Partenskyella glossopodia (Chlorarachniophyceae) possesses a nucleomorph genome of approximately 1 Mbp

The nucleomorph genome size of the recently described chlorarachniophyte Partenskyella glossopodia, which forms an independent lineage in the phylogeny of chlorarachniophytes, was analyzed by pulse‐field gel electrophoresis and Southern hybridization. These analyses showed that the nucleomorph genome of P. glossopodia is composed of three linear chromosomes that are about 445 kbp, 313 kbp, and 275 kbp in size. Thus, the total genome size is approximately 1033 kbp, which is significantly larger than the known size of chlorarachniophyte nucleomorph genomes, i.e. 330–610 kbp. This is the first study to report a nucleomorph genome that reaches approximately 1 Mbp in size.     

Nucleomorph ribosomal DNA and telomere dynamics in chlorarachniophyte algae

Chlorarachniophytes are enigmatic marine unicellular algae that acquired photosynthesis by secondary endosymbiosis. Chlorarachniophytes are unusual in that the nucleus of the engulfed algal cell (a green alga) persists in a miniaturized form, termed a nucleomorph. The nucleomorph genome of the model chlorarachniophyte, Bigelowiella natans CCMP621, is 373 kilobase pairs (kbp) in size, the smallest nuclear genome characterized to date. The B. natans nucleomorph genome is composed of three chromosomes, each with canonical eukaryotic telomeres and sub-telomeric ribosomal DNA (rDNA) operons transcribed away from the chromosome end. Here we present the complete rDNA operon and telomeric region from the nucleomorph genome of Lotharella oceanica CCMP622, a newly characterized chlorarachniophyte strain with a genome ～610 kbp in size, significantly larger than all other known chlorarachniophytes. We show that the L. oceanica rDNA operon is in the opposite chromosomal orientation to that of B. natans. Furthermore, we determined the rDNA operon orientation of five additional chlorarachniophyte strains, the majority of which possess the same arrangement as L. oceanica, with the exception of Chlorarachnion reptans and those very closely related to B. natans. It is thus possible that the ancestral rDNA operon orientation of the chlorarachniophyte nucleomorph genome might have been the same as in the independently evolved, red algal-derived, nucleomorph genomes of cryptophytes. A U2 small nuclear RNA gene was found adjacent to the telomere in Gymnochlora stellata CCMP2057 and Chlorarachnion sp. CCMP2014. This feature may represent a useful evolutionary character for inferring the relationships among extant lineages.     

Nucleomorph and plastid genome sequences of the chlorarachniophyte Lotharella oceanica: convergent reductive evolution and frequent recombination in nucleomorph-bearing algae

Background

Nucleomorphs are residual nuclei derived from eukaryotic endosymbionts in chlorarachniophyte and cryptophyte algae. The endosymbionts that gave rise to nucleomorphs and plastids in these two algal groups were green and red algae, respectively. Despite their independent origin, the chlorarachniophyte and cryptophyte nucleomorph genomes share similar genomic features such as extreme size reduction and a three-chromosome architecture. This suggests that similar reductive evolutionary forces have acted to shape the nucleomorph genomes in the two groups. Thus far, however, only a single chlorarachniophyte nucleomorph and plastid genome has been sequenced, making broad evolutionary inferences within the chlorarachniophytes and between chlorarachniophytes and cryptophytes difficult. We have sequenced the nucleomorph and plastid genomes of the chlorarachniophyte Lotharella oceanica in order to gain insight into nucleomorph and plastid genome diversity and evolution.

Results

The L. oceanica nucleomorph genome was found to consist of three linear chromosomes totaling ~610 kilobase pairs (kbp), much larger than the 373 kbp nucleomorph genome of the model chlorarachniophyte Bigelowiella natans. The L. oceanica plastid genome is 71 kbp in size, similar to that of B. natans. Unexpectedly long (~35 kbp) sub-telomeric repeat regions were identified in the L. oceanica nucleomorph genome; internal multi-copy regions were also detected. Gene content analyses revealed that nucleomorph house-keeping genes and spliceosomal intron positions are well conserved between the L. oceanica and B. natans nucleomorph genomes. More broadly, gene retention patterns were found to be similar between nucleomorph genomes in chlorarachniophytes and cryptophytes. Chlorarachniophyte plastid genomes showed near identical protein coding gene complements as well as a high level of synteny.

