Enantioselective Synthesis of α-Aminoboronic Acid Derivatives via Copper-Catalyzed N-Alkylation.

Due to burgeoning interest in the pharmaceutical industry in exploiting optically active α-aminoboronic derivatives as bioisosteres of α-amino acid derivatives, the discovery of methods for their catalytic asymmetric synthesis is an important challenge. Herein, we establish that a chiral copper catalyst (generated in situ from commercially available components) can achieve the enantioselective synthesis of α-aminoboronic derivatives via the coupling of two readily available partners, a carbamate and a racemic α-chloroboronate ester. Furthermore, we describe mechanistic studies that played a key role in the development of this new method and that provide insight into the optimized process.

Hidden diversity in high-latitude Southern Hemisphere environments: Reinstatement of the genus Rama and description of Vandenhoekia gen. nov. (Cladophoraceae, Ulvophyceae, Chlorophyta), two highly variable genera.

The continental coasts and remote islands in the high-latitude Southern Hemisphere, including the subantarctic region, are characterized by many endemic species, high abundance of taxa, and intermediate levels of biodiversity. The macroalgal flora of these locations has received relatively little attention. Filamentous green algae are prolific in the intertidal of southern islands, but the taxonomy, distribution, and evolutionary history of these taxa are yet to be fully explored, mostly due to the difficulty of access to some of these locations. In this study, we examined specimens of the order Cladophorales from various locations in the high-latitude Southern Hemisphere including the subantarctic (the Auckland Islands, Bounty Islands, Campbell Island, Macquarie Island, and Kerguelen Islands), as well as mainland New Zealand, the Chatham Islands, Chile, and Tasmania. The analyses of the rDNA sequences of the samples revealed the existence of two new clades in a phylogeny of the Cladophoraceae. One of these clades is described as the novel genus Vandenhoekia gen. nov., which contains three species that are branched or unbranched. The amended genus Rama is reinstated to accommodate the other clade, and contains four species, including the Northern Hemisphere "Cladophora rupestris." In Rama both branched and unbranched morphologies are found. It is remarkable that gross morphology is not a predictor for generic affiliations in these algae. This study illustrates that much can still be learned about diversity in the Cladophorales and highlights the importance of new collections, especially in novel locations.

Gene-rich plastid genomes of two parasitic red algal species, Laurencia australis and L. verruciformis (Rhodomelaceae, Ceramiales), and a taxonomic revision of Janczewskia.

Parasitic red algae are an interesting system for investigating the genetic changes that occur in parasites. These parasites have evolved independently multiple times within the red algae. The functional loss of plastid genomes can be investigated in these multiple independent examples, and fine-scale patterns may be discerned. The only plastid genomes from red algal parasites known so far are highly reduced and missing almost all photosynthetic genes. Our study assembled and annotated plastid genomes from the parasites Janczewskia tasmanica and its two Laurencia host species (Laurencia elata and one unidentified Laurencia sp. A25) from Australia and Janczewskia verruciformis, its host species (Laurencia catarinensis), and the closest known free-living relative (Laurencia obtusa) from the Canary Islands (Spain). For the first time we show parasitic red algal plastid genomes that are similar in size and gene content to free-living host species without any gene loss or genome reduction. The only exception was two pseudogenes (moeB and ycf46) found in the plastid genome of both isolates of J. tasmanica, indicating potential for future loss of these genes. Further comparative analyses with the three highly reduced plastid genomes showed possible gene loss patterns, in which photosynthetic gene categories were lost followed by other gene categories. Phylogenetic analyses did not confirm monophyly of Janczewskia, and the genus was subsumed into Laurencia. Further investigations will determine if any convergent small-scale patterns of gene loss exist in parasitic red algae and how these are applicable to other parasitic systems.

Hydrogen peroxide signalling mediates fertilization and post-fertilization development in the red alga Bostrychia moritziana.

