Biology subjects

Michael, T.

Publications and source records attributed to Michael, T..

7 recordsLinked to original sources

bioRxiv2022

Characterization of EDS1-independent plant defense responses against bacterial pathogens using Duckweed/Pseudomonas pathosystems

ENHANCED DISEASE SUSCEPTIBILITY 1 (EDS1) mediates the induction of defense responses against pathogens in most land plants. However, it has recently been shown that a few species have lost EDS1. It is unknown how defense against disease unfolds and evolves in the absence of EDS1. Here we utilize duckweeds; a collection of aquatic species that lack EDS1, to investigate this question. We successfully established duckweed-Pseudomonas pathosystems and were able to characterize pathogen-induced responses in an immune system that lacks the EDS1 signaling pathway. We show that the copy number of infection-associated genes and the infection-induced transcriptional responses of duckweeds differ from that of other model species. Moreover, we show that the conservation of canonical Microbe Triggered Immunity and Effector Triggered Immunity pathways varies between duckweed species. This work shows that pathogen defense has evolved along different trajectories and uncovers alternative genomic and transcriptional reprogramming. Specifically, the miAMP1 domain containing proteins, which are absent in Arabidopsis, show pathogen responsive upregulation in duckweeds. Despite such divergence between Arabidopsis and duckweed species, we find evidence for the conservation of upregulation of certain genes and the role of hormones in response to disease. Our work highlights the importance of expanding the pool of model species to study defense responses that have evolved in the plant kingdom, including those independent of EDS1.

plant biology↗

bioRxiv2021

LoopViz: A uLoop Assembly Clone Verification Tool for Nanopore Sequencing Reads

Cloning has been an integral part of most laboratory research questions and continues to be an essential tool in defining the genetic elements determining life. Cloning can be difficult and time consuming as each plasmid is unique to a particular project and each sequence must be carefully selected, cloned and sequenced to determine correctness. Loop assembly (uLOOP) is a recursive, Golden Gate-like assembly method that allows rapid cloning of domesticated DNA fragments to robustly refactor novel pathways. With uLOOP methodologies, one can clone several sequences directionally to generate a library of transcriptional units (TUs) in plasmids within a single reaction but analysis of the plasmid population has been impeded by current sequencing and analysis methods. Here we develop LoopViz, a quality control tool that quantifies and visualizes results from assembly reactions using long-read Oxford Nanopore Technologies (ONT) sequencing. LoopViz identifies full length reads originating from a single plasmid in the population, and visualizes them in terms of a user input DNA fragments file, and provides QC statistics. This methodology enables validation and analysis of cloning and sequencing reactions in less than a day, determination of the entire plasmids sequence, and sequencing through repetitive meta-regions that cannot be meaningfully assembled. Finally, LoopViz represents a new paradigm in determining plasmid sequences that is rapid, cost-effective and performed in-lab. LoopViz is made publicly available at https://gitlab.com/marielelensink325/loopseq

bioinformatics↗

bioRxiv2020

Unusual predominance of maintenance DNA methylation in Spirodela polyrhiza

5-methylcytosine (5mC) is a modified base often described as necessary for the proper regulation of genes and transposons and for the maintenance of genome integrity in plants. However, the extent of this dogma is limited by the current phylogenetic sampling of land plant species diversity. Here, we report that a monocot plant, Spirodela polyrhiza, has lost CG gene body methylation, genome-wide CHH methylation, and the presence or expression of several genes in the highly conserved RNA-directed DNA methylation (RdDM) pathway. It has also lost the CHH methyltransferase CHROMOMETHYLASE 2. Consequently, the transcriptome is depleted of 24-nucleotide, heterochromatic, small interfering RNAs that act as guides for the deposition of 5mC to RdDM-targeted loci in all other currently sampled angiosperm genomes. Although the genome displays low levels of genome-wide 5mC primarily at LTR retrotransposons, CG maintenance methylation is still functional. In contrast, CHG methylation is weakly maintained even though H3K9me2 is present at loci dispersed throughout the euchromatin and highly enriched at regions likely demarcating pericentromeric regions. Collectively, these results illustrate that S. polyrhiza is maintaining CG and CHG methylation mostly at repeats in the absence of small RNAs. S. polyrhiza reproduces rapidly through clonal propagation in aquatic environments, which we hypothesize may enable low levels of maintenance methylation to persist in large populations. Significance StatementDNA methylation is a widespread chromatin modification that is regularly found in all plant species. By examining one aquatic duckweed species, Spirodela polyrhiza, we find that it has lost highly conserved genes involved in methylation of DNA at sites often associated with repetitive DNA, and within genes, however DNA methylation and heterochromatin is maintained during cell division at other sites. Consequently, small RNAs that normally guide methylation to silence repetitive DNA like retrotransposons are diminished. Despite the loss of a highly conserved methylation pathway, and the reduction of small RNAs that normally target repetitive DNA, transposons have not proliferated in the genome, perhaps due in part to the rapid, clonal growth lifestyle of duckweeds.