Conclusions

We have provided insight into the process of nucleomorph genome evolution by elucidating the fine-scale dynamics of sub-telomeric repeat regions. Homologous recombination at the chromosome ends appears to be frequent, serving to expand and contract nucleomorph genome size. The main factor influencing nucleomorph genome size variation between different chlorarachniophyte species appears to be expansion-contraction of these telomere-associated repeats rather than changes in the number of unique protein coding genes. The dynamic nature of chlorarachniophyte nucleomorph genomes lies in stark contrast to their plastid genomes, which appear to be highly stable in terms of gene content and synteny.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-374) contains supplementary material, which is available to authorized users.     

Nucleomorph genome diversity and its phylogenetic implications in cryptomonad algae

The relationship between phylogeny and nucleomorph genome size was examined in 16 strains of cryptomonad algae using pulsed‐field gel electrophoresis, Southern hybridization and phylogenetic analyses. Our results suggest that all cryptomonads examined in this study contain three nucleomorph chromosomes and their total genome size ranges from 495 to 750 kb. In addition, we estimated the plastid genome size of the respective organisms. The plastid genomes of photosynthetic strains were approximately 120–160 kb in size, whereas the non‐photosynthetic Cryptomonas paramecium NIES715 possesses a genome of approximately 70 kb. Phylogenetic analysis of the nuclear small subunit ribosomal DNA (SSU rDNA) gene showed that nucleomorph genome size varies considerably within closely related strains. This result indicates that the reduction of nucleomorph genomes is a rapid phenomenon that occurred multiple times independently during cryptomonad evolution. The nucleomorph genome sizes of Cryptomonas rostratiformis NIES277 appeared to be approximately 495 kb. This is smaller than that of Guillardia theta CCMP327, which until now was thought to have the smallest known nucleomorph genome size among photosynthetic cryptomonads.     

Proceedings of the SMBE Tri-National Young Investigators' Workshop 2005. Insight into the diversity and evolution of the cryptomonad nucleomorph genome

The cryptomonads are an enigmatic group of marine and freshwater unicellular algae that acquired their plastids through the engulfment and retention of a eukaryotic ("secondary") endosymbiont. Together with the chlorarachniophyte algae, the cryptomonads are unusual in that they have retained the nucleus of their endosymbiont in a miniaturized form called a nucleomorph. The nucleomorph genome of the cryptomonad Guillardia theta has been completely sequenced and with only three chromosomes and a total size of 551 kb, is a model of nuclear genome compaction. Using this genome as a reference, we have investigated the structure and content of nucleomorph genomes in a wide range of cryptomonad algae. In this study, we have sequenced nine new cryptomonad nucleomorph 18S ribosomal DNA (rDNA) genes and four heat shock protein 90 (hsp90) gene fragments, and using pulsed-field gel electrophoresis and Southern hybridizations, have obtained nucleomorph genome size estimates for nine different species. We also used long-range polymerase chain reaction to obtain nucleomorph genomic fragments from Hanusia phi CCMP325 and Proteomonas sulcata CCMP704 that are syntenic with the subtelomeric region of nucleomorph chromosome I in G. theta. Our results indicate that (1) the presence of three chromosomes is a common feature of the nucleomorph genomes of these organisms, (2) nucleomorph genome size varies dramatically in the cryptomonads examined, (3) unidentified cryptomonad species CCMP1178 has the largest nucleomorph genome identified to date at approximately 845 kb, (4) nucleomorph genome size reductions appear to have occurred multiple times independently during cryptomonad evolution, (5) the relative positions of the 18S rDNA, ubc4, and hsp90 genes are conserved in three different cryptomonad genera, and (6) interchromosomal recombination appears to be rapidly changing the size and sequence of a repetitive subtelomeric region of the nucleomorph genome between the 18S rDNA and ubc4 loci. These results provide a glimpse into the genetic diversity of nucleomorph genomes in cryptomonads and set the stage for more comprehensive sequence-based studies in closely and distantly related taxa.     