Reactive oxygen species (ROS) signalling has a multitude of roles in cellular processes throughout biology. We hypothesized that red algal fertilization may offer an interesting model to study ROS-mediated signalling, as the stages of fertilization are complex and unique. We detected the localization of ROS production microscopically and monitored the expression of three homologues of NADPH oxidase in reproductive cells during fertilization. ROS were instantaneously produced by spermatia (sperm) when they attached to female trichogynes, diffused across the cell membrane in the form of H2O2, and triggered ROS generation in the carpogonium (egg) as well as carpogonial branch cells which are not in direct contact with spermatia. The expression of NADPH oxidase homologues, RESPIRATORY BURST OXIDASE HOMOLOGUES (BmRBOHs), began to be up-regulated in the female plant upon gamete binding, peaking during the fertilization process and descending back to their original level after fertilization. Pre-treatment with diphenylene iodonium or caffeine blocked gene expression as well as H2O2 production. Post-fertilization development was also inhibited when the redox state of the plants was perturbed with H2O2 at any time before or after the fertilization. Our results suggest that H2O2 acts as an auto-propagating signalling molecule, possibly through Ca2+ channel activation, and regulates gene expression in fertilization as well as post-fertilization development in red algae.

Sex-Specific Genes and their Expression in the Life History of the Red Alga Bostrychia moritziana (Ceramiales, Rhodomelaceae).

Diverse sex determination mechanisms have been reported in eukaryotes, but little is known about the genetic pathways leading to sex determination in red algae. Sex-specific genes that could be involved in sex determination and sexual differentiation were investigated in the red alga Bostrychia moritziana by analyzing the transcriptomes of various phases including males, females, and tetrasporophytes. Sex dominantly expressed genes which showed >10-fold difference between sexes was isolated using comparative RNA-seq analysis. We found 19 gene homologues, 10 from males, and nine from females, that were found only in one sex in genomic amplification using strains collected from five different localities. Most of the sex-specific genes are involved in important cellular processes including chromosome segregation, nucleo-cytoplasmic protein shuttling, or tRNA modification. Quantitative PCR analysis showed that some sex-specific genes were differently regulated during critical events of sexual reproduction like fertilization and carposporophyte development. We could localize the expression of a male-specific gene in spermatia before and after gamete binding using RNA in situ hybridization. Amino acid sequence identity between male and female homologues of importin alpha gene and PreQ(0) reductase were highly divergent (75% and 74%, respectively), suggesting that these divergent homologues are on non-recombining UV-type chromosomes in their respective sexes. Another set of transcripts were found that were sex dominantly expressed, but not sex-specific. Nineteen out of 39 sex dominantly expressed transcripts were annotated to transposable elements. Our results suggest that sexual differentiation in B. moritziana may be achieved by multi-level regulation of cellular processes, both from genes present only in one sex and differential expression of shared genes.

Divergence times and plastid phylogenomics within the intron-rich order Erythropeltales (Compsopogonophyceae, Rhodophyta).

The advent of high-throughput sequencing (HTS) has allowed for the use of large numbers of coding regions to produce robust phylogenies. These phylogenies have been used to highlight relationships at ancient diversifications (subphyla, class) and highlight the evolution of plastid genome structure. The Erythropeltales are an order in the Compsopogonophyceae, a group with unusual plastid genomes but with low taxon sampling. We use HTS to produce near complete plastid genomes of all genera, and multiple species within some genera, to produce robust phylogenies to investigate character evolution, dating of divergence in the group, and plastid organization, including intron patterns. Our results produce a fully supported phylogeny of the genera in the Erythropeltales and suggest that morphologies (upright versus crustose) have evolved multiple times. Our dated phylogeny also indicates that the order is very old (~800 Ma), with diversification occurring after the ice ages of the Cryogenian period (750-635 Ma). Plastid gene order is congruent with phylogenetic relationships and suggests that genome architecture does not change often. Our data also highlight the abundance of introns in the plastid genomes of this order. We also produce a nearly complete plastid genome of Tsunamia transpacifica (Stylonematophyceae) to add to the taxon sampling of genomes of this class. The use of plastid genomes clearly produces robust phylogenetic relationships that can be used to infer evolutionary events, and increased taxon sampling, especially in less well-known red algal groups, will provide additional insights into their evolution.

