Serpins in plants and green algae

Control of proteolysis is important for plant growth, development, responses to stress, and defence against insects and pathogens. Members of the serpin protein family are likely to play a critical role in this control through irreversible inhibition of endogenous and exogenous target proteinases. Serpins have been found in diverse species of the plant kingdom and represent a distinct clade among serpins in multicellular organisms. Serpins are also found in green algae, but the evolutionary relationship between these serpins and those of plants remains unknown. Plant serpins are potent inhibitors of mammalian serine proteinases of the chymotrypsin family in vitro but, intriguingly, plants and green algae lack endogenous members of this proteinase family, the most common targets for animal serpins. An Arabidopsis serpin with a conserved reactive centre is now known to be capable of inhibiting an endogenous cysteine proteinase. Here, knowledge of plant serpins in terms of sequence diversity, inhibitory specificity, gene expression and function is reviewed. This was advanced through a phylogenetic analysis of amino acid sequences of expressed plant serpins, delineation of plant serpin gene structures and prediction of inhibitory specificities based on identification of reactive centres. The review is intended to encourage elucidation of plant serpin functions.     

Glycolate metabolism in green algae

Using ¹⁴C-labelled substrates, the succession of the single steps in the glycolate metabolism was investigated in Mougeotia scalaris and Eremosphaera viridis , which, within the group of green algae, are representatives of the evolutionary lines of Charophyta and Chlorophyta , respectively. In both algae the same metabolites are formed as in higher plants, although in Eremosphaera , which in contrast to Mougeotia does not possess leaf peroxisomes, all reactions are exclusively mitochondrial. Concomitant with the oxidation of glycolate, the synthesis of ATP was demonstrated in Eremosphaera . Formation of tartronic semi-aldehyde or other products different from those in land plants could not be demonstrated in either of these algae. Excretion of glycolate by Mougeotia and Eremosphaera is enhanced by decreasing the CO₂ concentration as well as by increasing the light intensity, but is completely stopped about 14 h later. Whereas increasing enzyme activities of the glycolate pathway apparently reduces glycolate excretion in Mougeotia , activation of CO₂ pumps seems to be the dominant reaction to prevent glycolate excretion in Eremosphaera . Mesostigma viride is one of the phylogenetically oldest algae in the group of Charophyceae . As this alga has already been demonstrated to contain microbodies with enzymes of leaf peroxisomes, the peroxisomal glycolate pathway must have originated at a very early stage. Surprisingly, the organelles from Mesostigma contain also the glyoxysomal marker enzyme isocitrate lyase suggesting these microbodies to be prototypes from which both glyoxysomes and leaf peroxisomes evolved.     

Chloroplast evolution in algae and land plants

Eukaryotes contain a chimeric assembly of genomes, each localized in a specialized subcellular compartment. The successful survival of an organism requires that these sequestered genomes be viewed as dependent variables in a coevolutionary complex. This discussion focuses on chloroplast evolution. A selected review of information available on chloroplast diversity is presented, followed by an analysis of the genetic modifications which may have occurred in the conversion of a free-living ancestral photosynthetic prokaryote into an organelle that has an obligately dependent and highly efficient interplay with the nuclear genome.     

Green algae to land plants: An evolutionary transition

Studies focused upon the evolutionary transition from ancestral green algae to the earliest land plants are important from a range of ecological, molecular and evolutionary perspectives. A substantial suite of ultrastructural, biochemical and molecular data supports the concept that land plants (embryophytes) are monophyletically derived from an ancestral charophycean alga. However, the details of phylogenetic branching patterns linking extant charophytes and seedless embryophytes are currently unclear. Moreover, the fossil record has so far been mute regarding the algae-land plant transition. Nevertheless, an accurate reflection of major evolutionary events in the history of the earliest land plants can be obtained by comparative paleontological-neontological studies, and comparative molecular, cellular and developmental investigations of extant charophytes and bryophytes. This review focuses upon research progress toward understanding three clade-specific adaptations that were important in the successful colonization of land by plants: the histogenetic apical meristem, the matrotrophic embryo, and decay-resistant cell wall polymers.     

