Large Non-coding Region Research Articles

Mitogenomics reveals two subspecies in Coelomactra antiquata (Mollusca: Bivalvia)

The mitochondrial genome sequence of Coelomactra antiquata (Mollusca: Bivalvia) in Zhangzhou (zz-mtDNA) was fully sequenced and compared with that in Rizhao (rz-mtDNA) in this study. A tRNA (tRNAMet) located between tRNAAla and cox1 genes was identified in zz-mtDNA but not in rz-mtDNA. The largest non-coding region (NCR; MNR) contained 11 copies 99nt tandem repeat sequences exclusively in rz-mtDNA, while the second largest NCR with 400 bp between tRNAAla and tRNAMet in zz-mtDNA was absent in rz-mtDNA. Secondary structures of ZZ and RZ C. antiquata rRNAs are significantly different. The mitochondrial genomic characteristics clearly indicate that there are at least two subspecies in C. antiquata.

Complete mitochondrial genome of Coelomactra antiquata (Mollusca: Bivalvia): The first representative from the family Mactridae with novel gene order and unusual tandem repeats

The complete mitochondrial genome plays an important role in the accurate inference of phylogenetic relationships among metazoans. Mactridae, also known as trough shells or duck clams, is an important family of marine bivalve clams in the order Veneroida. Here we present the complete mitochondrial genome sequence of the Xishishe Coelomactra antiquata (Mollusca: Bivalvia), which is the first representative from the family Mactridae. The mitochondrial genome of C. antiquata is of 17,384bp in length, and encodes 35 genes, including 12 protein-coding, 21 transfer RNA, and 2 ribosomal RNA genes. Compared with the typical gene content of animal mitochondrial genomes, atp8 and tRNAS2 are missing. Gene order of the mitochondrial genome of C. antiquata is unique compared with others from Veneroida. In the mitochondrial genome of the C. antiquata, a total of 2189bp of non-coding nucleotides are scattered among 26 non-coding regions. The largest non-coding region contains one section of tandem repeats (99 bp×11), which is the second largest tandem repeats found in the mitochondrial genomes from Veneroida. The phylogenetic trees based on mitochondrial genomes support the monophyly of Veneridae and Lucinidae, and the relationship at the family level: ((Veneridae+Mactridae)+(Cardiidae+Solecurtidae))+Lucinidae. The phylogenetic result is consistent with the morphological classification. Meanwhile, bootstrap values are very high (BP=94–100), suggesting that the evolutionary relationship based on mitochondrial genomes is very reliable.

The complete mitochondrial genome sequence of Euphausia pacifica (Malacostraca: Euphausiacea) reveals a novel gene order and unusual tandem repeats

Euphausiid krill are dominant organisms in the zooplankton population and play a central role in marine ecosystems. Euphausia pacifica (Malacostraca: Euphausiacea) is one of the most important and dominant crustaceans in the North Pacific Ocean. In this paper, we described the gene content, organization, and codon usage of the E. pacifica mitochondrial genome. The mitochondrial genome of E. pacifica is 16 898 bp in length and contains a standard set of 13 protein-coding genes, 2 ribosomal RNA genes, and 22 transfer RNA genes. Translocation of three transfer RNAs (trnL(1), trnL(2), and trnW) was found in the E. pacifica mitochondrial genome when comparing with the pancrustacean ground pattern. The rate of K(a)/K(s) in 13 protein-coding genes among three krill is much less than 1, which indicates a strong purifying selection within this group. The largest noncoding region in the E. pacifica mitochondrial genome contains one section with tandem repeats (4.7 x 154 bp), which are the largest tandem repeats found in malacostracan mitochondrial genomes so far. All analyses based on nucleotide and amino acid data strongly support the monophyly of Stomatopoda, Penaeidae, Caridea, Brachyura, and Euphausiacea. The Bayesian analysis of nucleotide and amino acid datasets strongly supports the close relationship between Euphausiacea and Decapoda, which confirms traditional findings. The maximum likelihood analysis based on amino acid data strongly supports the close relationship between Euphausiacea and Penaeidae, which destroys the monophyly of Decapoda.

