Biology subjects

Sharon, I.

Publications and source records attributed to Sharon, I..

14 recordsLinked to original sources

bioRxiv2025

Nuclear lamina-associated domain biogenesis is regulated by nuclear pore density during embryogenesis and mediates UV protection

Lamina-associated domains (LADs) are critical for genome organization and function, but their formation during development is not well understood. Here, we use DamID and image analysis to reveal the dynamics of LAD biogenesis during C. elegans embryogenesis. At early stages, DNA at the lamina is transcriptionally active and lacks lamina-associated heterochromatin. This state depends on abundant nuclear pores, which prevent heterochromatin accumulation at the nuclear periphery. As development proceeds, pore numbers decline, enabling heterochromatin to access the lamina. Reducing nuclear-pore components induces precocious accumulation of heterochromatin to the lamina. Functionally, we find that heterochromatic LADs confer protection against ultraviolet (UV) radiation. Older embryos are resistant to UV light and concentrate DNA damage at the nuclear periphery, whereas early embryos are UV sensitive and accumulate damage throughout nuclei when unshielded by their mothers in utero. These findings identify embryonic dissipation of nuclear pores as a key step in heterochromatic LAD assembly, allowing older embryos to withstand exposure to UV irradiation.

developmental biology↗

bioRxiv2025

Rubisco is slow across the tree of life

Rubisco is the main gateway through which inorganic carbon enters the biosphere, catalyzing the vast majority of carbon fixation on Earth. This pivotal enzyme has long been observed to be kinetically constrained. Yet, this impression is based on kinetic measurements heavily focused on eukaryotic rubiscos, a rather conserved group of low genetic diversity. Moreover, the fastest rubiscos that we know of so far were found among the sparsely sampled prokaryotes. Could there be yet faster rubiscos among the uncharted regions of rubiscos phylogenetic diversity? Here, we perform a characterization of more than 250 rubiscos from a wide range of bacteria and archaea, thereby doubling the coverage of the diversity of this key enzyme. We assess the distribution of the carboxylation rates at saturating levels of CO2, and establish that rubisco is a relatively slow enzyme across the tree of life, never exceeding {approx}30 reactions per second at 30{degrees}C. We show that relatively faster subclades share similar evolutionary contexts, involving micro-oxygenic environments or a CO2 concentrating mechanism. Leveraging a simple machine learning model trained on this dataset, we predict the carboxylation rate for all {approx}68,000 sequenced rubisco variants found in nature to date. This study provides the largest and most diverse dataset of natural variants for an enzyme and their associated rates, establishing a solid benchmark for future efforts to predict catalytic rates from sequence data. SignificanceDiscovering a fast carboxylating rubisco has been a long-standing challenge in the scientific community, given its potential impact on sustainable food and fuel production. Yet, only a small fraction of rubiscos natural diversity has been kinetically characterized. Here, we present a large-scale kinetic survey covering the entire spectrum of rubiscos diversity found in nature. Focusing on genetic clusters with above-average rates, we show that rubiscos catalytic rate does not exceed {approx}30 reactions per second at 30{degrees}C. Supported by a machine-learning predictive model, we extend this finding to all sequenced natural variants. Our study provides the most comprehensive kinetic dataset for a single enzyme to date.

systems biology↗

bioRxiv2024

Pear flower and leaf microbiome dynamics during the naturally occurring spread of Erwinia amylovora