Ribosomal protein L10 is encoded in the mitochondrial genome of many land plants and green algae

Background  

The mitochondrial genomes of plants generally encode 30-40 identified protein-coding genes and a large number of lineage-specific ORFs. The lack of wide conservation for most ORFs suggests they are unlikely to be functional. However, an ORF, termed orf-bryo1, was recently found to be conserved among bryophytes suggesting that it might indeed encode a functional mitochondrial protein.     

轮藻和陆地植物系统发育及其进化

Charophytic algae and land plants together make up a monophyletic group, streptophytes, which represents one of the main lineages of multicellular eukaryotes and has contributed greatly to the change of the environment on earth in the Phanerozoic Eon. Significant progress has been made to understand phylogenetic relationships among members of this group by phylogenetic studies of morphological and molecular data over the last twenty-five years. Mesostigma viride is now regarded as among the earliest diverging unicellular organisms in streptophytes. Characeae are the sister group to land plants. Liverworts represent the first diverging lineage of land plants. Hornworts and lycophytes are extant representatives of bryophytes and vascular plants, respectively, when early land plants changed from gametophyte to sporophyte as the dominant generation in the life cycle. Equisetum, Psilotaceae, and ferns constitute the monophyletic group of monilophytes, which are sister to seed plants. Gnetales are related to conifers, not to angiosperms as previously thought. Amborella, Nymphaeales, Hydatellaceae, Illiciales, Trimeniaceae, and Austrobaileya represent the earliest diverging lineages of extant angiosperms. These phylogenetic results, together with recent progress on elucidating genetic and developmental aspects of the plant life cycle, multicellularity, and gravitropism, will facilitate evolutionary developmental studies of these key traits, which will help us to gain mechanistic understanding on how plants adapted to environmental challenges when they colonized the land during one of the major transitions in evolution of life.     

Molecular genetics of xanthophyll-dependent photoprotection in green algae and plants

The involvement of excited and highly reactive intermediates in oxygenic photosynthesis inevitably results in the generation of reactive oxygen species. To protect the photosynthetic apparatus from oxidative damage, xanthophyll pigments are involved in the quenching of excited chlorophyll and reactive oxygen species, namely 1Chl*, 3Chl*, and 1O2*. Quenching of 1Chl* results in harmless dissipation of excitation energy as heat and is measured as non-photochemical quenching (NPQ) of chlorophyll fluorescence. The multiple roles of xanthophylls in photoprotection are being addressed by characterizing mutants of Chlarnydomonas reinhardtii and Arabidopsis thaliana. Analysis of Arabidopsis mutants that are defective in 1Chl* quenching has shown that, in addition to specific xanthophylls, the psbS gene is necessary for NPQ. Double mutants of Chlamydomonas and Arabidopsis that are deficient in zeaxanthin, lutein and NPQ undergo photo-oxidative bleaching in high light. Extragenic suppressors of the Chlamydomonas npq1 lor1 double mutant identify new mutations that restore varying levels of zeaxanthin accumulation and allow survival in high light.     

Molecular cloning and evolutionary analysis of the calcium-modulated contractile protein,centrin, in green algae and land plants

Centrin (= caltractin) is a ubiquitous, cytoskeletal protein which is a member of the EF-hand superfamily of calcium-binding proteins. A centrin-coding cDNA was isolated and characterized from the prasinophyte green alga Scherffelia dubia. Centrin PCR amplification primers were used to isolate partial, homologous cDNA sequences from the green algae Tetraselmis striata and Spermatozopsis similis. Annealing analyses suggested that centrin is a single-copy-coding region in T. striata and S. similis and other green algae studied. Centrin-coding regions from S. dubia, S. similis and T. striata encode four colinear EF-hand domains which putatively bind calcium. Phylogenetic analyses, including homologous sequences from Chlamydomonas reinhardtii and the land plant Atriplex nummularia, demonstrate that the domains of centrins are congruent and arose from the two-fold duplication of an ancestral EF hand with Domains 1+3 and Domains 2+4 clustering. The domains of centrins are also congruent with those of calmodulins demonstrating that, like calmodulin, centrin is an ancient protein which arose within the ancestor of all eukaryotes via gene duplication. Phylogenetic relationships inferred from centrin-coding region comparisons mirror results of small subunit ribosomal RNA sequence analyses suggesting that centrin-coding regions are useful evolutionary markers within the green algae.     

