Separating specific cell phenotypes from a heterotypic mixture is a critical step in many research projects. Traditional methods usually require a large sample volume and a complex preparation ...process that may alter cell property during the sorting process. Here we present the use of electrical impedance as an indicator of cell health and for identifying specific microalgal phenotypes. We developed a microfluidic platform for measuring electrical impedance at different frequencies using the bacterium-sized green alga Picochlorum SE3. The cells were cultured under different salinity conditions and sampled at four different time points. Our results demonstrate the utility of electrical impedance as an indicator of cell phenotype by providing results that are consistent with known changes in cell size and physiology. Outliers in the cell data distribution are particularly useful because they represent phenotypes that have the ability to maintain size and/or membrane ionic permeability under prolonged salt stress. This suggests that our device can be used to identify and sort desired (e.g., experimentally evolved, mutant) cell phenotypes based on their electrical impedance properties.
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Abstract
Changes in atmospheric CO
2
concentration have played a central role in algal and plant adaptation and evolution. The commercially important red algal genus,
Pyropia
(Bangiales) appears to ...have responded to inorganic carbon (C
i
) availability by evolving alternating heteromorphic generations that occupy distinct habitats. The leafy gametophyte inhabits the intertidal zone that undergoes frequent emersion, whereas the sporophyte conchocelis bores into mollusk shells. Here, we analyze a high-quality genome assembly of
Pyropia yezoensis
to elucidate the interplay between C
i
availability and life cycle evolution. We find horizontal gene transfers from bacteria and expansion of gene families (e.g. carbonic anhydrase, anti-oxidative related genes), many of which show gametophyte-specific expression or significant up-regulation in gametophyte in response to dehydration. In conchocelis, the release of HCO
3
-
from shell promoted by carbonic anhydrase provides a source of C
i
. This hypothesis is supported by the incorporation of
13
C isotope by conchocelis when co-cultured with
13
C-labeled CaCO
3
.
Plants have adapted to the diurnal light-dark cycle by establishing elaborate transcriptional programs that coordinate many metabolic, physiological, and developmental responses to the external ...environment. These transcriptional programs have been studied in only a few species, and their function and conservation across algae and plants is currently unknown. We performed a comparative transcriptome analysis of the diurnal cycle of nine members of Archaeplastida, and we observed that, despite large phylogenetic distances and dramatic differences in morphology and lifestyle, diurnal transcriptional programs of these organisms are similar. Expression of genes related to cell division and the majority of biological pathways depends on the time of day in unicellular algae but we did not observe such patterns at the tissue level in multicellular land plants. Hence, our study provides evidence for the universality of diurnal gene expression and elucidates its evolutionary history among different photosynthetic eukaryotes.
Corals comprise a biomineralizing cnidarian, dinoflagellate algal symbionts, and associated microbiome of prokaryotes and viruses. Ongoing efforts to conserve coral reefs by identifying the major ...stress response pathways and thereby laying the foundation to select resistant genotypes rely on a robust genomic foundation. Here we generated and analyzed a high quality long-read based ~886 Mbp nuclear genome assembly and transcriptome data from the dominant rice coral, Montipora capitata from Hawai'i. Our work provides insights into the architecture of coral genomes and shows how they differ in size and gene inventory, putatively due to population size variation. We describe a recent example of foreign gene acquisition via a bacterial gene transfer agent and illustrate the major pathways of stress response that can be used to predict regulatory components of the transcriptional networks in M. capitata. These genomic resources provide insights into the adaptive potential of these sessile, long-lived species in both natural and human influenced environments and facilitate functional and population genomic studies aimed at Hawaiian reef restoration and conservation.
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A 44-year-old female with a vision of 10/200 in the right eye had double pits in the temporal segment of the optic disc with serous macular detachment. Spectral-domain optical coherence tomography ...(SD-OCT) confirmed serous retinal detachment, an outer layer hole, and double optic disc pits. The patient underwent pars plana vitrectomy with modified ILM flap surgery involving fovea-sparing internal limiting membrane peeling (FSIP) technique with inverted ILM flap tucking with gas tamponade. Post surgery, the communications between perineural and intraretinal spaces were obliterated with flaps of ILM covering the pits, with reduced serous macular detachment and BCVA of 20/120. FSIP with inverted internal limiting membrane flap tuck can be an effective technique to manage rare cases of double ODP-M.
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The role and extent of horizontal gene transfer (HGT) in eukaryotes are hotly disputed topics that impact our understanding of the origin of metabolic processes and the role of organelles in cellular ...evolution. We addressed this issue by analyzing 10 novel Cyanidiales genomes and determined that 1% of their gene inventory is HGT-derived. Numerous HGT candidates share a close phylogenetic relationship with prokaryotes that live in similar habitats as the Cyanidiales and encode functions related to polyextremophily. HGT candidates differ from native genes in GC-content, number of splice sites, and gene expression. HGT candidates are more prone to loss, which may explain the absence of a eukaryotic pan-genome. Therefore, the lack of a pan-genome and cumulative effects fail to provide substantive arguments against our hypothesis of recurring HGT followed by differential loss in eukaryotes. The maintenance of 1% HGTs, even under selection for genome reduction, underlines the importance of non-endosymbiosis related foreign gene acquisition.
