known as a group I intron‐rich group along with rhodophytes and fungi. NC64A is particularly rich in the introns; five introns were newly identified from the LSU rDNA sequence, which we named Cnc.L200, Cnc.L1688, Cnc.L1926, Cnc.L2184 and Cnc.L2437, following the insertion positions. In the present study we analyzed these introns with three others (Cnc.S943, Cnc.S1367 and Cnc.S1512) that had already been found in NC64A SSU rDNA. Secondary structure modeling placed these introns in the group I intron family, with four introns belonging to subgroup C1 and the other four introns belonging to subgroup E. Five of the intron insertion positions are unique to the paramecian symbiont, which may indicate relatively recent events of intron infections that includes transpositions. Intron phylogeny showed unprecedented relationships; four Cnc. IC1 introns made a clade with some green algal introns with insertions at nine different positions, whereas four Cnc. IE introns made a clade with the S651 intron (Chlorella sp. AN 1–3), which lay as a sister to the S516 insertion position subfamily.  相似文献   

6.
Group I introns are widespread in eukaryotic organelles and nuclear- encoded ribosomal DNAs (rDNAs). The green algae are particularly rich in rDNA group I introns. To better understand the origins and phylogenetic relationships of green algal nuclear-encoded small subunit rDNA group I introns, a secondary structure-based alignment was constructed with available intron sequences and 11 new subgroup ICI and three new subgroup IB3 intron sequences determined from members of the Trebouxiophyceae (common phycobiont components of lichen) and the Ulvophyceae. Phylogenetic analyses using a weighted maximum-parsimony method showed that most group I introns form distinct lineages defined by insertion sites within the SSU rDNA. The comparison of topologies defining the phylogenetic relationships of 12 members of the 1512 group I intron insertion site lineage (position relative to the E. coli SSU rDNA coding region) with that of the host cells (i.e., SSU rDNAs) that contain these introns provided insights into the possible origin, stability, loss, and lateral transfer of ICI group I introns. The phylogenetic data were consistent with a viral origin of the 1512 group I intron in the green algae. This intron appears to have originated, minimally, within the SSU rDNA of the common ancestor of the trebouxiophytes and has subsequently been vertically inherited within this algal lineage with loss of the intron in some taxa. The phylogenetic analyses also suggested that the 1512 intron was laterally transferred among later-diverging trebouxiophytes; these algal taxa may have coexisted in a developing lichen thallus, thus facilitating cell- to-cell contact and the lateral transfer. Comparison of available group I intron sequences from the nuclear-encoded SSU rDNA of phycobiont and mycobiont components of lichens demonstrated that these sequences have independent origins and are not the result of lateral transfer from one component to the other.   相似文献   

7.
The spread of LAGLIDADG homing endonuclease genes in rDNA   总被引:5,自引:0,他引:5       下载免费PDF全文
Group I introns that encode homing endonuclease genes (HEGs) are highly invasive genetic elements. Their movement into a homologous position in an intron-less allele is termed homing. Although the mechanism of homing is well understood, the evolutionary relationship between HEGs and their intron partners remains unclear. Here we have focused on the largest family of HEGs (encoding the protein motif, LAGLIDADG) to understand how HEGs and introns move in rDNA. Our analysis shows the phylogenetic clustering of HEGs that encode a single copy of the LAGLIDADG motif in neighboring, but often evolutionarily distantly related, group I introns. These endonucleases appear to have inserted into existing introns independent of ribozymes. In contrast, our data support a common evolutionary history for a large family of heterologous introns that encode HEGs with a duplicated LAGLIDADG motif. This finding suggests that intron/double-motif HEG elements can move into heterologous sites as a unit. Our data also suggest that a subset of the double-motif HEGs in rDNA originated from the duplication and fusion of a single-motif HEG encoded by present-day ribozymes in LSU rDNA.  相似文献   

