Biology subjects

Williams, D. J.

Publications and source records attributed to Williams, D. J..

9 recordsLinked to original sources

bioRxiv2023

Ribosomal DNA copy number is associated with body mass in humans and other mammals

Body mass results from a complex interplay between genetics and environment. The contribution of genetic variation to body mass has been extensively studied, but due to the technical limitations of platforms used for population scale studies, repetitive parts of the genome have not previously been considered. Here we applied genome-wide approaches to identify an association between adult body mass and the copy number (CN) of 45S-ribosomal DNA (rDNA). rDNA codes for the rRNA components of the ribosome and exists in hundreds of copies/cell in mammals. Inter-individual variation in rDNA CN has not previously been documented but has not been associated with a mammalian phenotype. Here, we show that germ-line inherited rDNA CN is associated with post-pubertal growth rate in rats and BMI in adult humans. rDNA CN variation is not associated with rRNA transcription rates in adult tissues, suggesting the mechanistic link occurs earlier in development. This is supported by our observation that it is growth rate and body mass, rather than obesity that are associated with copy number and that these phenotypes emerge prior to adulthood. We present hypotheses for future investigation into the mechanistic basis of this association.

genomics↗

bioRxiv2023

Behavioral control through the direct, focal silencing of neuronal activity

Voltage-gated sodium channel (NaV) activity underlies electrical signaling, synaptic release, circuit function, and, ultimately, behavior. Molecular tools that enable precise control of NaV subpopulations make possible temporal regulation of neuronal activity and cellular communication. To rapidly modulate NaV currents, we have rendered a potent NaV inhibitor, saxitoxin, transiently inert through chemical protection with a novel nitrobenzyl-derived photocleavable group. Light-induced uncaging of the photocaged toxin, STX-bpc, effects rapid inhibitor release and focal NaV block. We demonstrate the efficacy of this reagent for manipulating action potentials in mammalian neurons and brain slice and for altering locomotor behavior in larval zebrafish. Photo-uncaging of STX-bpc is a non-invasive, effective method for reversible, spatiotemporally precise tuning of NaV currents, application of which requires no genetic manipulation of the biological sample.

neuroscience↗

bioRxiv2022

Population History and Genome Wide Association Studies of Birth Weight in a Native High Altitude Ladakhi Population

Pathological low birth weight due to fetal growth restriction (FGR)is an important predictor of adverse obstetric and neonatal outcomes. It is more common amongst native lowlanders when gestating in the hypoxic environment of high altitude, whilst populations who have resided at high altitude for many generations are relatively protected. Genetic study of pregnant populations at high altitude permits exploration of the role of hypoxic in FGR pathogenesis, and perhaps of FGR pathogenesis more broadly. We studied the umbilical cord blood DNA of 316 neonates born to pregnant women managed at the Sonam Norboo Memorial Hospital, Ladakh (altitude 3540m) between February 2017-January 2019. Principal component, admixture and genome wide association studies (GWAS) were applied to dense single nucleotide polymorphism (SNP) genetic data, to explore ancestry and genetic predictors of low birth weight. Our findings support Tibetan ancestry in the Ladakhi population, with subsequent admixture with neighboring Indo-Aryan populations. Fetal growth protection was evident in Ladakhi neonates. Although no variants achieved genome wide significance, we observed nominal association of seven variants across genes (ZBTB38, ZFP36L2, HMGA2, CDKAL1, PLCG1) previously associated with birthweight.

genetics↗

bioRxiv2022

The phylogenomic landscape of the genus Serratia

The genus Serratia has been studied for over a century and includes clinically-important and diverse environmental members. Despite this, there is a paucity of genomic information across the genus and a robust whole genome-based phylogenetic framework is lacking. Here, we have assembled and analysed a representative set of 664 genomes from across the genus, including 215 historic isolates originally used in defining the genus. Phylogenomic analysis of the genus reveals a clearly-defined population structure which displays deep divisions and aligns with ecological niche, as well as striking congruence between historical biochemical phenotyping data and contemporary genomics data. We show that Serratia is a diverse genus which displays striking plasticity and ability to adapt to its environment, including a highly-varied portfolio of plasmids, and provide evidence of different patterns of gene flow across the genus. This work provides an essential platform for understanding the emergence of clinical and other lineages of Serratia.

microbiology↗

bioRxiv2021

Human stem cell model of HNF1A deficiency shows uncoupled insulin to C-peptide secretion with accumulation of abnormal insulin granules