Complete nucleotide sequence and organization of the mitochondrial genome of eri-silkworm, Samia cynthia ricini (Lepidoptera: Saturniidae)

Samia cynthia ricini is a commercial silk-producing insect that is now reared year-round in Korea, with the expectation of being utilized for diverse purposes. In this report, we present the complete mitochondrial genome (mitogenome) of S. c. ricini. The 15,384-bp long S. cynthia ricini mitogenome was amplified into 26 short fragments using three long overlapping fragments using primers designed from reported lepidopteran mitogenome sequences. The genome comprises 37 genes (13 protein-coding genes, two rRNA genes, and 22 tRNA genes), and one large non-coding region termed the A + T-rich region. The A/T content of the third codon position was 91.7%, which was 18.8% and 21.6% higher than those of first and second codon positions, respectively. The high A/T content in the genome is reflected in codon usage, accounting for 39.5% of A/T-composed codons (TTA, ATT, TTT, and ATA). Unlike a previous report on the start codon for the COI gene, the S. c. ricini COI gene commences with a typical ATT codon. A total of 221 bp of non-coding sequences are dispersed in 17 regions, ranging in size from 1 to 54 bp, which comprise 1.4% of the total genome. One of the non-coding sequence located between tRNA Gln and ND2 (54 bp) has 77% sequence homology with the 5′-sequence of the neighboring ND2 gene, suggesting partial duplication of the sequence during evolution. The 361-bp long A + T-rich region contains an 18 bp-long poly-T stretch, ATAGA motif, ATTTA element, microsatellite-like A/T sequence, poly-A stretch and one tRNA-like sequence, as typically found in Lepidoptera including Bombycoidea.

The mitochondrial genome of Polylabris halichoeres (Monogenea: Microcotylidae)

This study presents the complete mitochondrial (mt) genome of Polylabris halichoeres, which is the largest mt genome sequenced of monogeneans so far and the second complete sequence after Microcotyle sebastis from the Microcotylidae. It is basically similar to that of M. sebastis, with the exception of a high level of gene rearrangement located between trnC and trnL(UUR), a translocation of trnM and trnH, as well as a highly repetitive region (HRR) in the large non-coding region (NCR). We also find a series of trnI pseudogenes (ΨI) and one unknown short open reading frame (ORF) in the large NCR. Although the ORF cannot be unambiguously regarded as an atp8 gene, we cannot rule out the possibility that it has other functional importance, but it need further study in the future.

Distinctive mitochondrial genome of Calanoid copepod Calanus sinicus with multiple large non-coding regions and reshuffled gene order: Useful molecular markers for phylogenetic and population studies

BackgroundCopepods are highly diverse and abundant, resulting in extensive ecological radiation in marine ecosystems. Calanus sinicus dominates continental shelf waters in the northwest Pacific Ocean and plays an important role in the local ecosystem by linking primary production to higher trophic levels. A lack of effective molecular markers has hindered phylogenetic and population genetic studies concerning copepods. As they are genome-level informative, mitochondrial DNA sequences can be used as markers for population genetic studies and phylogenetic studies.ResultsThe mitochondrial genome of C. sinicus is distinct from other arthropods owing to the concurrence of multiple non-coding regions and a reshuffled gene arrangement. Further particularities in the mitogenome of C. sinicus include low A + T-content, symmetrical nucleotide composition between strands, abbreviated stop codons for several PCGs and extended lengths of the genes atp6 and atp8 relative to other copepods. The monophyletic Copepoda should be placed within the Vericrustacea. The close affinity between Cyclopoida and Poecilostomatoida suggests reassigning the latter as subordinate to the former. Monophyly of Maxillopoda is rejected. Within the alignment of 11 C. sinicus mitogenomes, there are 397 variable sites harbouring three 'hotspot' variable sites and three microsatellite loci.ConclusionThe occurrence of the circular subgenomic fragment during laboratory assays suggests that special caution should be taken when sequencing mitogenomes using long PCR. Such a phenomenon may provide additional evidence of mitochondrial DNA recombination, which appears to have been a prerequisite for shaping the present mitochondrial profile of C. sinicus during its evolution. The lack of synapomorphic gene arrangements among copepods has cast doubt on the utility of gene order as a useful molecular marker for deep phylogenetic analysis. However, mitochondrial genomic sequences have been valuable markers for resolving phylogenetic issues concerning copepods. The variable site maps of C. sinicus mitogenomes provide a solid foundation for population genetic studies.