NUCLEOMORPH KARYOTYPE DIVERSITY IN THE FRESHWATER CRYPTOPHYTE GENUS CRYPTOMONAS1

Cryptophytes are unicellular, biflagellate algae with plastids (chloroplasts) derived from the uptake of a red algal endosymbiont. These organisms are unusual in that the nucleus of the engulfed red alga persists in a highly reduced form called a nucleomorph. Nucleomorph genomes are remarkable in their small size (<1,000 kilobase pairs [kbp]) and high degree of compaction (～1 kbp per gene). Here, we investigated the molecular and karyotypic diversity of nucleomorph genomes in members of the genus Cryptomonas. 18S rDNA genes were amplified, sequenced, and analyzed from C. tetrapyrenoidosa Skuja CCAP979/63, C. erosa Ehrenb. emmend. Hoef‐Emden CCAP979/67, Cryptomonas sp. CCAP979/52, C. lundii Hoef‐Emden et Melkonian CCAP979/69, and C. lucens Skuja CCAP979/35 in the context of a large set of publicly available nucleomorph 18S rDNA sequences. Pulsed‐field gel electrophoresis (PFGE) was used to examine the nucleomorph genome karyotype of each of these strains. Individual chromosomes ranged from ～160 to 280 kbp in size, with total genome sizes estimated to be ～600–655 kbp. Unexpectedly, the nucleomorph karyotype of Cryptomonas sp. CCAP979/52 is significantly different from that of C. tetrapyrenoidosa and C. lucens, despite the fact that their 18S rDNA genes are >99% identical to one another. These results suggest that nucleomorph karyotype similarity is not a reliable indicator of evolutionary affinity and provides a starting point for further investigation of the fine‐scale dynamics of nucleomorph genome evolution within members of the genus Cryptomonas.     

Novel nucleomorph genome architecture in the cryptomonad genus hemiselmis

Cryptomonads are ubiquitous aquatic unicellular eukaryotes that acquired photosynthesis through the uptake and retention of a red algal endosymbiont. The nuclear genome of the red alga persists in a highly reduced form termed a nucleomorph. The nucleomorph genome of the model cryptomonad Guillardia theta has been completely sequenced and is a mere 551 kilobases (kb) in size, spread over three chromosomes. The presence of three chromosomes appears to be a universal characteristic of nucleomorph genomes in cryptomonad algae as well as in the chlorarachniophytes, an unrelated algal lineage with a nucleomorph and plastid genome derived from a green algal endosymbiont. Another feature of nucleomorph genomes in all cryptomonads and chlorarachniophytes examined thus far is the presence of subtelomeric ribosomal DNA (rDNA) repeats at the ends of each chromosome. Here we describe the first exception to this canonical nucleomorph genome architecture in the cryptomonad Hemiselmis rufescens CCMP644. Using pulsed-field gel electrophoresis (PFGE), we estimate the size of the H. rufescens nucleomorph genome to be approximately 580 kb, slightly larger than the G. theta genome. Unlike the situation in G. theta and all other known cryptomonads, sub-telomeric repeats of the rDNA cistron appear to be absent on both ends of the second largest chromosome in H. rufescens and two other members of this genus. Southern hybridizations using a variety of nucleomorph protein gene probes against PFGE-separated H. rufescens chromosomes indicate that recombination has been a major factor in shaping the karyotype and genomic structure of cryptomonad nucleomorphs.     

