Structure and organization of Marchantia polymorpha chloroplast genome. III. Gene organization of the large single copy region from rbcL to trnI(CAU)

The nucleotide sequence (25,320 base-pairs) of a part of the large single-copy region of chloroplast DNA from the liverwort Marchantia polymorpha was determined. This region encodes putative genes for four tRNAs, isoleucine tRNA(CAU), arginine tRNA(CCG), proline tRNA(UGG) and tryptophan tRNA(CCA); eight photosynthetic polypeptides, the large subunit of ribulose bisphosphate carboxylase/oxygenase (rbcL), 51,000 Mr photosystem II chlorophyll alpha apoprotein (psbB), apocytochrome b-559 polypeptides (psbE and psbF), 10,000 Mr phosphoprotein (psbH), cytochrome f preprotein (petA), cytochrome b6 polypeptide (petB), and cytochrome b6/f complex subunit 4 polypeptide (petD); 13 ribosomal proteins (L2, L14, L16, L20, L22, L23, L33, S3, S8, S11, S12, S18 and S19); initiation factor 1 (infA); ribosome-associating polypeptide (secX); and alpha subunit of RNA polymerase (rpoA). Functionally related genes were located in several clusters in this region of the genome. There were two ribosomal protein gene clusters: rpl23-rpl2-rps19-rpl22-rps3-rpl16-+ ++rpl14-rps8-infA-secX-rps11-rpoA, with a gene arrangement similar to that of the Escherichia coli S10-spc-alpha operons, and the rps12'-rpl20-rps18-rpl33 cluster. There were gene clusters encoding photosynthesis components such as the psbB-psbH-petB-petD and the psbE-psbF clusters. Thirteen open reading frames, ranging in length from 31 to 434 amino acid residues, remain to be identified.     

Structure and organization of Marchantia polymorpha chloroplast genome. IV. Inverted repeat and small single copy regions

We characterized the genes in the regions of large inverted repeats (IRA and IRB, 10,058 base-pairs each) and a small single copy (SSC 19,813 bp) of chloroplast DNA from Marchantia polymorpha. The inverted repeat (IR) regions contain genes for four ribosomal RNAs (16 S, 23 S, 4.5 S and 5 S rRNAs) and five transfer RNAs (valine tRNA(GAC), isoleucine tRNA(GAU), alanine tRNA(UGC), arginine tRNA(ACG) and asparagine tRNA(GUU)). The gene organization of the IR regions in the liverwort chloroplast genome is conserved, although the IR regions are smaller (10,058 base-pairs) than any reported in higher plant chloroplasts. The small single-copy region (19,813 base-pairs) encoded genes for 17 open reading frames, a leucine tRNA(UAG) and a proline tRNA(GGG)-like sequence. We identified 12 open reading frames by homology of their coding sequences to a 4Fe-4S-type ferredoxin protein, a bacterial nitrogenase reductase component (Fe-protein), five human mitochondrial components of NADH dehydrogenase (ND1, ND4, ND4L, ND5 and ND6), two Escherichia coli ribosomal proteins (S15 and L21), two putative proteins encoded in the kinetoplast maxicircle DNA of Leishmania tarentolae (LtORF 3 and LtORF 4), and a bacterial permease inner membrane component (encoded by malF in E. coli or hisQ in Salmonella typhimurium).     

Gene organization deduced from the complete sequence of liverwort Marchantia polymorpha mitochondrial DNA. A primitive form of plant mitochondrial genome.