Identification of two novel equine papillomavirus sequences suggests three genera in one cluster

The number of recognized papillomavirus (PV) species and potential PV genera has dramatically been increasing throughout the past decade. It seems that every host species might potentially harbour a large set of PVs, while the PVs of each species appear to belong to only a few genera.In horses at least three conditions beside the equine sarcoid have been described, being supposedly PV induced namely classical equine papillomas, genital papillomas and aural plaques. We were able to identify the DNA of novel equine PVs (EcPVs) in the two latter disorders where PV involvement had been predicted. Both PV genomes were entirely cloned and sequenced. Both EcPV genomes, one derived from a penile papilloma, the other derived from an ear papilloma contain the characteristic open reading frames (ORFs) E6, E7, E1, E2, L2 and L1, a large non-coding region between the late and early region as well as a small non-coding region between the early and the late region. The viruses were consequently designated as EcPV2 and EcPV3.The genomes of the three equine PVs were analysed and compared with each other and further PVs. Upon phylogenetic analyses the equine PVs group well together. Pairwise alignment of the L1 nucleotide sequences reveals that EcPV1 shares 54.9% identities with EcPV2 and 53.2% with EcPV3. EcPV2 and EcPV3 share 51.3% identities. As the three EcPVs share less than 60% of nucleotide identities in L1, they may be regarded as belonging to different genera.

Designing L-Systems for making three and six open reading frames from the leading strand of a single DNA molecule

AbstractCurrent molecular biology dogma (except alternative splicing) suggests that leading strand of DNA produces mRNA and usually the leading strand gives one ORF (ORF) among the available six ORFs. We attempted to break this rule where three and six ORFs from the leading strand will be formed by designing L-Systems, originally proposed to study symmetry of biological world. We thought this would revolutionize the world of synthetic biology. While doing so, we found L-System containing A→T, T→G, C→A, and G→C, i.e., single nucleotide production rule will stop growing after one nucleotide. L-System containing either (a) A→TG, T→GC, C→GA, G→TC or (b) A→CTG, T→GAC, C→TAC, G→TGC will lead to generation of repetitive DNA sequence after few iteration. These repetitive sequences might become easy target for enzyme like restriction endonucleases and transposase or RNAse but may give repetitive DNA sequences which are normally found in the large non-coding region of the genome. Here we show that a number of L-Systems like with (Axiom: A) A→ATGC, C→CTCA, T→GCAG, G→CGAA and A→GTAC, C→TGCA, T→ CTGG, G→ATAT if used to generate either a short (~ 2000 bp) or long (~ 16000 bp) can give rise to three ORFs and six ORFs per DNA molecule when conceptually translated through Expasy tools respectively. To our surprise we find that L-System production rule with only four letter codes (not one, two, three, five, six) can produce this miraculous results which nothing but follows Young Fibonacci graph. We find that production rule with higher order of magnitude like 10 bp also give similar results but not 20 bp containing production rules.

Mitochondrial genome analysis of the predatory mitePhytoseiulus persimilisand a revisit of theMetaseiulus occidentalismitochondrial genome

In this study we sequenced and analysed the complete mitochondrial (mt) genome of the Chilean predatory mite Phytoseiulus persimilis Athias-Henriot (Chelicerata: Acari: Mesostigmata: Phytoseiidae: Amblyseiinae). The 16 199 bp genome (79.8% AT) contains the standard set of 13 protein-coding and 24 RNA genes. Compared with the ancestral arthropod mtDNA pattern, the gene order is extremely reshuffled (35 genes changed position) and represents a novel arrangement within the arthropods. This is probably related to the presence of several large noncoding regions in the genome. In contrast with the mt genome of the closely related species Metaseiulus occidentalis (Phytoseiidae: Typhlodrominae) - which was reported to be unusually large (24 961 bp), to lack nad6 and nad3 protein-coding genes, and to contain 22 tRNAs without T-arms - the genome of P. persimilis has all the features of a standard metazoan mt genome. Consequently, we performed additional experiments on the M. occidentalis mt genome. Our preliminary restriction digests and Southern hybridization data revealed that this genome is smaller than previously reported. In addition, we cloned nad3 in M. occidentalis and positioned this gene between nad4L and 12S-rRNA on the mt genome. Finally, we report that at least 15 of the 22 tRNAs in the M. occidentalis mt genome can be folded into canonical cloverleaf structures similar to their counterparts in P. persimilis.