Nucleomorph genomes: structure, function, origin and evolution

The cryptomonads and chlorarachniophytes are two unicellular algal lineages with complex cellular structures and fascinating evolutionary histories. Both groups acquired their photosynthetic abilities through the assimilation of eukaryotic endosymbionts. As a result, they possess two distinct cytosolic compartments and four genomes--two nuclear genomes, an endosymbiont-derived plastid genome and a mitochondrial genome derived from the host cell. Like mitochondrial and plastid genomes, the genome of the endosymbiont nucleus, or 'nucleomorph', of cryptomonad and chlorarachniophyte cells has been greatly reduced through the combined effects of gene loss and intracellular gene transfer. This article focuses on the structure, function, origin and evolution of cryptomonad and chlorarachniophyte nucleomorph genomes in light of recent comparisons of genome sequence data from both groups. It is now possible to speculate on the reasons that nucleomorphs persist in cryptomonads and chlorarachniophytes but have been lost in all other algae with plastids of secondary endosymbiotic origin.     

Molecular, morphological and phylogenetic characterization of six chlorarachniophyte strains

We compare and contrast the morphological and molecular features of six chlorarachniophyte strains, and examine their evolutionary origins. Electron microscopical studies of nucleomorphs and chloroplasts, characterization of nucleomorph karyotypes, and phylogenetic analyses of small subunit ribosomal RNA (srRNA) genes derived from the nucleomorph and host cell genomes have been used to separate the six strains into three distinct groups. One group, dubbed the‘beast group’, contains the strains Chlorarachnion sp. 242, Chlor-arachnion sp. 621, Chlorarachnion sp. 1408 and Chlorarachnion sp. 1481. Members of the beast group have a novel flagellate form and are apparently picoplank-tonic. The other two groups currently contain only one species each: Chlorarachnion reptans and Lotharella sp. 240. All chlorarachniophyte nucleomorphs examined house three small linear chromosomes each furnished with telomeres and srRNA genes.     

Lotharella reticulosa sp. nov.: a highly reticulated network forming chlorarachniophyte from the Mediterranean Sea

A new chlorarachniophyte Lotharella reticulosa sp. nov. is described from a culture isolated from the Mediterranean Sea. This strain is maintained as strain RCC375 at the Roscoff Culture Collection, France. This species presents a multiphasic life cycle: vegetative cells of this species were observed to be coccoid, but amoeboid cells with filopodia and globular suspended cells were also present in the life cycle, both of which were not dominant phases. Flagellate cells were also observed but remained very rare in culture. The vegetative cells were 9-16 μm in diameter and highly vacuolated, containing several green chloroplasts with a projecting pyrenoid, mitochondria, and a nucleus. The chloroplast was surrounded by four membranes possessing a nucleomorph in the periplastidial compartment near the pyrenoid base. According to ultrastructural observations of the pyrenoid and nucleomorph, the present species belongs to the genus Lotharella in the phylum Chlorarachniophyta. This taxonomic placement is consistent with the molecular phylogenetic trees of the 18S rRNA gene and ITS sequences. This species showed a unique colonization pattern. Clusters of cells extended cytoplasmic strands radially. Then, amoeboid cells being born proximately moved distally along the cytoplasmic strand like on a "railway track". Subsequently the amoeboid cell became coccoid near the strand. In this way, daughter cells were dispersed evenly on the substratum. We also observed that the present species regularly formed a structure of filopodial nodes in mid-stage and later-stage cultures, which is a novel phenotype in chlorarachniophytes. The unique colonization pattern and other unique features demonstrate that RCC375 is a new chlorarachniophyte belonging to genus Lotharella, which we describe as Lotharella reticulosa sp. nov.     

Nucleus- and nucleomorph-targeted histone proteins in a chlorarachniophyte alga

The plastid of chlorarachniophytes is distinguished by the retention of a relict nucleus (nucleomorph) derived from a green algal endosymbiont, which is located in the periplastidal compartment (PPC). The nucleomorph genome of a chlorarachniophyte, Bigelowiella natans, encodes several plastid-targeted proteins and hundreds of housekeeping proteins, but it lacks many fundamental genes to maintain itself. Here we report the first two host nucleus-encoded genes for proteins targeted to the nucleomorph, histone H2A and H2B. We identified 20 histone genes from the host nuclear genome, and based on phylogenetic analyses predicted that most of these are derived from the host, but that two histone genes are symbiont-derived. The genes both encode N-terminal extensions resembling PPC targeting signals, further suggesting they function in the nucleomorph. Using green fluorescent protein (GFP) fusion proteins expressed in transformed cells, we confirmed that the putative symbiont H2A and H2B were targeted into the nucleomorph, whereas putative host proteins were localized to the host nucleus. Furthermore, we have developed a method to temporarily synchronize B. natans cells, and confirmed that both host and symbiont histone expression is controlled during the cell cycle. Our findings provide the first evidence of how the nucleomorph may be regulated by host-encoded gene products.     