8.
Although hypotheses have been proposed and developed to interpret the origins and functions of introns, substantial controversies remain about the mechanism of intron evolution. The availability of introns in the intermediate state is quite helpful for resolving this debate. In this study, a new strain of diatom (denominated as DB21‐1) was isolated and identified as Olifantiella sp., which possesses multiple types of 18S rDNAs (obtained from genomic DNA; lengths ranged from 2,056 bp to 2,988 bp). Based on alignments between 18S rDNAs and 18S rRNA (obtained from cDNA; 1,783 bp), seven intron insertion sites (IISs) located in the 18S rDNA were identified, each of which displayed the polymorphism of intron presence/absence. Specific primers around each IIS were designed to amplify the introns and the results indicated that introns in the same IIS varied in lengths, while terminal sequences were conserved. Our study showed that the process of intron loss happens via a series of successive steps, and each step could derive corresponding introns under intermediate states. Moreover, these results indicate that the mechanism of genomic deletion that occurs at DNA level can also lead to exact intron loss.  相似文献   

9.
The origins of fungal group I introns within nuclear small-subunit (nSSU) rDNA are enigmatic. This is partly because they have never been reported in basal fungal phyla (Zygomycota and Chytridiomycota), which are hypothesized to be ancestral to derived phyla (Ascomycota and Basidiomycota). Here we report group I introns from the nSSU rDNA of two zygomycete fungi, Zoophagus insidians (Zoopagales) and Coemansia mojavensis (Kickxellales). Secondary structure analyses predicted that both introns belong to the IC1 subgroup and that they are distantly related to each other, which is also suggested by different insertion sites. Molecular phylogenetic analyses indicated that the IC1 intron of Z. insidians is closely related to the IC1 intron inserted in the LSU rDNA of the basidiomycete fungus Clavicorona taxophila, which strongly suggests interphylum horizontal transfer. The IC1 intron of C. mojavensis has a low phylogenetic affinity to other fungal IC1 introns inserted into site 943 of nSSU rDNA (relative to E. coli 16S rDNA). It is noteworthy that this intron contains a putative ORF containing a His–Cys box motif in the antisense strand, a hallmark for nuclear-encoded homing endonucleases. Overall, molecular phylogenetic analyses do not support the placement of these two introns in basal fungal IC1 intron lineages. This result leads to the suggestion that fungal IC1 introns might have invaded or been transferred laterally after the divergence of the four major fungal phyla. Received: 8 February 2001 / Accepted: 1 November 2001  相似文献   

10.
The recent transfer of a homing endonuclease gene   总被引:1,自引:0,他引:1       下载免费PDF全文
The myxomycete Didymium iridis (isolate Panama 2) contains a mobile group I intron named Dir.S956-1 after position 956 in the nuclear small subunit (SSU) rRNA gene. The intron is efficiently spread through homing by the intron-encoded homing endonuclease I-DirI. Homing endonuclease genes (HEGs) usually spread with their associated introns as a unit, but infrequently also spread independent of introns (or inteins). Clear examples of HEG mobility are however sparse. Here, we provide evidence for the transfer of a HEG into a group I intron named Dir.S956-2 that is inserted into the SSU rDNA of the Costa Rica 8 isolate of D.iridis. Similarities between intron sequences that flank the HEG and rDNA sequences that flank the intron (the homing endonuclease recognition sequence) suggest that the HEG invaded the intron during the recent evolution in a homing-like event. Dir.S956-2 is inserted into the same SSU site as Dir.S956-1. Remarkably, the two group I introns encode distantly related splicing ribozymes with phylogenetically related HEGs inserted on the opposite strands of different peripheral loop regions. The HEGs are both interrupted by small spliceosomal introns that must be removed during RNA maturation.  相似文献   