Analysis of the mitochondrial DNA of a liverwort Marchantia polymorpha by electron microscopy and restriction endonuclease mapping indicated that the liverwort mitochondrial genome was a single circular molecule of about 184,400 base-pairs. We have determined the complete sequence of the liverwort mitochondrial DNA and detected 94 possible genes in the sequence of 186,608 base-pairs. These included genes for three species of ribosomal RNA, 29 genes for 27 species of transfer RNA and 30 open reading frames (ORFs) for functionally known proteins (16 ribosomal proteins, 3 subunits of H(+)-ATPase, 3 subunits of cytochrome c oxidase, apocytochrome b protein and 7 subunits of NADH ubiquinone oxidoreductase). Three ORFs showed similarity to ORFs of unknown function in the mitochondrial genomes of other organisms. Furthermore, 29 ORFs were predicted as possible genes by using the index of G + C content in first, second and third letters of codons (42.0 +/- 10.9%, 37.0 +/- 13.2% and 26.4 +/- 9.4%, respectively) obtained from the codon usages of identified liverwort genes. To date, 32 introns belonging to either group I or group II intron have been found in the coding regions of 17 genes including ribosomal RNA genes (rrn18 and rrn26), a transfer RNA gene (trnS) and a pseudogene (psi nad7). RNA editing was apparently lacking in liverwort mitochondria since the nucleotide sequences of the liverwort mitochondrial DNA were well-conserved at the DNA level.     

Genome walking with consensus primers: application to the large single copy region of chloroplast DNA

Conserved chloroplast (cp) DNA primer pairs are useful in plant molecular genetics, evolution and ecology. We have designed 20 conserved cpDNA primer pairs that, in combination with 18 previously described ones, amplify overlapping fragments (mean size of 2.5 kb) spanning the large single copy (LSC) region from Eudicots. These 38 primer pairs as well as eight primer pairs flanking cpDNA microsatellites were tested on 20 plant species belonging to 13 families. At least 79% and up to 100% of the LSC (> 86 kb) can be amplified. Many primer pairs are robust and work with all species.     

Gene clusters for ribosomal proteins in the mitochondrial genome of a liverwort, Marchantia polymorpha.

We detected 16 genes for ribosomal proteins in the complete sequence of the mitochondrial DNA from a liverwort, Marchantia polymorpha. The genes formed two major clusters, rps12-rps7 and rps10-rpl2-rps19-rps3-rpl16-rpl5- rps14-rps8- rpl6-rps13-rps11-rps1, very similar in organization to Escherichia coli ribosomal protein operons (str and S10-spc-alpha operons, respectively). In contrast, rps2 and rps4 genes were located separately in the liverwort mitochondrial genome (the latter was part of the alpha operon in E. coli). Furthermore, several ribosomal proteins encoded by the liverwort mitochondrial genome differed substantially in size from their counterparts in E. coli and liverwort chloroplast.     

Structure and evolution of junctions between inverted repeat and small single copy regions of chloroplast genome in non-core Caryophyllales

The structure of the junction between inverted repeat (IR) and small single copy (SSC) regions of the chloroplast genome in the representatives of non-core Caryophyllales is investigated in this work. It was found that for two families—Polygonaceae and Plumbaginaceae—the extension of inverted region is characteristic. This extension is due to the duplication of the part of the ycf1 gene that is partly located in the small single copy region in plants with typical structure of IR/SSC junctions. Comparison of the position of IR/SSC junctions in different species of Polygonaceae has shown that their exact position is not correlated with the affinity of these species inferred from molecular and morphological data. Possible mechanisms leading to the change in position of IR/SSC junctions observed in this work are discussed.     

Composition and physiological function of the chloroplast NADH dehydrogenase‐like complex in Marchantia polymorpha