DNA BARCODING OF CHLORARACHNIOPHYTES USING NUCLEOMORPH ITS SEQUENCES1

Chlorarachniophytes are a small group of marine photosynthetic protists. They are best known as examples of an intermediate stage of secondary endosymbiosis: their plastids are derived from green algae and retain a highly reduced nucleus, called a nucleomorph, between the inner and outer pairs of membranes. Chlorarachniophytes can be challenging to identify to the species level, due to their small size, complex life cycles, and the fact that even genus‐level diagnostic morphological characters are observable only by EM. Few species have been formally described, and many available culture collection strains remain unnamed. To alleviate this difficulty, we have developed a barcoding system for rapid and accurate identification of chlorarachniophyte species in culture, based on the internal transcribed spacer (ITS) region of the nucleomorph rRNA cistron. Although this is a multicopy locus, encoded in both subtelomeric regions of each chromosome, interlocus variability is low due to gene conversion by homologous recombination in this region. Here, we present barcode sequences for 39 cultured strains of chlorarachniophytes (>80% of currently available strains). Based on barcode data, other published molecular data, and information from culture records, we were able to recommend names for 21 out of the 24 unidentified, partially identified, or misidentified chlorarachniophyte strains in culture. Most strains could be assigned to previously described species, but at least two to as many as five new species may be present among cultured strains.     

Variation of osteocyte lacunae size within the tetrapod skeleton: implications for palaeogenomics

Recent studies have emphasized the ability to reconstruct genome sizes (C-values) of extinct organisms such as dinosaurs, using correlations between known genome sizes and bone cell (osteocyte lacunae) volumes. Because of the established positive relationship between cell size and genome size in extant vertebrates, osteocyte lacunae volume is a viable proxy for reconstructing C-values in the absence of any viable genetic material. However, intra-skeletal osteocyte lacunae size variation, which could cause error in genome size estimation, has remained unexplored. Here, 11 skeletal elements of one individual from each of four major clades (Mammalia, Amphibia, Aves, Reptilia) were examined histologically. Skeletal elements in all four clades exhibit significant differences in the average sizes of their lacunae. This variation, however, generally does not cause a significant difference in the estimated genome size when common phylogenetic estimation methods are employed. On the other hand, the spread of the estimations illustrates that this method may not be precise. High variance in genome size estimations remains an outstanding problem. Additionally, a suite of new methods is introduced to further automate the measurement of bone cells and other microstructural features on histological thin sections.     

Five major nuclear ribosomal repeats represent a large and variable fraction of the genomic DNA of Picea rubens and P. mariana.

The nuclear ribosomal repeats for the 18S, 5.8S, and 26S RNAs of two closely related Picea (spruce) species were characterized by restriction mapping and Southern blot hybridization. Restriction polymorphisms were identified in the IGS and ITS sequences; however, no polymorphism was species specific. As many as five different rDNA repeat units were observed in individual genomes. The repeat size for these gymnosperms ranged from a minimum of 32 kbp to greater than 40 kbp, two- to threefold larger than the typical angiosperm rDNA unit. Slot-blot hybridizations were used to determine the nuclear rDNA copy concentration. Among P. rubens individuals threefold variation was observed in the rDNA copy concentration, and among P. mariana individuals such variation was as much as sixfold. At a size greater than 32 kbp and at a concentration averaging 1.2-1.3 x 10(4) copies/pg, the rDNA constitutes approximately 4% of the total genome. Regression analysis revealed a significant relationship between copy concentration of the rDNA repeat unit in P. rubens and geographic origins. Differences in the rDNA content in Picea could contribute to the variation, in overall genome size, that has been observed within conifer species.     