Erwinia amylovora is the causal pathogen of fire blight, a contagious disease that affects apple and pear trees and other members of the family Rosaceae. In this study, we investigated the population dynamics of the pear flower microbiome in an agricultural setting during the naturally occurring infection of E. amylovora. Five potential factors were considered: collection date, the flowers phenological stage, location on the tree, location within the orchard, and pear cultivar. The phenological stage and the collection date were identified as the most important factors associated with pear flower microbiome composition. The location of the tree in the orchard and the flowers location on the tree had a marginal effect on the microbiome composition. The leaf microbiome reflected that of the abundant phenological stage on each date. The flower microbiome shifted towards E. amylovora, dominating the community as time and phenological stages progressed, leading to decreased community diversity. The strain population of E. amylovora remained similar throughout the entire collection period. In contrast, other taxa, including Pseudomonas, Pantoea, Lactobacillus, and Sphingomonas, were represented by dozens of amplicon sequence variants (ASVs), and different succession patterns in their populations were observed. Some of the taxa identified include known antagonists to E. amylovora. Overall, our results suggest that flower physiology and the interaction with the environment are strongly associated with the pear flower microbiome and should be considered separately. Strain succession patterns for the different taxa under E. amylovora spread may help in choosing candidates for antagonist-based treatments for fire blight. ImportanceThe spread of pathogens in plants is an important ecological phenomenon and has a significant economic impact on agriculture. Flowers serve as the entry point for E. amylovora, but members of the flower microbiome can inhibit or slow down the proliferation and penetration of the pathogen. Knowledge about leaf and flower microbiome response to the naturally occurring spread of E. amylovora is still lacking. The current study is the first to describe the flower microbiome dynamics during the naturally occurring infection of E. amylovora. Unlike previous studies, our experiment design enabled us to evaluate the contribution of five important environmental parameters to the community composition. We identified different strain succession patterns across different taxa in the flower consortia throughout the season. These results contribute to our understanding of plant microbial ecology during pathogen spread and can help to improve biological treatments for these diseases.

microbiology↗

bioRxiv2023

A metabolic modeling-based framework for predicting trophic dependencies in native rhizobiomes of crop plants

The exchange of metabolites (i.e., metabolic interactions) between bacteria in the rhizosphere determines various plant-associated functions. Systematically understanding the metabolic interactions in the rhizosphere, as well as in other types of microbial communities, would open the door to the optimization of specific pre-defined functions of interest, and therefore to the harnessing of the functionality of various types of microbiomes. However, mechanistic knowledge regarding the gathering and interpretation of these interactions is limited. Here, we present a framework utilizing genomics and constraint based modeling approaches, aiming to interpret the hierarchical trophic interactions in the soil environment. 243 genome-scale metabolic models of bacteria associated with a specific disease suppressive vs disease conducive apple rhizospheres were drafted based on genome resolved metagenomes, comprising an in-silico native microbial community. Iteratively simulating microbial community members growth in a metabolomics-based apple root-like environment produced novel data on potential trophic successions, used to form a network of communal trophic dependencies. Network-based analyses have characterized interactions associated with beneficial vs non-beneficial microbiome functioning, pinpointing specific compounds and microbial species as potential disease supporting and suppressing agents. This framework provides a means for capturing trophic interactions and formulating a range of testable hypotheses regarding the metabolic capabilities of microbial communities within their natural environment. Essentially, it can be applied to different environments and biological landscapes, elucidating the conditions for the targeted manipulation of various microbiomes, and the execution of countless predefined functions.

microbiology↗

bioRxiv2023

Systematic exploration of prokaryotic form I rubisco maximal carboxylation rates

Autotrophy is the basis for complex life on Earth. Central to this process is rubisco - the enzyme that catalyzes almost all carbon fixation on the planet. Yet, with only a small fraction of rubisco diversity kinetically characterized so far, the underlying biological factors driving the evolution of fast rubiscos in nature remain unclear. We conducted a high-throughput kinetic characterization of over 100 bacterial form I rubiscos, the most ubiquitous group of rubisco sequences in nature, to uncover the determinants of rubiscos carboxylation velocity. We show that the presence of a carboxysome CO2 concentrating mechanism correlates with faster rubiscos with a median 5-fold higher rate. In contrast to prior studies, we find that rubiscos originating from -cyanobacteria exhibit the highest carboxylation rates among form I enzymes ({approx}10 s-1 median versus <7 s-1 in other groups). Our study systematically reveals biological and environmental properties associated with kinetic variation across rubiscos from nature.