A Comment on Salomaki and Lane 2019 "Molecular Phylogenetics Supports a Clade of Red Algal Parasites Retaining Native Plastids: Taxonomy and Terminology Revised"1.

Salomaki and Lane (2019) proposed a new terminology to group red algal parasites either as parasites containing their own (native) reduced plastid: "archaeplastic" (allied to the old designation "alloparasite") or parasites that contain only a host plastid: "neoplastic" (similar to the older term "adelphoparasite"). We believe this is premature. There are examples that contradict their proposed grouping, and their proposal was based on work from the mid-1990s that should be re-evaluated. We also believe that grouping red algal parasites into two groups obscures both our lack of knowledge of these organisms and the diversity that is already seen in the few intensively studied parasites. Instead of making generalizations based on limited knowledge, further in-depth study should be encouraged and will be useful in understanding these intriguing organisms.

The Organelle Genomes in the Photosynthetic Red Algal Parasite Pterocladiophila hemisphaerica (Florideophyceae, Rhodophyta) Have Elevated Substitution Rates and Extreme Gene Loss in the Plastid Genome.

Comparative organelle genome studies of parasites can highlight genetic changes that occur during the transition from a free-living to a parasitic state. Our study focuses on a poorly studied group of red algal parasites, which are often closely related to their red algal hosts and from which they presumably evolved. Most of these parasites are pigmented and some show photosynthetic capacity. Here, we assembled and annotated the complete organelle genomes of the photosynthetic red algal parasite, Pterocladiophila hemisphaerica. The plastid genome is the smallest known red algal plastid genome at 68,701 bp. The plastid genome has many genes missing, including all photosynthesis-related genes. In contrast, the mitochondrial genome is similar in architecture to that of other free-living red algae. Both organelle genomes show elevated mutation rates and significant changes in patterns of selection, measured as dN/dS ratios. This caused phylogenetic analyses, even of multiple aligned proteins, to be unresolved or give contradictory relationships. Full plastid datasets interfered by selected best gene evolution models showed the supported relationship of P. hemisphaerica within the Ceramiales, but the parasite was grouped with support as sister to the Gracilariales when interfered under the GHOST model. Nuclear rDNA showed a supported grouping of the parasite within a clade containing several red algal orders including the Gelidiales. This photosynthetic parasite, which is unable to photosynthesize with its own plastid due to the total loss of all photosynthesis genes, raises intriguing questions on parasite-host organelle genome capabilities and interactions.

Comparative phylogeography of two Agarophyton species in the New Zealand archipelago.

Molecular studies have reported the coexistence of two species of Agarophyton in New Zealand: the newly described A.transtasmanicum with an apparently restricted distribution to some sites in the North Island, and the more widespread A.chilense. Here, we compared the distribution, genetic diversity, and structure of both Agarophyton species throughout the archipelago using sequences of the nuclear Internal Transcribed Spacer 2 (ITS2) marker. Agarophyton chilense's distribution was continuous and extensive along the North and South Islands, Stewart Island, and Chatham Island, and the genetic clusters were mostly concordant with boundaries between biogeographic regions. In contrast, specimens of A.transtasmanicum were collected in four sites broadly distributed in both the North and South Islands, with no clear spatial structure of the genetic diversity. Populations, where the species co-occurred, tended to display similar levels in genetic diversity for the two species. Demographic inferences supported a postglacial demographic expansion for two A.chilense genetic clusters, one present in the South Island and the eastern coast of the North Island, and the other present in northern South Island. A third genetic cluster located on the western coast of the North Island had a signature of long-term demographic stability. For A.transtasmanicum, the skyline plot also suggested a postglacial demographic expansion. Last, we developed a new molecular tool to quickly and easily distinguish between the two Agarophyton species, which could be used to ease future fine-scale population studies, especially in areas where the two species coexist.

Enantioselective Folding of Enynes by Gold(I) Catalysts with a Remote C2-Chiral Element.

