Biology subjects

Jackson, D.

Publications and source records attributed to Jackson, D..

4 recordsLinked to original sources

bioRxiv2018

On the design of CRISPR-based single cell molecular screens

Several groups recently reported coupling CRISPR/Cas9 perturbations and single cell RNA-seq as a potentially powerful approach for forward genetics. Here we demonstrate that vector designs for such screens that rely on cis linkage of guides and distally located barcodes suffer from swapping of intended guide-barcode associations at rates approaching 50% due to template switching during lentivirus production, greatly reducing sensitivity. We optimize a published strategy, CROP-seq, that instead uses a Pol II transcribed copy of the sgRNA sequence itself, doubling the rate at which guides are assigned to cells to 94%. We confirm this strategy performs robustly and further explore experimental best practices for CRISPR/Cas9-based single cell molecular screens.

genomics↗

bioRxiv2017

The CLAVATA receptor FASCIATED EAR2 responds to different CLE peptides by signaling through different downstream effectors.

Meristems contain groups of indeterminate stem cells that are critical for organ initiation throughout plant development. The shoot apical meristem (SAM) maintains itself and initiates all shoot organs, such as leaves, floral organs and axillary branch meristems. Development and balanced proliferation of the SAM is regulated by a feedback loop between CLAVATA (CLV) and WUSCHEL (WUS) signaling. CLV signaling is initiated by secretion of the CLV3 peptide ligand, which is perceived directly or indirectly by a number of Leucine-Rich-Repeat (LRR) receptor-like kinases, including CLV1 and BARELY ANY MERISTEM (BAM) 1-3, and RECEPTOR-LIKE PROTEIN KINASE 2 (RPK2), as well as the receptor-like protein CLV2 in a complex with the CORYNE (CRN) pseudokinase. However, CLV2, and its maize ortholog FASCIATED EAR2 (FEA2) appear to function in signaling by several related CLV3/EMBRYO-SURROUNDING REGION (CLE) peptide ligands, including CLV3. Nevertheless, it remains unknown how CLV2 or FEA2 transmit specific signals from distinct CLE peptides. Here we show that FEA2 is involved in signaling from at least 2 distinct CLE peptides, ZmCLE7, a maize CLV3 ortholog, and ZmFON2-LIKE CLE PROTEIN1 (ZmFCP1), a newly identified CLE peptide functioning in SAM regulation. Signaling from these 2 different CLE peptides appears to be transmitted through 2 different candidate downstream effectors, COMPACT PLANT2 (CT2), the alpha subunit of the maize heterotrimeric G protein, and maize CRN. Our data provide a novel framework to understand how diverse signaling peptides can activate different downstream pathways through common receptor proteins.

plant biology↗

bioRxiv2017

Genetic fingerprinting of salmon louse (Lepeophtheirus salmonis) populations in the North-East Atlantic using a random forest classification approach

Caligid sea lice represent a significant threat to salmonid aquaculture worldwide. Lepeophtheirus salmonis is the predominant species that occurs in the Northern Hemisphere. Dispersal of sea lice between marine aquaculture sites and geographic regions is thought to occur rapidly via planktonic transport of larvae. Population genetic analyses have consistently shown minimal population genetic structure in North Atlantic L. salmonis, frustrating efforts to track louse populations, improve targeted control measures and understand local adaption to environmental conditions. The aim of this study was to test the power of reduced representation library sequencing (IIb-RAD sequencing) coupled with random forest machine learning algorithms to define markers for fine-scale discrimination of louse populations. We identified 1286 robustly supported SNPs among four L. salmonis populations from Ireland (N=2, 27 individuals), Scotland (N=1, 11 individuals) and North Norway (N=1, 12 individuals). Weak global structure (FSC = 0.018, p<0.0001) and only one significant pairwise FST comparison was observed (Scotland vs Kenmare Bay, (FST = 0.018, p<0.0001)) using all 1286 SNPs. The application of a random forest machine-learning algorithm identified 98 discriminatory SNPs that dramatically improved population assignment (DAPC assignment probability = 1), increased global Fsc = 0.098, (p<0.0001) and resulted in pairwise comparisons that all showed highly significant Fst-values (range = 0.081 - 0.096, p<0.0001). Out of 19 SNPs found to be under directional selection between populations, 12 corresponded to the discriminatory SNPs identified using random forest. Taken together our data suggest that L. salmonis SNP diversity exists with which it is possible to discriminate differences between nearby populations given suitable marker selection approaches, and that such differences might have an adaptive basis. We discuss these data in light of sea lice adaption to anthropogenic and environmental pressures as well as novel approaches to track and predict sea louse dispersal.

ecology↗

bioRxiv2017

Chromatin accessibility dynamics of myogenesis at single cell resolution

Over a million DNA regulatory elements have been cataloged in the human genome, but linking these elements to the genes that they regulate remains challenging. We introduce Cicero, a statistical method that connects regulatory elements to target genes using single cell chromatin accessibility data. We apply Cicero to investigate how thousands of dynamically accessible elements orchestrate gene regulation in differentiating myoblasts. Groups of co-accessible regulatory elements linked by Cicero meet criteria of \"chromatin hubs\", in that they are physically proximal, interact with a common set of transcription factors, and undergo coordinated changes in histone marks that are predictive of gene expression. Pseudotemporal analysis revealed a subset of elements bound by MYOD in myoblasts that exhibit early opening, potentially serving as the initial sites of recruitment of chromatin remodeling and histone-modifying enzymes. The methodological framework described here constitutes a powerful new approach for elucidating the architecture, grammar and mechanisms of cis-regulation on a genome-wide basis.

genomics↗