Revisiting of Carex sect. Confertiflorae s.l. (Cyperaceae): New data from molecular and morphological evidence and first insights on Carex biogeography in East Asia

Carex sect. Confertiflorae s.l. is a medium-sized species group (ca. 40 species) with its center of diversity in E Asia (China and Japan). According to morphological traits, the section has been proposed to split into two sections (sects. Confertiflorae sensu Ohwi and Molliculae Ohwi) up to five different ones (sects. Confertiflorae s.s., Molliculae, Dispalatae Ohwi, Ischnostachyae Ohwi, and Alliiformes Akiyama). Recent phylogenetic reconstructions showed Confertiflorae s.l. not to be monophyletic, as species traditionally considered part of it were found to belong to other clades, whereas species traditionally ascribed to other sections were nested within it. In this study, we investigated the phylogenetic structure, morphological affinities, and biogeographic history of sect. Confertiflorae s.l. We employed a taxon-based approach to explore the morphological affinities of the species considered in sect. Confertiflorae and compared the micromorphology of the nutlets of almost all the taxa using SEM. We included 40 samples representing 31 species/subspecies of sect. Confertiflorae s.l. and used two nuclear (ETS and ITS) and three plastid (trnL-F, matK, and rpl32-trnL ^UAG) molecular markers to reconstruct the phylogeny of the group. The phylogenetic analyses confirmed the polyphyly of sect. Confertiflorae s.l., whose representatives were found within five distinct clades. From these, two clades, sect. Confertiflorae and sect. Molliculae, were found to be closely related and contained the majority of the species. The composition of the two clades agreed with the morphological structure of the group, and we confirmed an exclusive combination of features (namely color of basal sheaths, presence of bract sheath, peduncle of lowest spike, inflorescence sex distribution, shape of pistillate glume apex, and color and veins of utricle, among others) characterizing each of the two clades. The origin of the two clades was found to be in the early Pliocene; however, the majority of the diversification events within each clade took place during the Pleistocene. This illustrates that although Asia has been regarded as having little potential ecological space for Carex to diversify due to its climate stability, groups of sedges sub-endemic from that area may have a fairly recent origin related to glaciations. We proposed the rearrangement of sect. Confertiflorae as previously conceived as three independent sections: the monotypic Alliiformes, sect. Molliculae, and sect. Paludosae.     

Macroevolutionary insights into sedges (Carex: Cyperaceae): The effects of rapid chromosome number evolution on lineage diversification

Changes in holocentric chromosome number due to fission and fusion have direct and immediate effects on genome structure and recombination rates. These, in turn, may influence ecology and evolutionary trajectories profoundly. Sedges of the genus Carex (Cyperaceae) comprise ca. 2000 species with holocentric chromosomes. The genus exhibits a phenomenal range in the chromosome number (2n = 10 − 132) with almost not polyploidy. In this study, we integrated the most comprehensive cytogenetic and phylogenetic data for sedges with associated climatic and morphological data to investigate the hypothesis that high recombination rates are selected when evolutionary innovation is required, using chromosome number evolution as a proxy for recombination rate. We evaluated Ornstein–Uhlenbeck models to infer shifts in chromosome number equilibrium and selective regime. We also tested the relationship between chromosome number and diversification rates. Our analyses demonstrate significant correlations between morphology and climatic niche and chromosome number in Carex. Nevertheless, the amount of chromosomal variation that we are able to explain is very small. We recognized a large number of shifts in mean chromosome number, but a significantly lower number in climatic niche and morphology. We also detected a peak in diversification rates near intermediate recombination rates. In combination, these analyses point toward the importance of chromosome evolution to the evolutionary history of Carex. Our work suggests that the effect of chromosome evolution on recombination rates, not just on reproductive isolation, may be central to the evolutionary history of sedges.     