It has long been recognized that a suite of proteins exists in coral skeletons that is critical for the oriented precipitation of calcium carbonate crystals, yet these proteins remain poorly ...characterized. Using liquid chromatography-tandem mass spectrometry analysis of proteins extracted from the cell-free skeleton of the hermatypic coral, Stylophora pistillata , combined with a draft genome assembly from the cnidarian host cells of the same species, we identified 36 coral skeletal organic matrix proteins. The proteome of the coral skeleton contains an assemblage of adhesion and structural proteins as well as two highly acidic proteins that may constitute a unique coral skeletal organic matrix protein subfamily. We compared the 36 skeletal organic matrix protein sequences to genome and transcriptome data from three other corals, three additional invertebrates, one vertebrate, and three single-celled organisms. This work represents a unique extensive proteomic analysis of biomineralization-related proteins in corals from which we identify a biomineralization “toolkit,” an organic scaffold upon which aragonite crystals can be deposited in specific orientations to form a phenotypically identifiable structure.
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Diatoms are unicellular algae that accumulate significant amounts of triacylglycerols as storage lipids when their growth is limited by nutrients. Using biochemical, physiological, bioinformatics, ...and reverse genetic approaches, we analyzed how the flux of carbon into lipids is influenced by nitrogen stress in a model diatom, Phaeodactylum tricornutum . Our results reveal that the accumulation of lipids is a consequence of remodeling of intermediate metabolism, especially reactions in the tricarboxylic acid and the urea cycles. Specifically, approximately one-half of the cellular proteins are cannibalized; whereas the nitrogen is scavenged by the urea and glutamine synthetase/glutamine 2-oxoglutarate aminotransferase pathways and redirected to the de novo synthesis of nitrogen assimilation machinery, simultaneously, the photobiological flux of carbon and reductants is used to synthesize lipids. To further examine how nitrogen stress triggers the remodeling process, we knocked down the gene encoding for nitrate reductase, a key enzyme required for the assimilation of nitrate. The strain exhibits 40–50% of the mRNA copy numbers, protein content, and enzymatic activity of the wild type, concomitant with a 43% increase in cellular lipid content. We suggest a negative feedback sensor that couples photosynthetic carbon fixation to lipid biosynthesis and is regulated by the nitrogen assimilation pathway. This metabolic feedback enables diatoms to rapidly respond to fluctuations in environmental nitrogen availability.
Significance When starved for nutrients, diatoms redirect carbon toward biosynthesis of storage lipids, triacylglycerols (TAGs). We examined how this modification is achieved in the diatom Phaeodactylum tricornutum. Under nitrogen stress, the cells cannibalized their photosynthetic apparatus while recycling intracellular nitrogen and redirecting it to synthesize nitrogen assimilation enzymes. Simultaneously, they allocated newly fixed carbon toward lipids. In contrast, a nitrate reductase knocked-down strain shunted ∼40% more carbon toward TAGs than the wild type without losing photosynthetic capacity. Our results show that diatoms can remodel their intermediate metabolism on environmental cues and reveal that a key signal in this remodeling is associated with nitrogen assimilation. This insight informs a strategy of developing a much more efficient pathway to produce algal-based biofuels.
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Diatoms and other chromalveolates are among the dominant phytoplankters in the world's oceans. Endosymbiosis was essential to the success of chromalveolates, and it appears that the ancestral plastid ...in this group had a red algal origin via an ancient secondary endosymbiosis. However, recent analyses have turned up a handful of nuclear genes in chromalveolates that are of green algal derivation. Using a genome-wide approach to estimate the "green" contribution to diatoms, we identified >1700 green gene transfers, constituting 16% of the diatom nuclear coding potential. These genes were probably introduced into diatoms and other chromalveolates from a cryptic endosymbiont related to prasinophyte-like green algae. Chromalveolates appear to have recruited genes from the two major existing algal groups to forge a highly successful, species-rich prtist lineage.
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The thecate amoeba Paulinella is a valuable model for understanding plastid organellogenesis because this lineage has independently gained plastids (termed chromatophores) of alpha-cyanobacterial ...provenance. Plastid primary endosymbiosis in Paulinella occurred relatively recently (90-140 million years ago, Mya), whereas the origin of the canonical Archaeplastida plastid occurred >1,500 Mya. Therefore, these two events provide independent perspectives on plastid formation on vastly different timescales. Here we generated the complete chromatophore genome sequence from P. longichromatophora (979,356 bp, GC-content = 38.8%, 915 predicted genes) and P. micropora NZ27 (977,190 bp, GC-content = 39.9%, 911 predicted genes) and compared these data to that from existing chromatophore genomes. Our analysis suggests that when a basal split occurred among photosynthetic Paulinella species ca. 60 Mya, only 35% of the ancestral orthologous gene families from the cyanobacterial endosymbiont remained in chromatophore DNA. Following major gene losses during the early stages of endosymbiosis, this process slowed down significantly, resulting in a conserved gene content across extant taxa. Chromatophore genes faced relaxed selection when compared to homologs in free-living alpha-cyanobacteria, likely reflecting the homogeneous intracellular environment of the Paulinella host. Comparison of nucleotide substitution and insertion/deletion events among different P. micropora strains demonstrates that increases in AT-content and genome reduction are ongoing and dynamic processes in chromatophore evolution.
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