Identification of transcribed and persistent variants of the psbA gene carried by plastid minicircles in a dinoflagellate

Plastid genomes of the peridinin-containing dinoflagellates are composed of a limited number of genes that are contained separately on small circular molecules (minicircles). It has been shown that occasionally aberrant minicircles are generated, but they are usually small and contain little coding information. In this study, we discovered multiple variants, a putative "gene family", of the gene psbA in the plastid minicircles of the dinoflagellate Alexandrium tamarense, which have persisted for almost 3 years in culture. Each variant, like the ordinary psbA, existed on distinct minicircles of similar size (5-6 kb). These psbA variants retained all, or almost all, the coding sequence of the ordinary gene, and all four were transcribed and edited after transcription, even though they could not encode the entire protein due to intervening or translocated sequences. Repeat elements were generally found in the relatively large non-coding region of these minicircles. Each psbA variant might have been generated by DNA recombination and/or replication slippage, as for previously reported aberrant minicircles. The fact that these minicircles are transcribed, individually edited and maintained in the genome suggests that they are functionally important, although their precise roles remain unclear.

The complete mitochondrial genome of the house dust mite Dermatophagoides pteronyssinus (Trouessart): a novel gene arrangement among arthropods.

BackgroundThe apparent scarcity of available sequence data has greatly impeded evolutionary studies in Acari (mites and ticks). This subclass encompasses over 48,000 species and forms the largest group within the Arachnida. Although mitochondrial genomes are widely utilised for phylogenetic and population genetic studies, only 20 mitochondrial genomes of Acari have been determined, of which only one belongs to the diverse order of the Sarcoptiformes. In this study, we describe the mitochondrial genome of the European house dust mite Dermatophagoides pteronyssinus, the most important member of this largely neglected group.ResultsThe mitochondrial genome of D. pteronyssinus is a circular DNA molecule of 14,203 bp. It contains the complete set of 37 genes (13 protein coding genes, 2 rRNA genes and 22 tRNA genes), usually present in metazoan mitochondrial genomes. The mitochondrial gene order differs considerably from that of other Acari mitochondrial genomes. Compared to the mitochondrial genome of Limulus polyphemus, considered as the ancestral arthropod pattern, only 11 of the 38 gene boundaries are conserved. The majority strand has a 72.6% AT-content but a GC-skew of 0.194. This skew is the reverse of that normally observed for typical animal mitochondrial genomes. A microsatellite was detected in a large non-coding region (286 bp), which probably functions as the control region. Almost all tRNA genes lack a T-arm, provoking the formation of canonical cloverleaf tRNA-structures, and both rRNA genes are considerably reduced in size. Finally, the genomic sequence was used to perform a phylogenetic study. Both maximum likelihood and Bayesian inference analysis clustered D. pteronyssinus with Steganacarus magnus, forming a sistergroup of the Trombidiformes.ConclusionAlthough the mitochondrial genome of D. pteronyssinus shares different features with previously characterised Acari mitochondrial genomes, it is unique in many ways. Gene order is extremely rearranged and represents a new pattern within the Acari. Both tRNAs and rRNAs are truncated, corroborating the theory of the functional co-evolution of these molecules. Furthermore, the strong and reversed GC- and AT-skews suggest the inversion of the control region as an evolutionary event. Finally, phylogenetic analysis using concatenated mt gene sequences succeeded in recovering Acari relationships concordant with traditional views of phylogeny of Acari.