plant biology↗

bioRxiv2020

A new Spirodela polyrhiza genome and proteome reveal a conserved chromosomal structure with high abundances of proteins favoring energy production

Duckweeds are a monophyletic group of rapidly reproducing aquatic monocots in the Lemnaceae family. Spirodela polyrhiza, the Greater Duckweed, has the largest body plan yet the smallest genome size in the family (1C = 150 Mb). Given their clonal, exponentially fast reproduction, a key question is whether genome structure is conserved across the species in the absence of meiotic recombination. We generated a highly contiguous, chromosome-scale assembly of Spirodela polyrhiza line Sp7498 using Oxford Nanopore plus Hi-C scaffolding (Sp7498_HiC) which is highly syntenic with a related line (Sp9509). Both the Sp7498_HiC and Sp9509 genome assemblies reveal large chromosomal misorientations in a recent PacBio assembly of Sp7498, highlighting the necessity of orthogonal long-range scaffolding techniques like Hi-C and BioNano optical mapping. Shotgun proteomics of Sp7498 verified the expression of [~]2,250 proteins and revealed a high abundance of proteins involved in photosynthesis and carbohydrate metabolism among other functions. In addition, a strong increase in chloroplast proteins was observed that correlated to chloroplast density. This Sp7498_HiC genome was generated cheaply and quickly with a single Oxford Nanopore MinION flow cell and one Hi-C library in a classroom setting. Combining these data with a mass spectrometry-generated proteome illustrates the utility of duckweed as a model for genomics- and proteomics-based education.

plant biology↗

bioRxiv2020

Gapless assembly of maize chromosomes using long read technologies

Creating gapless telomere-to-telomere assemblies of complex genomes is one of the ultimate challenges in genomics. We used long read technologies and an optical map based approach to produce a maize genome assembly composed of only 63 contigs. The B73-Ab10 genome includes gapless assemblies of chromosome 3 (236 Mb) and chromosome 9 (162 Mb), multiple highly repetitive centromeres and heterochromatic knobs, and 53 Mb of the Ab10 meiotic drive haplotype.

bioinformatics↗

bioRxiv2019

Sleep’s impact on emotional memory: A meta-analysis of whole-night, nap, and REM sleep effects

Numerous studies have shown that sleep enhances the consolidation of episodic memory. However, it remains unclear whether this consolidation benefit is moderated by the emotional valence of the learned material. To clarify whether sleep selectively enhances the consolidation of emotional material, we conducted a meta-analysis including N = 1,059 observations. Overall, our results do not support this hypothesis. When only studies with a sleep-group wake-group comparison were included in the analysis (k = 22), the retention advantage for emotional over neutral material was not significantly different between sleep and wake groups. When studies initially lacking a wake-control group were included in the analysis after statistical estimation of wake-group parameters, the retention advantage for emotional material was significantly larger in wake-groups than in sleep-groups (k = 34). Interestingly, however, an additional analysis of k = 8 studies investigating the selective effects of rapid-eye-movement sleep and slow-wave sleep on emotional memory consolidation provided evidence for a selective enhancement of emotional over neutral memory consolidation after rapid-eye-movement sleep compared to slow-wave sleep. These results suggest that sleep does not generally enhance emotional memory consolidation over neutral memory consolidation. However, specific sleep stages might preferentially enhance consolidation of emotional and neutral material, respectively.

neuroscience↗

bioRxiv2019

D-Cycloserine as Adjunct to Brief Computerised CBT for Spider Fear: Effects on Fear, Behaviour, and Cognitive Biases

In anxiety disorders, cognitive behavioural therapy (CBT) improves information-processing biases such as implicit fear evaluations and avoidance tendencies, which predicts treatment response, so they might constitute important treatment targets. This study investigated (i) whether information-processing biases changed following single-session computerised CBT for spider fear, and (ii) whether this effect could be augmented by administration of D-cycloserine (DCS). Spider-fearful individuals were randomized to receiving 250mg of DCS (n=21) or placebo (n=17) and spider fear was assessed using self-report, behavioural, and information-processing (Extrinsic Affective Simon Task & Approach Avoidance Task) measures. Linear mixed-effects analyses indicated improvements on self-report and behavioural spider fear following CBT, but not on cognitive bias measures. There was no evidence of an augmentation effect of DCS on any outcome. Cognitive biases at 1-day were not predictive of 1-month follow-up spider fear. These findings provide no evidence for information-processing biases relating to CBT response or augmentation with DCS.

neuroscience↗