Chiral gold(I) catalysts have been designed based on a modified JohnPhos ligand with a distal C2-2,5-diarylpyrrolidine that creates a tight binding cavity. The C2-chiral element is close to where the C-C bond formation takes place in cyclizations of 1,6-enynes. These chiral mononuclear catalysts have been applied for the enantioselective 5-exo-dig and 6-endo-dig cyclization of different 1,6-enynes as well as in the first enantioselective total synthesis of three members of the carexane family of natural products. Opposite enantioselectivities have been achieved in seemingly analogous reactions of 1,6-enynes, which result from different chiral folding of the substrates based on attractive aryl-aryl interactions.

Gold(I)-Catalyzed Intramolecular C(sp3 )-H Insertion by Decarbenation of Cycloheptatrienes.

A novel synthesis of indanes and dihydronaphtalenes based on the intramolecular insertion into C(sp3 )-H bonds of gold(I) carbenes generated by retro-Buchner reaction (decarbenation) has been developed. Deuterium-labeling and kinetic isotope effect experiments, DFT calculations, and generation of the proposed carbene intermediate from a well-characterized gold(I) carbenoid support the involvement of a three-center concerted mechanism for the C(sp3 )-H functionalization process.

Costatone C-A New Halogenated Monoterpene from the New Zealand Red Alga Plocamium angustum.

Red algae of the genus Plocamium have been a rich source of halogenated monoterpenes. Herein, a new cyclic monoterpene, costatone C (7), was isolated from the extract of P. angustum collected in New Zealand, along with the previously reported (1E,5Z)-1,6-dichloro-2-methylhepta-1,5-dien-3-ol (8). Elucidation of the planar structure of 7 was achieved through conventional NMR and (-)-HR-APCI-MS techniques, and the absolute configuration by comparison of experimental and DFT-calculated ECD spectra. The absolute configuration of 8 was determined using Mosher's method. Compound 7 showed mild antibacterial activity against Staphylococcus aureus and S. epidermidis. The state of Plocamium taxonomy and its implications upon natural product distributions, especially across samples from specimens collected in different countries, is also discussed.

The endemic Cladophorales (Ulvophyceae) of ancient Lake Baikal represent a monophyletic group of very closely related but morphologically diverse species.

Lake Baikal, the oldest lake in the world, is home to spectacular biodiversity and extraordinary levels of endemism. While many of the animal species flocks from Lake Baikal are famous examples of evolutionary radiations, the lake also includes a wide diversity of endemic algae that are not well investigated with regards to molecular-biological taxonomy and phylogeny. The endemic taxa of the green algal order Cladophorales show a range of divergent morphologies that led to their classification in four genera in two families. We sequenced partial large- and small-subunit rDNA as well as the internal transcribed spacer region of 14 of the 16 described endemic taxa to clarify their phylogenetic relationships. One endemic morphospecies, Cladophora kusnetzowii, was shown to be conspecific with the widespread Aegagropila linnaei. All other endemic morphospecies formed a monophyletic group nested within the genus Rhizoclonium (Cladophoraceae), a very surprising result, in stark contrast to their morphological affinities. The Baikal clade represents a species flock of closely related taxa with very low genetic differentiation. Some of the morphospecies were congruent with lineages recovered in the phylogenies, but due to the low phylogenetic signal in the rDNA sequences the relationships within the Baikal clade were not all well resolved. The Baikal clade appears to represent a recent radiation, based on the low molecular divergence within the group, and it is hypothesized that the large morphological variation results from diversification in sympatry from a common ancestor in Lake Baikal.

Molecular phylogeny and taxonomic revision of the genus Wittrockiella (Pithophoraceae, Cladophorales), including the descriptions of W. australis sp. nov. and W. zosterae sp. nov.