A unique genetic code change in the mitochondrial genome of the parasitic nematode Radopholus similis

BackgroundMitochondria (mt) contain their own autonomously replicating DNA, constituted as a small circular genome encoding essential subunits of the respiratory chain. Mt DNA is characterized by a genetic code which differs from the standard one. Interestingly, the mt genome of nematodes share some peculiar features, such as small transfer RNAs, truncated ribosomal RNAs and - in the class of Chromadorean nematodes - unidirectional transcription.FindingsWe present the complete mt genomic sequence (16,791 bp) of the plant-parasitic nematode Radopholus similis (class Chromadorea). Although it has a gene content similar to most other nematodes, many idiosyncrasies characterize the extremely AT-rich mt genome of R. similis (85.4% AT). The secondary structure of the large (16S) rRNA is further reduced, the gene order is unique, the large non-coding region contains two large repeats, and most interestingly, the UAA codon is reassigned from translation termination to tyrosine. In addition, 7 out of 12 protein-coding genes lack a canonical stop codon and analysis of transcriptional data showed the absence of polyadenylation. Northern blot analysis confirmed that only one strand is transcribed and processed. Furthermore, using nucleotide content bias methods, regions for the origin of replication are suggested.ConclusionThe extraordinary mt genome of R. similis with its unique genetic code appears to contain exceptional features correlated to DNA decoding. Therefore the genome may provide an incentive to further elucidate these barely understood processes in nematodes. This comprehension may eventually lead to parasitic nematode-specific control targets as healthy mitochondria are imperative for organism survival. In addition, the presented genome is an interesting exceptional event in genetic code evolution.

The complete mitochondrial DNA sequence of the monogenean Gyrodactylus thymalli (Platyhelminthes: Monogenea), a parasite of grayling ( Thymallus thymallus)

We present the complete mitochondrial (mt) genome of Gyrodactylus thymalli, a monogenean ectoparasite on grayling ( Thymallus thymallus). The circular genome is 14788 bp in size and includes all 35 genes recognized from other flatworm mt genomes. The overall A + T content of the mt genome is 62.8%. Twenty regions of non-coding DNA ranging from 1 to 111 bp in length were identified in addition to 2 highly conserved large non-coding regions 799 bp and 767 bp in size. Compared to the recently described mt DNA of the closely related G. salaris from Atlantic salmon from Signaldalselva, Norway, the mitochondrial genome of G. thymalli from Hnilec, Slovakia, differs on average by 2.2%.

Glass Sponges and Bilaterian Animals Share Derived Mitochondrial Genomic Features: A Common Ancestry or Parallel Evolution?

Glass sponges (Hexactinellida) are a group of deep-water benthic animals that have a unique syncytial organization and possess a characteristic siliceous skeleton. Although hexactinellids are traditionally grouped with calcareous and demosponges in the phylum Porifera, the monophyly of sponges and the phylogenetic position of the Hexactinellida remain contentious. We determined and analyzed the nearly complete mitochondrial genome sequences of the hexactinellid sponges Iphiteon panicea and Sympagella nux. Unexpectedly, our analysis revealed several mitochondrial genomic features shared between glass sponges and bilaterian animals, including an Arg --> Ser change in the genetic code, a characteristic secondary structure of one of the serine tRNAs, highly derived tRNA and rRNA genes, and the presence of a single large noncoding region. At the same time, glass sponge mtDNA contains atp9, a gene previously found only in the mtDNA of demosponges (among animals), and encodes a tRNA(Pro);(UGG) with an atypical A11-U24 pair that is also found in demosponges and placozoans. Most of our sequence-based phylogenetic analyses place Hexactinellida as the sister group to the Bilateria; however, these results are suspect given accelerated rates of mitochondrial sequence evolution in these groups. Thus, it remains an open question whether shared mitochondrial genomic features in glass sponges and bilaterian animals reflect their close phylogenetic affinity or provide a remarkable example of parallel evolution.

Complete mitochondrial genome of a Chinese scorpionMesobuthus martensii(Chelicerata, Scorpiones, Buthidae)

The complete mitochondrial genome (15,034 bp) of a Chinese scorpion Mesobuthus martensii (Buthidae) was sequenced and characterized in detail. The genome contains 13 protein-coding genes, 21 transfer RNA genes, two ribosomal RNA genes and a large non-coding region ( = CR). Its gene arrangement pattern is identical to that of Limulus polyphemus (Chelicerata, Xiphosura), with the exceptions of the tRNA(Glu)-tRNA(Ile)-tRNA(Met) (Q-I-M) arrangement and tRNA(Asp)-loss. Additional interesting features are found and discussed: high frequency of Leu(UUG) codon use, low A+T content of the genome (66.75%), and six repeat units (five 60-nt-long and one 58-nt-long repeats) in the 998-nt CR. Bayesian analysis based on amino acid sequences of the 12 proteincoding genes (excluding ATP8) reveals that the family Buthidae (Order Scorpiones) and the class Arachnida form strong monophyletic groups within Chelicerata, respectively. It indicated that the scorpions are the most ancestral arachnids.