systems biology↗

bioRxiv2023

Structure and function of a hexameric cyanophycin synthetase 2

Cyanophycin is a natural polymer composed of a poly-aspartate backbone with arginine attached to each of the aspartate sidechains. Produced by a wide range of bacteria, which mainly use it as a store of fixed nitrogen, it has many promising industrial applications. Cyanophycin can be synthesized from the amino acids Asp and Arg by the widespread cyanophycin synthetase 1 (CphA1), or from the dipeptide {beta}-Asp-Arg by the cyanobacterial enzyme cyanophycin synthetase 2 (CphA2). CphA2 enzymes display a range of oligomeric states, from dimers to dodecamers. Recently, the crystal structure of a CphA2 dimer was solved but could not be obtained in complex with substrate. Here, we report cryo-EM structures of the hexameric CphA2 from Stanieria sp. at ~2.8 [A] resolution, both with and without ATP and cyanophycin. The structures show a trimer-of-dimers hexameric architecture, and substrate-binding interactions that are similar to those of CphA1. Mutagenesis experiments demonstrate the importance of several conserved substrate-binding residues. We also find that a Q416A/R528G double mutation prevents hexamer formation and use this double mutant to show that hexamerization augments the rate of cyanophycin synthesis. Together, these results increase our mechanistic understanding of how an interesting green polymer is biosynthesized.

biochemistry↗

bioRxiv2023

Inflammation and bacteriophages affect DNA inversion states and functionality of the gut microbiota

Reversible genomic DNA-inversions control expression of numerous bacterial molecules in the human gut, but how this relates to disease remains uncertain. By analyzing metagenomic samples from six human Inflammatory Bowel Disease cohorts combined with mice experimentation, we identified multiple invertible regions where a particular orientation was correlated with disease. These include the promoter of the anti-inflammatory polysaccharide-A (PSA) of Bacteroides fragilis, which is mostly oriented OFF during inflammation but is present in the ON orientation when inflammation is resolved. We further detected increased abundances of B. fragilis-associated bacteriophages in patients with the PSA OFF orientation, and a significant reduction in the frequency of the ON orientation, in the presence of the B. fragilis-associated bacteriophage, thereby altering the bacterial induced immune modulation. Altogether, we reveal dynamic and reversible bacterial phase-variations driven both by bacteriophages and the host inflammatory state, signifying bacterial functional plasticity during inflammation and opening future research avenues.

microbiology↗

bioRxiv2023

Bioinformatics of cyanophycin metabolism genes and characterization of promiscuous isoaspartyl dipeptidases that catalyze the final step of cyanophycin degradation

Cyanophycin is a bacterial biopolymer used for storage of fixed nitrogen. It is composed of a backbone of L-aspartate residues with L-arginines attached to each of their side chains. Cyanophycin is produced by cyanophycin synthetase 1 (CphA1) using Arg, Asp and ATP, and is degraded in two steps. First, cyanophycinase breaks down the backbone peptide bonds, releasing {beta}-Asp-Arg dipeptides. Then, these dipeptides are broken down into free Asp and Arg by enzymes with isoaspartyl dipeptidase activity. Two bacterial enzymes are known to possess promiscuous isoaspartyl dipeptidase activity: isoaspartyl dipeptidase (IadA) and isoaspartyl aminopeptidase (IaaA). We performed a bioinformatic analysis to investigate whether genes for cyanophycin metabolism enzymes cluster together or are spread around the microbial genomes. Many genomes showed incomplete contingents of known cyanophycin metabolizing genes. Cyanophycin synthetase and cyanophycinase are usually clustered together when recognizable genes for each are found within a genome. Cyanophycinase and isoaspartyl dipeptidase genes typically cluster within genomes lacking cphA1. About one-third of genomes with genes for CphA1, cyanophycinase and IaaA show these genes clustered together, while the proportion is around one-sixth for CphA1, cyanophycinase and IadA. We used X-ray crystallography and biochemical studies to characterize an IadA and an IaaA from two such clusters. The enzymes retained their promiscuous nature, showing that being associated with cyanophycin-related genes did not make them specific for {beta}-Asp-Arg dipeptides derived from cyanophycin degradation.

biochemistry↗

bioRxiv2023

Heterologous production of cyanophycin with Tatumella morbirosei cyanophycin synthetase