11.
Studies of the distribution of the three group I introns (intron A, intron T, and intron AT) in the 26S rDNA of Gaeumannomyces graminis had suggested that they were transferred to a common ancestor of G. graminis var. avenae and var. tritici after it had branched off from var. graminis. Intron AT and intron A exhibited vertical inheritance and coevolved in concert with their hosts. Intron loss could occur after its acquisition. Loss of any one of the three introns could occur in var. tritici whereas only loss of intron T had been found in the majority of var. avenae isolates. The existence of isolates of var. tritici and var. avenae with three introns suggested that intron loss could be reversed by intron acquisition and that the whole process is a dynamic one. This process of intron acquisition and intron loss reached different equilibrium points for different varieties and subgroups, which explained the irregular distribution of these introns in G. graminis. Each of the three group I introns was more closely related to other intron sequences that share the same insertion point in the 26S rDNA than to each other. These introns in distantly related organisms appeared to have a common ancestry. This system had provided a good model for studies on both the lateral transfer and common ancestry of group I introns in the 26S rRNA genes. Received: 17 May 1996 / Accepted: 14 January 1997  相似文献   

12.
13.
14.
Self-splicing group I introns are being found in an increasing number of bacteriophages. Most introns contain an open reading frame coding for a homing endo-nuclease that confers mobility to both the intron and the homing endonuclease gene (HEG). The frequent occurrence of intron/HEG has raised questions whether group I introns are spread via horizontal transfer between phage populations. We have determined complete sequences for the known group I introns among T-even-like bacteriophages together with sequences of the intron-containing genes td, nrdB, and nrdD from phages with and without introns. A previously uncharacterized phage isolate, U5, is shown to contain all three introns, the only phage besides T4 found with a "full set" of these introns. Sequence analysis of td and nrdB genes from intron-containing and intronless phages provides evidence that recent horizontal transmission of introns has occurred among the phages. The fact that several of the HEGs have suffered deletions rendering them non-functional implies that the homing endonucleases are of no selective advantage to the phage and are rapidly degenerating and probably dependent upon frequent horizontal transmissions for maintenance within the phage populations. Several of the introns can home to closely related intronless phages during mixed infections. However, the efficiency of homing varies and is dependent on homology in regions flanking the intron insertion site. The occurrence of optional genes flanking the respective intron-containing gene can strongly affect the efficiency of homing. These findings give further insight into the mechanisms of propagation and evolution of group I introns among the T-even-like bacteriophages.  相似文献   

15.
The complete gene sequence and secondary structure of the mitochondrial LSU rRNA from the cultivated Basidiomycota Agrocybe aegerita was derived by chromosome walking. The A.aegerita LSU rRNA gene (13 526 nt) represents, to date, the longest described, due to the highest number of introns (eight) and the occurrence of six long nucleotidic extensions. Seven introns belong to group I, while the intronic sequence i5 constitutes the first typical group II intron reported in a fungal mitochondrial LSU rDNA. As with most fungal LSU rDNA introns reported to date, four introns (i5-i8) are distributed in domain V associated with the peptidyl-transferase activity. One intron (i1) is located in domain I, and three (i2-i4) in domain II. The introns i2-i8 possess homologies with other fungal, algal or protozoan introns located at the same position in LSU rDNAs. One of them (i6) is located at the same insertion site as most Ascomycota or algae LSU introns, suggesting a possible inheritance from a common ancestor. On the contrary, intron i1 is located at a so-far unreported insertion site. Among the six unusual nucleotide extensions, five are located in domain I and one in domain V. This is the first report of a mitochondrial LSU rRNA gene sequence and secondary structure for the whole Basidiomycota division.  相似文献   

16.
Homing endonucleases (HEs) of the LAGLIDADG family cleave intron/inteinless cognate DNA at, or near, the insertion site (IS) of their own intron/intein. Here, we describe a notable exception to this rule. Two introns, Pog.S1205 (length 32 bp) and Pog.S1213 (664 bp), whose ISs are 8 bp apart, exist within the 16S rRNA gene of the archaeon Pyrobaculum oguniense. Pog.S1213 harbors a nested open reading frame (ORF) encoding a 22 kDa monomeric protein, I-PogI, which contains two LAGLIDADG motifs and has optimal DNA cleavage activity at 90 degrees C. Intriguingly, I-PogI cleaves the Pog.S1205-less substrate DNA in the presence or absence of Pog.S1213. The cleavage site (CS) of I-PogI does not coincide with the IS of Pog.S1213 but with that of Pog.S1205. Thus, I-PogI activity both promotes the homing of its own intron, Pog.S1213, and guarantees co-conversion of the ORF-less intron Pog.S1205.  相似文献   