Geographical vs. ecological diversification in Carex section Phacocystis (Cyperaceae): Patterns hidden behind a twisted taxonomy

Carex section Phacocystis (Cyperaceae) is one of the most diverse and taxonomically complex groups of sedges (between 116 and 147 species), with a worldwide distribution in a wide array of biomes. It has a very complicated taxonomic history, with numerous disagreements among different treatments. We studied the biogeography and niche evolution in a phylogenetic framework to unveil the relative contribution of geographical and ecological drivers to diversification of the group. We used a large species sampling of the section (82% of extant species) to build a phylogeny based on four DNA regions, constrained with a phylogenomic HybSeq tree and dated with six fossil calibrations. Our phylogenetic results recovered section Phacocystis s.s. (core Phacocystis) as sister to section Praelongae. Ancestral area reconstruction points toward the N Pacific as the cradle for the crown diversification of section Phacocystis during the Middle Miocene. Wide distributions were recurrently inferred across deep nodes. Large Northern Hemisphere lineages with geographical congruence were retrieved, pointing toward the importance of allopatric divergence at deep phylogenetic levels, whereas within-area speciation emerges as the predominant pattern at shallow phylogenetic level. The Southern Hemisphere (Neotropics, SW Pacific) was colonized several times from the Northern Hemisphere. The global expansion of Carex section Phacocystis did not entail major ecological changes along the inner branches of the phylogeny. Nevertheless, ecological differentiation seems to gain importance toward recent times.     

Antidiabetic Potential of Protein Hydrolysates and Peptide Fractions from Lima Bean (Phaseolus lunatus L): An In Vitro Study

International Journal of Peptide Research and Therapeutics - Characterized by uncontrolled, long-term high blood sugar levels, diabetes mellitus affects ever increasing numbers of people worldwide....     

Passive Internalization of Bioactive β-Casein Peptides into Phospholipid (POPC) Bilayers. Free Energy Landscapes from Unbiased Equilibrium MD Simulations at μs-Time Scale

Food Biophysics - Absorption of bioactive peptides in the intestinal epithelium take place in the apical or the basolateral tight junctions of the cells. Depending on the peptide size and...     

Conformational insights on the molecular recognition processes of carbohydrate molecules by proteins and enzymes: A 3D view by using NMR

This review focuses, in a non-exhaustive manner, on the essential structural and conformational features of protein–carbohydrate interactions and on some applications of NMR spectroscopy to deal with this topic from different levels of complexity.     

Long-term biomonitoring of lichen and bryophyte biodiversity at Burnham Beeches SAC and global environmental change

Long-term monitoring began 20 years ago at Burnham Beeches Site of Special Interest (SSSI), National Nature Reserve (NNR) and European Special Area of Conservation (SAC) lying 40 km west of London as a consequence of the authorization of an application to extract gravel from an adjacent site lying north of Slough Trading Estate. Dust monitoring (sticky pads) and photographic monitoring, recording and image analysis was instigated in 1992 on Parmelion communities to assess changes in lichen growth, health and community composition. Long-term monitoring identifies that the lichen flora on free-standing trees has undergone rapid expansion from a near dominance by the SO₂-tolerant ‘acidophyte’ species Lecanora conizaeoides and Hypogymnia physodes following reductions in SO₂ concentrations. Long-term influences of low levels of eutrophication, gaseous pollutants (particularly globally rising background ozone concentrations) on lichen and bryophyte communities and succession under changing climatic conditions are unknown. Soil–plant relationships, lichen–invertebrate interactions and a pollution legacy must also be considered.     

The Design and Synthesis of N 4-Anthraniloyl-2′-dC,the Improved Syntheses of N 4-Carbamoyl-and N4- Ureidocarbamoyl-2′-dC,Incorporation into Oligonucleotides and Triplex Formation Testing

Abstract

Three modified nucleosides were designed with the aim of achieving triplet formation with the CG base pair of duplex DNA. Direct anthraniloylation of 2′-deoxycytidine, using isatoic anhydride, afforded the novel N ⁴-anthraniloyl-2′-deoxycytidine. Much improved preparations of N ⁴-carbamoyl-2′-deoxycytidine and of N ⁴-ureidocarbonyl-2′-deoxycytidine were accomplished. The modified nucleosides were incorporated into oligonucleotides. Thermal denaturation studies and gel mobility shift analysis suggest that these nucleosides do not form base triplets with any of the four base pairs of DNA.     