The chloroplast NADH dehydrogenase‐like (NDH) complex mediates cyclic electron transport and chloro‐respiration and consists of five sub‐omplexes, which in angiosperms further associate with photosystem I (PSI) to form a super‐complex. In Marchantia polymorpha, 11 plastid‐encoded subunits and all the nuclear‐encoded subunits of the A, B, membrane and ferredoxin‐binding sub‐complexes are conserved. However, it is unlikely that the genome of this liverwort encodes Lhca5 and Lhca6, both of which mediate NDH–PSI super‐complex formation. It is also unlikely that the subunits of the lumen sub‐complex, PnsL1–L4, are encoded by the genome. Consistent with this in silico prediction, the results of blue‐native gel electrophoresis showed that NDH subunits were detected in a protein complex with lower molecular mass in Marchantia than the NDH–PSI super‐complex in Arabidopsis. Using the plastid transformation technique, we knocked out the ndhB gene in Marchantia. Although the wild‐type genome copies were completely segregated out, the ΔndhB lines grew like the wild‐type photoautotrophically. A post‐illumination transient increase in chlorophyll fluorescence, which reflects NDH activity in vivo in angiosperms, was absent in the thalli of the ΔndhB lines. In ruptured chloroplasts, antimycin A‐insensitive, and ferredoxin‐dependent plastoquinone reduction was impaired, suggesting that chloroplast NDH mediates similar electron transport in Marchantia and Arabidopsis, despite its possible difference in structure. As in angiosperms, linear electron transport was not strongly affected in the ΔndhB lines. However, the plastoquinone pool was slightly more reduced at low light intensity, suggesting that chloroplast NDH functions in redox balancing of the inter system, especially under low light conditions.     

Transfer RNA genes in the mitochondrial genome from a liverwort, Marchantia polymorpha: the absence of chloroplast-like tRNAs.

Twenty-nine genes for 27 species of tRNAs were deduced from the complete nucleotide sequence of the mitochondrial genome from a liverwort, Marchantia polymorpha. One to three species of tRNA genes corresponded to each of 20 amino acids including three species for leucine and arginine, two species for serine and glycine, and one for the rest of the amino acids. Interestingly, all tRNA genes were located in the semicircle of the liverwort mitochondrial genome except for the trnY and trnR genes. The region containing these tRNA genes was originally duplicated, and two trnR genes have diverged from each other. On the other hand, trnY and trnfM are present as two identical copies. The G:U and U:N wobbling between the first nucleotide of the anticodon and the third nucleotide of the codon permit the 27 tRNA identified species to translate almost all codons. However, at least two additional tRNA genes, trnl-GAU for AUY codon and trnT-UGU for ACR codon, are required to read all codons used in the liverwort mitochondrial genome. All of the identified tRNA genes are 'native' in liverwort mitochondria, not 'chloroplast-like' tRNAs as are found in the mitochondria of higher plants. This result implies that the tRNA gene transfer from chloroplast to mitochondrial genome in higher plants has occurred after the divergence from bryophytes.     

Nucleotide sequences of chloroplast 5S ribosomal RNA from cell suspension cultures of the liverworts Marchantia polymorpha and Jungermannia subulata

The nucleotide sequences of chloroplast 5S rRNAs from cell suspension cultures of the liverworts Marchantia polymorpha and Jungermannia subulata were determined. Their nucleotide sequences, 119 nucleotides long, were highly homologous to each other (96% identity) and had high homology with those from chloroplast 5S rRNAs of two higher plants, tobacco (92% identity) and spinach (92-91% identity), but less homology (87-85% identity) with that from a lower plant, the fern Dryopteris acuminata.     

Associations between light-harvesting complexes and Photosystem II from Marchantia polymorpha L. determined by two- and three-dimensional electron microscopy

Assemblies of Photosystem II and light-harvesting proteins were purified from the liverwort Marchantia polymorpha and investigated by two- and three-dimensional transmission electron microscopy of negatively stained specimens. By single-particle analysis, it was determined that about 25% of the particles are rectangular or slightly S-shaped with dimensions of 285 A in length, 144 A in width, 84 A in height, while the membrane part is about 52 A thick. This structure reveals the same architecture as that of a Photosystem II-light-harvesting assembly from seed plants. An overlay of the projection structure of the liverwort's complex with a projection structure deduced from stained trimeric LHC II crystals from pea confirmed the locations of trimeric LHC II within the liverwort's complex. Remarkably tight associations of LHC II and other chlorophyll a/b binding proteins with the PS II core complex are observed. More than 50% of the Photosystem II particles from the liverwort carry one or two additional masses. These extra masses are found to consist of an additional LHC II trimer and probably a chlorophyll a/b binding protein. For the first time, a three-dimensional structure of such a large assembly is defined.     