The Charophycean green algae as model systems to study plant cell walls and other evolutionary adaptations that gave rise to land plants

The Charophycean green algae (CGA) occupy a key phylogenetic position as the evolutionary grade that includes the sister group of the land plants (embryophytes), and so provide potentially valuable experimental systems to study the development and evolution of traits that were necessary for terrestrial colonization. The nature and molecular bases of such traits are still being determined, but one critical adaptation is thought to have been the evolution of a complex cell wall. Very little is known about the identity, origins and diversity of the biosynthetic machinery producing the major suites of structural polymers (i. e., cell wall polysaccharides and associated molecules) that must have been in place for land colonization. However, it has been suggested that the success of the earliest land plants was partly based on the frequency of gene duplication, and possibly whole genome duplications, during times of radical habitat changes. Orders of the CGA span early diverging taxa retaining more ancestral characters, through complex multicellular organisms with morphological characteristics resembling those of land plants. Examination of gene diversity and evolution within the CGA could help reveal when and how the molecular pathways required for synthesis of key structural polymers in land plants arose.     

POT1 proteins in green algae and land plants: DNA-binding properties and evidence of co-evolution with telomeric DNA

Telomeric DNA terminates with a single-stranded 3′ G-overhang that in vertebrates and fission yeast is bound by POT1 (Protection Of Telomeres). However, no in vitro telomeric DNA binding is associated with Arabidopsis POT1 paralogs. To further investigate POT1–DNA interaction in plants, we cloned POT1 genes from 11 plant species representing major branches of plant kingdom. Telomeric DNA binding was associated with POT1 proteins from the green alga Ostreococcus lucimarinus and two flowering plants, maize and Asparagus. Site-directed mutagenesis revealed that several residues critical for telomeric DNA recognition in vertebrates are functionally conserved in plant POT1 proteins. However, the plant proteins varied in their minimal DNA-binding sites and nucleotide recognition properties. Green alga POT1 exhibited a strong preference for the canonical plant telomere repeat sequence TTTAGGG with no detectable binding to hexanucleotide telomere repeat TTAGGG found in vertebrates and some plants, including Asparagus. In contrast, POT1 proteins from maize and Asparagus bound TTAGGG repeats with only slightly reduced affinity relative to the TTTAGGG sequence. We conclude that the nucleic acid binding site in plant POT1 proteins is evolving rapidly, and that the recent acquisition of TTAGGG telomere repeats in Asparagus appears to have co-evolved with changes in POT1 DNA sequence recognition.     

2. Alicyclic acid metabolism in plants Enzymes related to the shikimate pathway in developing mung bean seedlings

A study was made of chages in the activities of enzymes relatedto the biosynthesis of aromatic compounds in etiolated mungbean seedlings during their growth. Shikimate: NADP oxidoreductaseactivity in the root-shoot axes increased rapidly to attainits highest activity the 4th day after sowing, and remainedat that level over the experimental period of 7 days. 5-Dehydroquinatehydro-lyase activity continuously increased for at least 7 days.In the cotyledons, a gradual decrease in the activities of theseenzymes occurred. Phenylalanine ammonia-lyase activity in root-shootaxes gradually increased showing a maximum on the 6th day. Thehighest specific activity, on a protein basis, of this enzymewas seen in the initial stage of growth. In the cotyledons,a rise in total activity appeared on the 2nd day. Tyrosine ammonia-lyaseactivity was very low as compared with phenylalanine ammonia-lyase.The enzyme activities of light-germinated seedlings were comparedto those of dark-germinated seedlings on the 7th day. Lighthad practically no significant effect on the appearance of shikimate:NADP oxidoreductase and 5-dehydroquinate hydro-lyase activities.On the other hand, a marked effect from the light on the riseof phenylalanine ammonia-lyase and tyrosine ammonia-lyase activitieswas found, especially in the epicotyl-plumules. The results are discussed with respect to the metabolism ofalicyclic acids such as shikimic acid in the developing mungbean seedlings. ¹This work was partly supported by a grant-in-aid from the Ministryof Education.     