Microbial production of biopolymers represents a promising, sustainable alternative to current approaches for plastic production. Cyanophycin synthetase 1 (CphA1) produces cyanophycin - an attractive biopolymer consisting of a poly-L-aspartic acid backbone decorated with L-arginine side groups. In this work, a series of CphA1 enzymes from different bacteria were screened for heterologous cyanophycin production in engineered Escherichia coli, from which it was found that CphA1 from Tatumella morbirosei (TmCphA1) was especially productive. TmCphA1 was capable of supporting up to ~2-fold greater yields of insoluble cyanophycin than any other tested CphA1 enzymes, including 10.8-times more than CphA1 from Synechocystis sp. PCC6308. Finally, using a bench-scale bioreactor, cyanophycin production by TmCphA1-expressing E. coli reached up to 1.9 g per liter of culture by 48 h.

molecular biology↗

bioRxiv2022

Differences in homologous recombination and maintenance of heteropolyploidy between Haloferax volcanii and Haloferax mediterranei

Polyploidy, the phenomenon of having more than one copy of the genome in an organism, is common among haloarchaea. While providing short-term benefits for DNA repair, polyploidy is generally regarded as an evolutionary trap that by the notion of the Mullers ratchet will inevitably conclude in the species decline or even extinction due to a gradual reduction in fitness. In most reported cases of polyploidy in archaea, the genetic state of the organism is considered as homoploidy i.e. all copies of the genome are identical. Here we demonstrate that while this is indeed the prevalent genetic status in the halophilic archaeon H. volcanii, its close relative H. mediterranei maintains a prolonged heteroploidy state in a non-selective environment once a second allele is introduced. Moreover, a strong genetic linkage was observed between two distant loci in H. mediterranei indicating a low rate of homologous recombination while almost no such linkage was shown in H. volcanii indicating a high rate of recombination in the latter species. We suggest that H. volcanii escapes Mullers ratchet by means of an effective chromosome-equalizing gene-conversion mechanism facilitated by highly active homologous recombination, whereas H. mediterranei must elude the ratchet via a different, yet to be elucidated mechanism.

genetics↗

bioRxiv2018

Hydrogen-based metabolism - An ancestral trait in lineages sibling to the Cyanobacteria

The metabolic machinery from which microbial aerobic respiration evolved is tightly linked to the origins of oxygenic Cyanobacteria (Oxyphotobacteria). Even though the majority of Oxyphotobacteria are photoautotrophs and can use carbohydrates with oxygen (O2) as the electron acceptor, all are fermenters under dark anoxic conditions. Studies suggest that the ancestor of Oxyphotobacteria may have used hydrogen (H2) as an electron donor and that two types of NiFe hydrogenases are essential for its oxidation. Melainabacteria and Sericytochromatia, close phylogenetic neighbors to Oxyphotobacteria comprise fermentative and aerobic representatives, or organisms capable of both. Margulisbacteria (candidate divisions RBX-1 and ZB3) and Saganbacteria (candidate division WOR-1), a novel cluster of bacteria phylogenetically related to Melainabacteria, Sericytochromatia and Oxyphotobacteria may further constrain the metabolic platform in which oxygenic photosynthesis and aerobic respiration arose. Here, we predict the metabolisms of Margulisbacteria and Saganbacteria from new and published metagenome-assembled genomes (MAGs) and single amplified genomes (SAGs), and compare them to their phylogenetic neighbors. Sediment-associated Margulisbacteria are predicted to have a fermentation-based metabolism featuring a variety of hydrogenases, a nitrogenase for nitrogen (N2) fixation, and electron bifurcating complexes involved in cycling of ferredoxin and NAD(P)H. Overall, the genomic features suggest the capacity for metabolic fine-tuning under strictly anoxic conditions. In contrast, the genomes of Margulisbacteria from the ocean ecosystem encode an electron transport chain that supports aerobic growth. Similarly, some Saganbacteria genomes encode various hydrogenases, and others may have the ability to use O2 under certain conditions via a putative novel type of heme copper O2 reductase. Like Melainabacteria and Sericytochromatia, Margulisbacteria and Saganbacteria have diverse energy metabolisms capable of fermentation, and aerobic or anaerobic respiration. In summary, our findings support the hypothesis that the ancestor of these groups was an anaerobe in which fermentation and H2 metabolism were central metabolic features. Our genomic data also suggests that contemporary lineages sibling to the Oxyphotobacteria may have acquired the ability to use O2 as a terminal electron acceptor under certain environmental conditions.