Identification and Quantification of AKT Isoforms and Phosphoforms in Breast Cancer Using a Novel Nanofluidic Immunoassay

Breast cancer subtype-specific molecular variations can dramatically affect patient responses to existing therapies. It is thought that differentially phosphorylated protein isoforms might be a useful prognostic biomarker of drug response in the clinic. However, the accurate detection and quantitative analysis of cancer-related protein isoforms and phospho-isoforms in tumors are limited by current technologies. Using a novel, fully automated nanocapillary electrophoresis immunoassay (NanoPro^TM 1000) designed to separate protein molecules based on their isoelectric point, we developed a reliable and highly sensitive assay for the detection and quantitation of AKT isoforms and phosphoforms in breast cancer. This assay enabled the measurement of activated AKT1/2/3 in breast cancer cells using protein produced from as few as 56 cells. Importantly, we were able to assign an identity for the phosphorylated S473 phosphoform of AKT1, the major form of activated AKT involved in multiple cancers, including breast, and a current focus in clinical trials for targeted intervention. The ability of our AKT assay to detect and measure AKT phosphorylation from very low amounts of total protein will allow the accurate evaluation of patient response to drugs targeting activated PI3K-AKT using scarce clinical specimens. Moreover, the capacity of this assay to detect and measure all three AKT isoforms using one single pan-specific antibody enables the study of the multiple and variable roles that these isoforms play in AKT tumorigenesis.Activation of the PI3K-AKT signaling pathway is one of the most common events in cancer (1, 2). Pathway activation can confer a number of advantages to the cancer cells, including enhanced proliferation and survival (1, 2). Multiple mechanisms exist by which the pathway may become activated, including amplification or activation of receptor tyrosine kinases (e.g. ERBB2 in breast and EGFR in lung tumors), mutation of the catalytic or regulatory subunits of PI3K (e.g. PIK3CA in colorectal and breast tumors), loss of the negative regulator PTEN (e.g. mutation in prostate and melanoma), and gain of function of AKT (e.g. amplification or mutation in breast and pancreatic tumors) (reviewed in Refs. 1 and 2).AKT represents a central node in the PI3K signaling cascade (3). AKT is recruited to the cell membrane via its pleckstrin homology domain when PI3K phosphorylates PIP₂ to form PIP₃ (4, 5). Following recruitment, AKT is phosphorylated by PDK1 and the rictor-mTOR complex, resulting in conformational changes and activation of the protein (5–8). Multiple studies have shown that the phosphorylation of AKT leads to the phosphorylation and activation of downstream effectors of the signaling pathway, such as mTOR complex 1 and S6K (reviewed in Ref. 1). The central role of this pathway in cancer is further underscored by the efforts of multiple pharmaceutical companies that have developed inhibitors against AKT as potential anti-oncogenic therapeutics (9).Despite the importance of AKT in growth and survival signaling in cancer, there are surprisingly few data that address the specific roles played in growth and survival by the multiple AKT family members (AKT-1, -2, and -3) and different phosphorylation and putative phosphorylation sites that can potentially activate the protein. Western blot analysis has been the foundation of most AKT studies, but in many cases pan-AKT antibodies have been employed that fail to distinguish between the different AKT isoforms. Recent siRNA silencing studies have indicated distinct functions for different AKT family members within a cell (10, 11). Moreover, there is evidence in breast cancer that the three isoforms exhibit different localizations and therefore must have at least partially distinct functions (12). Similarly, evidence is mounting for multiple phosphorylation sites in AKT beyond the two most studied phosphorylation events (Thr-308 and Ser-473) (5–8). Phosphorylation at serine and threonine residues at Thr-72 and Ser-246 may be required for the activation or regulation of kinase activity (13). The functional significance of constitutive phosphorylation of Ser-124 and Thr-450 is still unknown (14). Finally, there is evidence that phosphorylation of tyrosine residues at Tyr-315 and Tyr-326 is required for full kinase activity (15).Analysis of such phospho- and isoform-specific activation often requires complicated in-depth analyses using large quantities of proteins, purified recombinant protein, immunoprecipitation, incorporation of ³²P isotopes, and/or mass spectroscopy, which makes such studies more difficult to perform and not easily adaptable to clinical specimens. Thus, better methods are required for the accurate assessment of both phosphoform and isoform usage in cells with an activated PI3K-AKT pathway and the effects of pathway inhibitors using relatively small amounts of starting material. We describe here the development of such an assay using nanocapillary-based isoelectric focusing (16). This approach allows the separation of AKT into distinct peaks that correspond to different iso- and phosphoforms using a small amount of starting material and a single pan-specific antibody. This approach should allow for more accurate determinations of isoform usage in different cell types, as well as of changes in phosphorylation states in response to pathway inhibition, including in clinical specimens.