Codon usage in higher plants, green algae, and cyanobacteria

Codon usage is the selective and nonrandom use of synonymous codons by an organism to encode the amino acids in the genes for its proteins. During the last few years, a large number of plant genes have been cloned and sequenced, which now permits a meaningful comparison of codon usage in higher plants, algae, and cyanobacteria. For the nuclear and organellar genes of these organisms, a small set of preferred codons are used for encoding proteins. Codon usage is different for each genome type with the variation mainly occurring in choices between codons ending in cytidine (C) or guanosine (G) versus those ending in adenosine (A) or uridine (U). For organellar genomes, chloroplastic and mitochrondrial proteins are encoded mainly with codons ending in A or U. In most cyanobacteria and the nuclei of green algae, proteins are encoded preferentially with codons ending in C or G. Although only a few nuclear genes of higher plants have been sequenced, a clear distinction between Magnoliopsida (dicot) and Liliopsida (monocot) codon usage is evident. Dicot genes use a set of 44 preferred codons with a slight preference for codons ending in A or U. Monocot codon usage is more restricted with an average of 38 codons preferred, which are predominantly those ending in C or G. But two classes of genes can be recognized in monocots. One set of monocot genes uses codons similar to those in dicots, while the other genes are highly biased toward codons ending in C or G with a pattern similar to nuclear genes of green algae. Codon usage is discussed in relation to evolution of plants and prospects for intergenic transfer of particular genes.     

Nitrogen-assimilating enzymes in land plants and algae: phylogenic and physiological perspectives

An important biochemical feature of autotrophs, land plants and algae, is their incorporation of inorganic nitrogen, nitrate and ammonium, into the carbon skeleton. Nitrate and ammonium are converted into glutamine and glutamate to produce organic nitrogen compounds, for example proteins and nucleic acids. Ammonium is not only a preferred nitrogen source but also a key metabolite, situated at the junction between carbon metabolism and nitrogen assimilation, because nitrogen compounds can choose an alternative pathway according to the stages of their growth and environmental conditions. The enzymes involved in the reactions are nitrate reductase (EC 1.6.6.1-2), nitrite reductase (EC 1.7.7.1), glutamine synthetase (EC 6.3.1.2), glutamate synthase (EC 1.4.1.13-14, 1.4.7.1), glutamate dehydrogenase (EC 1.4.1.2-4), aspartate aminotransferase (EC 2.6.1.1), asparagine synthase (EC 6.3.5.4), and phosphoenolpyruvate carboxylase (EC 4.1.1.31). Many of these enzymes exist in multiple forms in different subcellular compartments within different organs and tissues, and play sometimes overlapping and sometimes distinctive roles. Here, we summarize the biochemical characteristics and the physiological roles of these enzymes. We also analyse the molecular evolution of glutamine synthetase, glutamate synthase and glutamate dehydrogenase, and discuss the evolutionary relationships of these three enzymes.     

The quadripolar microtubule system in lower land plants

The quadripolar microtubule system (QMS) is a complex array that is associated with predivision establishment of quadripolarity in sporocytes of lower plants (bryophytes and lycopsids). The QMS unerringly predicts the polarity of the two meiotic divisions and plays a central role in development of both the mitotic apparatus (MA) and cytokinetic apparatus (CA) which together accomplish quadripartitioning of the sporocyte into four haploid spores. The QMS is typically, but not exclusively, associated with monoplastidy and precocious quadrilobing of the cytoplasm. In early meiotic prophase the single plastid divides and the resultant plastids migrate so that either the tips of two plastids or the four plastids resulting from a second division are located in the future spore domains. Microtubules that emanate from the plastid tips or from individual plastids in the spore domains interact in the future planes of cytokinesis and give rise to the QMS. The QMS, which encages the prophase nucleus, consists of at least four and usually six (when spore domains are in tetrahedral arrangement) bipolar spindle-like arrays of microtubules presumably with minus ends at plastids in spore domains and plus ends interacting in the future plane of cytokinesis. Each of the six arrays is essentially like the single axial microtubule system (AMS) that intersects the division site and is transformed into the spindle in monoplastidic mitosis in hornworts. As comparative data accumulate, it appears that the AMS is not unique to monoplastidic cell division but instead represents a basic microtubule arrangement that survives as spindle and phragmoplast in cell division of higher plants.