The complete sequences and gene organisation of the mitochondrial genomes of the heterodont bivalves Acanthocardia tuberculata and Hiatella arctica – and the first record for a putative Atpase subunit 8 gene in marine bivalves

BackgroundMitochondrial (mt) gene arrangement is highly variable among molluscs and especially among bivalves. Of the 30 complete molluscan mt-genomes published to date, only one is of a heterodont bivalve, although this is the most diverse taxon in terms of species numbers. We determined the complete sequence of the mitochondrial genomes of Acanthocardia tuberculata and Hiatella arctica, (Mollusca, Bivalvia, Heterodonta) and describe their gene contents and genome organisations to assess the variability of these features among the Bivalvia and their value for phylogenetic inference.ResultsThe size of the mt-genome in Acanthocardia tuberculata is 16.104 basepairs (bp), and in Hiatella arctica 18.244 bp. The Acanthocardia mt-genome contains 12 of the typical protein coding genes, lacking the Atpase subunit 8 (atp8) gene, as all published marine bivalves. In contrast, a complete atp8 gene is present in Hiatella arctica. In addition, we found a putative truncated atp8 gene when re-annotating the mt-genome of Venerupis philippinarum. Both mt-genomes reported here encode all genes on the same strand and have an additional trnM. In Acanthocardia several large non-coding regions are present. One of these contains 3.5 nearly identical copies of a 167 bp motive. In Hiatella, the 3' end of the NADH dehydrogenase subunit (nad)6 gene is duplicated together with the adjacent non-coding region. The gene arrangement of Hiatella is markedly different from all other known molluscan mt-genomes, that of Acanthocardia shows few identities with the Venerupis philippinarum. Phylogenetic analyses on amino acid and nucleotide levels robustly support the Heterodonta and the sister group relationship of Acanthocardia and Venerupis. Monophyletic Bivalvia are resolved only by a Bayesian inference of the nucleotide data set. In all other analyses the two unionid species, being to only ones with genes located on both strands, do not group with the remaining bivalves.ConclusionThe two mt-genomes reported here add to and underline the high variability of gene order and presence of duplications in bivalve and molluscan taxa. Some genomic traits like the loss of the atp8 gene or the encoding of all genes on the same strand are homoplastic among the Bivalvia. These characters, gene order, and the nucleotide sequence data show considerable potential of resolving phylogenetic patterns at lower taxonomic levels.

Molecular Mechanisms for the Variation of Mitochondrial Gene Content and Gene Arrangement Among Chigger Mites of the Genus Leptotrombidium (Acari: Acariformes)

The gene content of a mitochondrial (mt) genome, i.e., 37 genes and a large noncoding region (LNR), is usually conserved in Metazoa. The arrangement of these genes and the LNR is generally conserved at low taxonomic levels but varies substantially at high levels. We report here a variation in mt gene content and gene arrangement among chigger mites of the genus Leptotrombidium. We found previously that the mt genome of Leptotrombidium pallidum has an extra gene for large-subunit rRNA (rrnL), a pseudo-gene for small-subunit rRNA (PrrnS), and three extra LNRs, additional to the 37 genes and an LNR typical of Metazoa. Further, the arrangement of mt genes of L. pallidum differs drastically from that of the hypothetical ancestor of the arthropods. To find to what extent the novel gene content and gene arrangement occurred in Leptotrombidium, we sequenced the entire or partial mt genomes of three other species, L. akamushi, L. deliense, and L. fletcheri. These three species share the arrangement of all genes with L. pallidum, except trnQ (for tRNA-glutamine). Unlike L. pallidum, however, these three species do not have extra rrnL or PrrnS and have only one extra LNR. By comparison between Leptotrombidium species and the ancestor of the arthropods, we propose that (1) the type of mt genome present in L. pallidum evolved from the type present in the other three Leptotrombidium species, and (2) three molecular mechanisms were involved in the evolution of mt gene content and gene arrangement in Leptotrombidium species.