Wittrockiella is a small genus of filamentous green algae that occurs in habitats with reduced or fluctuating salinities. Many aspects of the basic biology of these algae are still unknown and the phylogenetic relationships within the genus have not been fully explored. We provide a phylogeny based on three ribosomal markers (ITS, LSU, and SSU rDNA) of the genus, including broad intraspecific sampling for W. lyallii and W. salina, recommendations for the use of existing names are made, and highlight aspects of their physiology and life cycle. Molecular data indicate that there are five species of Wittrockiella. Two new species, W. australis and W. zosterae, are described, both are endophytes. Although W. lyallii and W. salina can be identified morphologically, there are no diagnostic morphological characters to distinguish between W. amphibia, W. australis, and W. zosterae. A range of low molecular weight carbohydrates were analyzed but proved to not be taxonomically informative. The distribution range of W. salina is extended to the Northern Hemisphere as this species has been found in brackish lakes in Japan. Furthermore, it is shown that there are no grounds to recognize W. salina var. kraftii, which was described as an endemic variety from a freshwater habitat on Lord Howe Island, Australia. Culture experiments indicate that W. australis has a preference for growth in lower salinities over full seawater. For W. amphibia and W. zosterae, sexual reproduction is documented, and the split of these species is possibly attributable to polyploidization.

Molecular phylogeny of the Cladophoraceae (Cladophorales, Ulvophyceae), with the resurrection of Acrocladus Nägeli and Willeella Børgesen, and the description of Lurbica gen. nov. and Pseudorhizoclonium gen. nov.

The taxonomy of the Cladophoraceae, a large family of filamentous green algae, has been problematic for a long time due to morphological simplicity, parallel evolution, phenotypic plasticity, and unknown distribution ranges. Partial large subunit (LSU) rDNA sequences were generated for 362 isolates, and the analyses of a concatenated dataset consisting of unique LSU and small subunit (SSU) rDNA sequences of 95 specimens greatly clarified the phylogeny of the Cladophoraceae. The phylogenetic reconstructions showed that the three currently accepted genera Chaetomorpha, Cladophora, and Rhizoclonium are polyphyletic. The backbone of the phylogeny is robust and the relationships of the main lineages were inferred with high support, only the phylogenetic position of both Chaetomorpha melagonium and Cladophora rupestris could not be inferred unambiguously. There have been at least three independent switches between branched and unbranched morphologies within the Cladophoraceae. Freshwater environments have been colonized twice independently, namely by the freshwater Cladophora species as well as by several lineages of the Rhizoclonium riparium clade. In an effort to establish monophyletic genera, the genera Acrocladus and Willeella are resurrected and two new genera are described: Pseudorhizoclonium and Lurbica.

Rhodenigma contortum, an obscure new genus and species of Rhodogorgonales (Rhodophyta) from Western Australia.

An unknown microscopic, branched filamentous red alga was isolated into culture from coral fragments collected in Coral Bay, Western Australia. It grew well unattached or attached to glass with no reproduction other than fragmentation of filaments. Cells of some branch tips became slightly contorted and digitated, possibly as a substrate-contact-response seen at filament tips of various algae. Attached multicellular compact disks on glass had a very different cellular configuration and size than the free filaments. In culture the filaments did not grow on or in coral fragments. Molecular phylogenies based on four markers (rbcL, cox1, 18S, 28S) clearly showed it belongs to the order Rhodogorgonales, as a sister clade of Renouxia. Based on these results, the alga is described as the new genus and species Rhodenigma contortum in the Rhodogorgonaceae. It had no morphological similarity to either of the other genera in Rhodogorgonaceae and illustrates the unknown diversity in cryptic habitats such as tropical coral rubble.

Contrasting patterns of population structure and demographic history in cryptic species of Bostrychia intricata (Rhodomelaceae, Rhodophyta) from New Zealand.