microbiology↗

bioRxiv2018

A novel oceanic uncultured temperate cyanophage lineage

Marine cyanobacteria are important contributors to primary production in the ocean and their viruses (cyanophages) affect the ocean microbial communities. Despite reports of lysogeny in marine cyanobacteria, the genome sequence of such temperate cyanophages remains unknown although genomic analysis indicate potential for lysogeny in certain marine cyanophages. Using assemblies from Red Sea and Tara Oceans metagenomes, we recovered genomes of a novel uncultured marine cyanophage lineage, which contain, in addition to common cyanophage genes, a phycobilisome degradation protein NblA, an integrase and a split DNA polymerase. The DNA polymerase forms a monophyletic clade with a DNA polymerase from a genomic island in Synechococcus WH8016. The island contains a relic prophage that does not resemble any previously reported cyanophage but shares several genes with the newly identified cyanophages reported here. Metagenomic recruitment indicates that the novel cyanophages are widespread, albeit at low abundance. Here we describe a novel potentially lysogenic cyanophage family, their abundance and distribution in the marine environment.\n\nOriginality-Significance StatementMarine cyanobacteria are major contributors to primary production in the ocean. Despite reports of lysogeny in marine cyanobacteria, genomes from lysogenic marine cyanophages have not been reported yet. Using metagenomics assemblies, we recovered complete genomes of a novel uncultured marine cyanophage lineage. Remarkably, the DNA polymerase of these uncultured phages forms a monophyletic clade with the DNA polymerase from a genomic island in Synechococcus WH8016. The genomic island contains a putative relic prophage that does not resemble any known cultured cyanophage but shares several genes with the newly identified cyanophage family. These findings provide both phylogenomic and abundance estimates that are missing from current ecological models of this important group of marine viruses.

microbiology↗

bioRxiv2017

Cyanophage-encoded lipid-desaturases: oceanic distribution, diversity and function

Cyanobacteria are among the most abundant photosynthetic organisms in the oceans; viruses infecting cyanobacteria (cyanophages) can alter cyanobacterial populations, and therefore affect the local food web and global biochemical cycles. These phages carry auxiliary metabolic genes (AMGs), which rewire various metabolic pathways in the infected host cell, resulting in increased phage fitness. Coping with stress resulting from photodamage appears to be a central necessity of cyanophages, yet the overall mechanism is poorly understood. Here we report a novel, widespread cyanophage AMG, encoding a fatty acid desaturase (FAD), found in two genotypes with distinct geographical distribution. FADs are capable of modulating the fluidity of the hosts membrane, a fundamental stress response in living cells. We show that both viral fatty acid desaturases (vFADs) families are {Delta}9 lipid desaturases, catalyzing the desaturation at carbon 9 in C16 fatty acid chains. In addition, we present the first fatty acid profiling for marine cyanobacteria, which suggests a unique desaturation pathway of medium to long chain fatty acids no longer than C16, in accordance to the vFADs activity. Our findings suggest that cyanophages fiddle with the infected hosts cell, leading to increased photoprotection and potentially enhancing viral-encoded photosynthetic proteins, resulting in a new viral metabolic network.

microbiology↗

bioRxiv2017

Novel Abundant Oceanic Viruses of Uncultured Marine Group II Euryarchaeota Identified by Genome-Centric Metagenomics

Marine Group II Euryarchaeota (MGII) are among the most abundant microbes in the oceanic surface waters. So far, however, representatives of MGII have not been cultivated, and no viruses infecting these organisms have been described. Here we present complete genomes for 3 distinct groups of viruses assembled from metagenomic sequence datasets highly enriched for MGII. These novel viruses, which we denote Magroviruses, possess double-stranded DNA genomes of 65 to 100 kilobase in size that encode a structural module characteristic of head-tailed viruses and, unusually for archaeal and bacterial viruses, a nearly complete replication apparatus of apparent archaeal origin. The newly identified Magroviruses are widespread and abundant, and therefore are likely to be major ecological agents.

microbiology↗