17.
The protein encoded by intron 1 of the single 23S rRNA gene of the archaeal hyperthermophile Pyrobaculum organotrophum was isolated and shown to constitute a homing-type DNA endonuclease, I-PorI. It cleaves the intron- 23S rDNA of the closely related organism Pyrobaculum islandicum near the site of intron insertion in Pb.organotrophum. In contrast, no endonuclease activity was detected for the protein product of intron 2 of the same gene of Pb.organotrophum which, like I-PorI, carries the LAGLI-DADG motif, common to group I intron-encoded homing enzymes. I-PorI cleaves optimally at 80 degrees C, with kcat and Km values of about 2 min-1 and 4 nM, respectively, and generates four nucleotide 3'-overhangs and 5'-phosphates. It can cleave a 25 base pair DNA fragment encompassing the intron insertion site. A mutation-selection study established the base pair specificity of the endonuclease within a 17 bp region, from positions -6 to +11 with respect to the intron-insertion site. Four of the essential base pairs encode the sequence involved in the intron-exon interaction in the pre-rRNA that is required for recognition by the RNA splicing enzymes. Properties of the enzyme are compared and contrasted with those of eucaryotic homing endonucleases.  相似文献   

18.
M. M. Parker  D. A. Court  K. Preiter    M. Belfort 《Genetics》1996,143(3):1057-1068
Many group I introns encode endonucleases that promote intron homing by initiating a double-strand break-mediated homologous recombination event. A td intron-phage λ model system was developed to analyze exon homology effects on intron homing and determine the role of the λ 5'-3' exonuclease complex (Redαβ) in the repair event. Efficient intron homing depended on exon lengths in the 35- to 50-bp range, although homing levels remained significantly elevated above nonbreak-mediated recombination with as little as 10 bp of flanking homology. Although precise intron insertion was demonstrated with extremely limiting exon homology, the complete absence of one exon produced illegitimate events on the side of heterology. Interestingly, intron inheritance was unaffected by the presence of extensive heterology at the double-strand break in wild-type λ, provided that sufficient homology between donor and recipient was present distal to the heterologous sequences. However, these events involving heterologous ends were absolutely dependent on an intact Red exonuclease system. Together these results indicate that heterologous sequences can participate in double-strand break-mediated repair and imply that intron transposition to heteroallelic sites might occur at break sites within regions of limited or no homology.  相似文献   

19.
20.
Pp LSU3 is a mobile group I intron in the extrachromosomal nuclear ribosomal DNA (rDNA) of Physarum polycephalum. As found for other mobile introns, Pp LSU3 encodes a site-specific endonuclease, I-Ppo, which mediates "homing" to unoccupied target sites in Physarum rDNA. The recognition sequence for this enzyme is conserved in all eucaryotic nuclear rDNAs. We have introduced this intron into a heterologous species, Saccharomyces cerevisiae, in which nuclear group I introns have not been detected. The expression of Pp LSU3, under control of the inducible GAL10 promoter, was found to be lethal as a consequence of double-strand breaks in the rDNA. However, surviving colonies that are resistant to the lethal effects of I-Ppo because of alterations in the rDNA at the cleavage site were recovered readily. These survivors are of two classes. The first comprises cells that acquired one of three types of point mutations. The second comprises cells in which Pp LSU3 became inserted into the rDNA. In both cases, each resistant survivor appears to carry the same alterations in all approximately 150 rDNA repeats. When it is embedded in yeast rDNA, Pp LSU3 leads to the synthesis of I-Ppo and appears to be mobile in appropriate genetic crosses. The existence of yeast cells carrying a mobile intron should allow dissection of the steps that allow expression of the highly unusual I-Ppo gene.  相似文献   

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