Spatial patterns of genetic diversity provide insight into the demography and history of species. Morphologically similar but genetically distinct "cryptic" species are increasingly being recognized in marine organisms through molecular analyses. Such species are, on closer inspection, often discovered to display contrasting life histories or occasionally minor morphological differences; molecular tools can thus be useful indicators of diversity. Bostrychia intricata, a marine red alga, is widely distributed throughout the Southern Hemisphere and comprises many cryptic species. We used mitochondrial cytochrome c oxidase I gene sequences to assess the genetic variation, population genetic structure, and demographic history of B. intricata in New Zealand. Our results supported the existence of three cryptic species of B. intricata (N2, N4, and N5) in New Zealand. Cryptic species N4, which was found throughout New Zealand, showed a higher genetic diversity and wider distribution than the other two species, which were only found in the North Island and northern South Island. Our analyses showed low to moderate genetic differentiation among eastern North Island populations for cryptic species N2, but high differentiation among North and South Island populations for N4, suggesting different population structure between these cryptic species. Data also indicated that N2 has recently undergone population expansion, probably since the Last Glacial Maximum (LGM), while the higher genetic diversity in N4 populations suggests persistence in situ through the LGM. The contrasting population structures and inferred demographic histories of these species highlight that life history can vary greatly even among morphologically indistinguishable taxa.

Stereodivergent total synthesis of Δ9-tetrahydrocannabinols.

All four stereoisomers of Δ(9)-tetrahydrocannabinol (Δ(9)-THC) were synthesized in concise fashion using stereodivergent dual catalysis. Thus, following identical synthetic sequences and applying identical reaction conditions to the same set of starting materials, selective access to the four stereoisomers of THC was achieved in five steps.

Past climate-driven range shifts structuring intraspecific biodiversity levels of the giant kelp (Macrocystis pyrifera) at global scales.

The paradigm of past climate-driven range shifts structuring the distribution of marine intraspecific biodiversity lacks replication in biological models exposed to comparable limiting conditions in independent regions. This may lead to confounding effects unlinked to climate drivers. We aim to fill in this gap by asking whether the global distribution of intraspecific biodiversity of giant kelp (Macrocystis pyrifera) is explained by past climate changes occurring across the two hemispheres. We compared the species' population genetic diversity and structure inferred with microsatellite markers, with range shifts and long-term refugial regions predicted with species distribution modelling (SDM) from the last glacial maximum (LGM) to the present. The broad antitropical distribution of Macrocystis pyrifera is composed by six significantly differentiated genetic groups, for which current genetic diversity levels match the expectations of past climate changes. Range shifts from the LGM to the present structured low latitude refugial regions where genetic relics with higher and unique diversity were found (particularly in the Channel Islands of California and in Peru), while post-glacial expansions following ~ 40% range contraction explained extensive regions with homogenous reduced diversity. The estimated effect of past climate-driven range shifts was comparable between hemispheres, largely demonstrating that the distribution of intraspecific marine biodiversity can be structured by comparable evolutionary forces across the global ocean. Additionally, the differentiation and endemicity of regional genetic groups, confers high conservation value to these localized intraspecific biodiversity hotspots of giant kelp forests.

Enantioselective Catalysis with Pyrrolidinyl Gold(I) Complexes: DFT and NEST Analysis of the Chiral Binding Pocket.

A new generation of chiral gold(I) catalysts based on variations of complexes with JohnPhos-type ligands with a remote C2-symmetric 2,5-diarylpyrrolidine have been synthesized with different substitutions at the top and bottom aryl rings: from replacing the phosphine by a N-heterocyclic carbene (NHC) to increasing the steric hindrance with bis- or tris-biphenylphosphine scaffolds, or by directly attaching the C2-chiral pyrrolidine in the ortho-position of the dialkylphenyl phosphine. The new chiral gold(I) catalysts have been tested in the intramolecular [4+2] cycloaddition of arylalkynes with alkenes and in the atroposelective synthesis of 2-arylindoles. Interestingly, simpler catalysts with the C2-chiral pyrrolidine in the ortho-position of the dialkylphenyl phosphine led to the formation of opposite enantiomers. The chiral binding pockets of the new catalysts have been analyzed by DFT calculations. As revealed by non-covalent interaction plots, attractive non-covalent interactions between substrates and catalysts direct specific enantioselective folding. Furthermore, we have introduced the open-source tool NEST, specifically designed to account for steric effects in cylindrical-shaped complexes, which allows predicting experimental enantioselectivities in our systems.