The mitochondrial genome ofGyrodactylus salaris(Platyhelminthes: Monogenea), a pathogen of Atlantic salmon (Salmo salar)

In the present study, we describe the complete mitochondrial (mt) genome of the Atlantic salmon parasite Gyrodactylus salaris, the first for any monogenean species. The circular genome is 14,790 bp in size. All of the 35 genes recognized from other flatworm mitochondrial genomes were identified, and they are transcribed from the same strand. The protein-coding and ribosomal RNA (rRNA) genes share the same gene arrangement as those published previously for neodermatan mt genomes (representing cestodes and digeneans only), and the genome has an overall A+T content of 65%. Three transfer RNA (tRNA) genes overlap with other genes, whereas the secondary structure of 3 tRNA genes lack the DHU arm and 1 tRNA gene lacks the TphiC arm. Eighteen regions of non-coding DNA ranging from 4 to 112 bp in length, totalling 278 bp, were identified as well as 2 large non-coding regions (799 bp and 768 bp) that were almost identical to each other. The completion of the mt genome offers the opportunity of defining new molecular markers for studying evolutionary relationships within and among gyrodactylid species.

Multiple promoters control expression of the Yersinia enterocolitica phage-shock-protein A (pspA) operon

The widely conserved phage-shock-protein A (pspA) operon encodes an extracytoplasmic stress response system that is essential for virulence in Yersinia enterocolitica, and has been linked to other important phenotypes in Escherichia coli, Salmonella enterica and Shigella flexneri. Regulation of pspA operon expression is mediated through a promoter upstream of pspA that depends on sigma factor RpoN (sigma(54)) and the enhancer binding protein PspF. PspA, PspB and PspC, encoded within the pspA operon, also regulate expression by participating in a putative signal transduction pathway that probably serves to modulate PspF activity. All of this suggests that appropriate expression of the pspA operon is critical. Previous genetic analysis of the Y. enterocolitica pspA operon suggested that an additional level of complexity might be mediated by PspF/RpoN-independent expression of some psp genes. Here, an rpoN null mutation and interposon analysis were used to confirm that PspF/RpoN-independent gene expression does originate within the psp locus. Molecular genetic approaches were used to systematically analyse the two large non-coding regions within the psp locus. Primer extension, control region deletion and site-directed mutagenesis experiments led to the identification of RpoN-independent promoters both upstream and downstream of pspA. The precise location of the PspF/RpoN-dependent promoter upstream of pspA was also determined. The discovery of these RpoN-independent promoters reveals yet another level of transcriptional complexity for the Y. enterocolitica pspA operon that may function to allow low-level constitutive expression of psp genes and/or additional regulation under some conditions.

Relaxed Transcription in Arabidopsis Mitochondria Is Counterbalanced by RNA Stability Control Mediated by Polyadenylation and Polynucleotide Phosphorylase

Plant mitochondrial genomes are extraordinarily large and complex compared to their animal counterparts, due to the presence of large noncoding regions. Multiple promoters are common for plant mitochondrial genes, and transcription exhibits little or no modulation. Mature functional RNAs are produced through various posttranscriptional processes, and control of RNA stability has a major impact on RNA abundance. This control involves polyadenylation which targets RNA for degradation by polynucleotide phosphorylase (PNPase). Here, we have analyzed polyadenylated RNA fragments from Arabidopsis plants down-regulated for PNPase (PNP- plants). Because of their polyadenylated status and the accumulation of the corresponding RNA in PNP- versus wild-type plants, these sequences represent mitochondrial RNA degradation tags. Analysis of these tags revealed that PNPase is involved in degrading rRNA and tRNA maturation by-products but also RNA transcribed from regions that are in some cases highly expressed although lacking known functional genes. Some of these transcripts, such as RNA containing chimeric open reading frames created by recombination or antisense RNA transcribed on the opposite strand of a known gene, may present potential detrimental effects to mitochondrial function. Taken together, our data show that the relaxed transcription in Arabidopsis mitochondria is counterbalanced by RNA stability control mediated by polyadenylation and PNPase.