Nucleotide sequence of Marchantia polymorpha chloroplast DNA: a region possibly encoding three tRNAs and three proteins including a homologue of E. coli ribosomal protein S14.

The nucleotide sequence of a region of Marchantia polymorpha chloroplast DNA was determined. On this DNA sequence (3.38kb), three open reading frames (ORFs) and three putative tRNA genes were detected in the following order: -ORF701-tRNASer(UGA)-ORF702-tRNAGly(GCC)-initiator tRNAMet(CAU)-ORF703-. The ORF703 is composed of 100 codons in which those for lysine (15%) and arginine (11%) are abundant, and could be accounted for as a counterpart of E. coli ribosomal protein S14 since they share 45% homology in the amino acid sequences. The ORF701 appears to code for a membrane protein, showing a periodic appearance of seven clusters of hydrophobic amino acids. Although the mechanisms remain unknown, the ORF701 causes a streptomycin-sensitive phenotype in resistant mutants of E. coli. The ORFs and tRNA genes are separated from each other by extremely AT-rich spacers containing sequences of dyad symmetry. The third letter positions of the codons in the ORFs are also rich in A and T residues.     

地钱生殖托形态结构新观察及其对有性生殖的适应

用显微观察技术对苔类植物地钱（Marchantia polymorpha）生殖托的形态结构进行了研究,本研究的新观察包括：（1）生殖托具有明显的背腹分化,被认为是叶状体为适应有性生殖而高度特化的直立枝;（2）雌托盘的9～11个指状裂瓣中边缘两个稍不同,除了两个边缘裂瓣外的其他裂瓣间具雌苞。托柄具有背腹之分,雌托柄背面具光合组织,并向两侧扩展形成纵沟,雌托柄的腹面具2条被鳞片重叠覆盖相互平行的纵沟,内具假根;（3）与雌托柄不同,雄托柄外观平滑,背面无光合组织及纵沟。观察发现,伞状的雌托能滞留水分,并沿雌托柄的纵沟缓慢释放连续的水流,游动精子沿着纵沟内的水流到达雌托下面的颈卵器。上述观察表明地钱生殖托的结构是对有性生殖的一种适应,这有助于我们理解地钱受精作用的机理。     

Biochemical Characterization of Casein Kinases II (CK-II) from Cultured Cells of the Liverwort, Marchantia polymorpha

Monomeric and oligomeric forms of CK-II have been purified froma 1.0 M KC1 extract of the liverwort, Marchantia polymorpha,by means of heparin-agarose column chromatography and gel filtrationon Superose 6HR (HPLC). It was found that (i) a monomeric kinase(approximately 38 kDa) is the main form of CK-II in the cells;and (ii) the enzymatic properties of oligomeric kinase (approximately140 kDa), which cross-reacts with anti-serum against DrosophilaCK-IIß, are similar to those of CK-II (₂ß₂)in various animal cells. (Received November 10, 1992; Accepted February 18, 1993)     

Identification and characterization of photosystem II chlorophyll a/b binding proteins in Marchantia polymorpha L.

The minor chlorophyll a/b-binding (CAB) proteins of the liverwort Marchantia polymorpha L. were investigated in order to compare the antenna organization and the light-acclimation potential in lower and higher plants. Homologues to the minor CAB proteins CP24, CP26 and CP29 were identified by the following criteria: enrichment in photosystem II preparations, immunological cross-reactivities, spectroscopic properties and protein-fragment amino acid sequences. The high violaxanthin content of the minor CAB proteins in M. polymorpha indicates that their role in protection from high light is comparable in lower and higher plants. Considerably more-alkaline isoelectric points are found for the minor CAB proteins of M. polymorpha than for their higher-plant counterparts. This might be due to a higher content of basic amino acids. While the N-terminal sequence of angiosperm CP29 contains a threonine that becomes phosphorylated during cold stress, this amino acid is substituted by valine in M. polymorpha. Therefore, the regulatory properties of this protein could differ in lower and higher plants. Received: 25 March 1997 / Accepted: 21 July 1997