Eukaryotic non-coding DNA is functional: evidence from the differential scaling of cryptomonad genomes

Genic DNA functions are commonplace: coding for proteins and specifying non-messenger RNA structure. Yet most DNA in the biosphere is non-genic, existing in nuclei as non-coding or secondary DNA. Why so much secondary DNA exists and why its amount per genome varies over orders of magnitude (correlating positively with cell volume) are central biological problems. A novel perspective on secondary DNA function comes from natural eukaryote eukaryote chimaeras (cryptomonads and chlorarachneans) where two phylogenetically distinct nuclei have coevolved within one cell for hundreds of millions of years. By comparing cryptomonad species differing 13-fold in cell volume, we show that nuclear and nucleomorph genome sizes obey fundamentally different scaling laws. Following a more than 125-fold reduction in DNA content, nucleomorph genomes exhibit little variation in size. Furthermore, the present lack of significant amounts of nucleomorph secondary DNA confirms that selection can readily eliminate functionless nuclear DNA, refuting 'selfish' and 'junk' theories of secondary DNA. Cryptomonad nuclear DNA content varied 12-fold: as in other eukaryotes, larger cells have extra DNA, which is almost certainly secondary DNA positively selected for a volume-related function. The skeletal DNA theory explains why nuclear genome size increases with cell volume and, using new evidence on nucleomorph gene functions, why nucleomorph genomes do not.     

Evidence for nucleomorph to host nucleus gene transfer: light-harvesting complex proteins from cryptomonads and chlorarachniophytes

Cryptomonads and chlorarachniophytes acquired photosynthesis independently by engulfing and retaining eukaryotic algal cells. The nucleus of the engulfed cells (known as a nucleomorph) is much reduced and encodes only a handful of the numerous essential plastid proteins normally encoded by the nucleus of chloroplast-containing organisms. In cryptomonads and chlorarachniophytes these proteins are thought to be encoded by genes in the secondary host nucleus. Genes for these proteins were potentially transferred from the nucleomorph (symbiont nucleus) to the secondary host nucleus; nucleus to nucleus intracellular gene transfers. We isolated complementary DNA clones (cDNAs) for chlorophyll-binding proteins from a cryptomonad and a chlorarachniophyte. In each organism these genes reside in the secondary host nuclei, but phylogenetic evidence, and analysis of the targeting mechanisms, suggest the genes were initially in the respective nucleomorphs (symbiont nuclei). Implications for origins of secondary endosymbiotic algae are discussed.     

Evolution of genome size in Brassicaceae

BACKGROUND AND AIMS: Brassicaceae, with nearly 340 genera and more than 3350 species, anchors the low range of angiosperm genome sizes. The relatively narrow range of DNA content (0.16 pg < 1C < 1.95 pg) was maintained in spite of extensive chromosomal change. The aim of this study was to erect a cytological and molecular phylogenetic framework for a selected subset of the Brassicacae, and use this as a template to examine genome size evolution in Brassicaceae. METHODS: DNA contents were determined by flow cytometry and chromosomes were counted for 34 species of the family Brassicaceae and for ten Arabidopsis thaliana ecotypes. The amplified and sequenced ITS region for 23 taxa (plus six other taxa with known ITS sequences) were aligned and used to infer evolutionary relationship by parsimony analysis. KEY RESULTS: DNA content in the species studied ranged over 8-fold (1C = 0.16-1.31 pg), and 4.4-fold (1C = 0.16-0.71 pg) excluding allotetraploid Brassica species. The 1C DNA contents of ten Arabidopsis thaliana ecotypes showed little variation, ranging from 0.16 pg to 0.17 pg. CONCLUSIONS: The tree roots at an ancestral genome size of approximately 1x = 0.2 pg. Arabidopsis thaliana (1C = 0.16 pg; approximately 157 Mbp) has the smallest genome size in Brassicaceae studied here and apparently represents an evolutionary decrease in genome size. Two other branches that represent probable evolutionary decreases in genome size terminate in Lepidium virginicum and Brassica rapa. Branches in the phylogenetic tree that represent probable evolutionary increases in genome size terminate in Arabidopsis halleri, A. lyrata, Arabis hirsuta, Capsella rubella, Caulanthus heterophyllus, Crucihimalaya, Lepidium sativum, Sisymbrium and Thlaspi arvense. Branches within one clade containing Brassica were identified that represent two ancient ploidy events (2x to 4x and 4x to 6x) that were predicted from published comparative mapping studies.     

Genome-based reconstruction of the protein import machinery in the secondary plastid of a chlorarachniophyte alga

Most plastid proteins are encoded by their nuclear genomes and need to be targeted across multiple envelope membranes. In vascular plants, the translocons at the outer and inner envelope membranes of chloroplasts (TOC and TIC, respectively) facilitate transport across the two plastid membranes. In contrast, several algal groups harbor more complex plastids, the so-called secondary plastids, which are surrounded by three or four membranes, but the plastid protein import machinery (in particular, how proteins cross the membrane corresponding to the secondary endosymbiont plasma membrane) remains unexplored in many of these algae. To reconstruct the putative protein import machinery of a secondary plastid, we used the chlorarachniophyte alga Bigelowiella natans, whose plastid is bounded by four membranes and still possesses a relict nucleus of a green algal endosymbiont (the nucleomorph) in the intermembrane space. We identified nine homologs of plant-like TOC/TIC components in the recently sequenced B. natans nuclear genome, adding to the two that remain in the nucleomorph genome (B. natans TOC75 [BnTOC75] and BnTIC20). All of these proteins were predicted to be localized to the plastid and might function in the inner two membranes. We also show that the homologs of a protein, Der1, that is known to mediate transport across the second membrane in the several lineages with secondary plastids of red algal origin is not associated with plastid protein targeting in B. natans. How plastid proteins cross this membrane remains a mystery, but it is clear that the protein transport machinery of chlorarachniophyte plastids differs from that of red algal secondary plastids.     

Phylogenetic clusters of rhizobia revealed by genome structures

Rhizobia, bacteria that fix atmospheric nitrogen, are important agricultural resources. In order to establish the evolutionary relationships among rhizobia isolated from different geographic regions and different plant hosts for systematic studies, we evaluated the use of physical structure of the rhizobial genomes as a phylogenetic marker to categorize these bacteria. In this work, we analyzed the features of genome structures of 64 rhizobial strains. These rhizobial strains were divided into 21 phylogenetic clusters according to the features of genome structures evaluated by the endonuclease I-Ceul. These clusters were supported by 16S rRNA comparisons and genomic sequences of four rhizobial strains, but they are largely different from those based on the current taxonomic scheme (except 16S rRNA).     

The considerable genome size variation of Hordeum species (poaceae) is linked to phylogeny, life form, ecology, and speciation rates

Genome size variation in plants is thought to be correlatedwith cytological, physiological, or ecological characters. However,conclusions drawn in several studies were often contradictory.To analyze nuclear genome size evolution in a phylogenetic framework,DNA contents of 134 accessions, representing all but one speciesof the barley genus Hordeum L., were measured by flow cytometry.The 2C DNA contents were in a range from 6.85 to 10.67 pg indiploids (2n = 14) and reached up to 29.85 pg in hexaploid species(2n = 42). The smallest genomes were found in taxa from theNew World, which became secondarily annual, whereas the largestdiploid genomes occur in Eurasian annuals. Genome sizes of polyploidtaxa equaled mostly the added sizes of their proposed progenitorsor were slightly (1% to 5%) smaller. The analysis of ancestralgenome sizes on the base of the phylogeny of the genus revealedlineages with decreasing and with increasing genome sizes. Correlationsof intraspecific genome size variation with the length of vegetationperiod were found in H. marinum populations from Western Europebut were not significant within two species from South America.On a higher taxonomical level (i.e., for species groups or theentire genus), environmental correlations were absent. Thiscould mostly be attributed to the superimposition of life-formchanges and phylogenetic constraints, which conceal ecogeographicalcorrelations.