Mutations in HNF1A cause Maturity Onset Diabetes of the Young type 3 (MODY3), the most prevalent form of monogenic diabetes. We generated stem cell-derived pancreatic endocrine cells from human embryonic stem cells (hESCs) with induced hypomorphic mutations in HNF1A. Using these cells, we show that HNF1A orchestrates a transcriptional program required for distinct aspects of {beta}-cell fate and function. During islet cell differentiation, HNF1A deficiency biases islet endocrine cells towards an -cell fate associated with PAX4 down-regulation. HNF1A- deficient {beta}-cells display impaired basal and glucose stimulated-insulin secretion in association with a reduction in CACNA1A and intracellular calcium levels, and impaired insulin granule exocytosis in association with SYT13 down-regulation. Knockout of PAX4, CACNA1A and SYT13 reproduce the relevant phenotypes. Reduction of insulin secretion is associated with accumulation of enlarged secretory granules, and altered stoichiometry of secreted insulin to C-peptide. In HNF1A deficient {beta}-cells, glibenclamide, a sulfonylurea drug used in the treatment of MODY3 patients, increases intracellular calcium to levels beyond those achieved by glucose, and restores C-peptide and insulin secretion to a normal stoichiometric ratio. To study HNF1A deficiency in the context of a human disease model, we also generated stem cell-derived pancreatic endocrine cells from two MODY3 patients induced pluripotent stem cells (iPSCs). While insulin secretion defects are constitutive in cells with complete HNF1A loss of function, {beta}-cells heterozygous for hypomorphic HNF1A mutations are initially normal, but lose the ability to secrete insulin and acquire abnormal stoichiometric secretion ratios. Importantly, the defects observed in these stem cell models are also seen in circulating proportions of insulin:C-peptide in nine MODY3 patients. One sentence of summaryDeficiency of the transcription factor HNF1A biases islet endocrine cell fate towards -cells, impairs intracellular calcium homeostasis and insulin exocytosis, alters the stoichiometry of insulin to C-peptide release, and leads to an accumulation of abnormal insulin secretory granules in {beta}-cells.

cell biology↗

bioRxiv2020

Regulation of intracellular signaling and neuron function by Bardet-Biedl Syndrome proteins in patient-specific iPSC-derived neurons

Bardet-Biedl Syndrome (BBS) is a rare autosomal recessive disorder caused by mutations in genes encoding components of the primary cilium and characterized by hyperphagic obesity. We developed a cellular model of BBS using induced pluripotent stem cell (iPSCs)-derived hypothalamic arcuate-like neurons. BBS mutations BBS1M390R and BBS10C91fsX95 did not affect neuron differentiation efficiency but caused morphological defects including impaired neurite outgrowth and longer primary cilia. Expression of intact BBS10 normalized cilia length. Single-cell RNA sequencing (scRNA-seq) of BBS1M390R hypothalamic neurons identified several down regulated pathways including insulin and cAMP signaling, and axon guidance. In agreement with scRNA-seq data, insulin-induced AKT phosphorylation at Thr308 was reduced in BBS1M390R and BBS10c91fsX95 human fibroblasts and iPSC-derived neurons, as well as in BBS10 knockdown iPSC-derived neurons. Overexpression of intact BBS10 fully restored insulin receptor tyrosine phosphorylation in BBS10c91fsX95 neurons. Mutations in BBS1 and BBS10 impaired leptin-mediated p-STAT3 activation in both human primary fibroblasts and iPSC-derived hypothalamic neurons. Correction of the BBS mutation by CRISPR rescued leptin signaling. POMC expression in BBS1M390R and BBS10 C91fsX95 iPSC-derived hypothalamic neurons was downregulated, as was hypothalamic Pomc in BBS1M390R knockin (KI) mice. In the aggregate, these data provide insights into the anatomic and functional mechanisms by which components of the BBsome in CNS primary cilia mediate effects on energy homeostasis.

cell biology↗

bioRxiv2020

Expression and function of murine WFDC2 in the respiratory tract.

WFDC2/HE4 encodes a poorly characterised secretory protein that shares structural similarity with multifunctional host defence proteins through possession of two conserved Whey Acidic Protein/four disulphide-core (WFDC) domains. WFDC2 is expressed in multiple epithelia and although its function remains unresolved, it is also overexpressed in a number of human cancers and has an established role as a cancer marker. Currently, little is known about the distribution of WFDC2 in the mouse and thus we have systematically analysed the mouse wfdc2 gene, its expression and distribution. We have used recombinant WFDC2 for functional studies. Wfdc2 is the most highly expressed family member in the lung and is enriched in the nasopharynx. Wfdc2 is the most highly expressed family member in differentiated epithelial cells isolated from the trachea, nasal passages and middle ear. Wfdc2 consists of 5 exons with exon 3 encoding an unstructured linker region that separates the two WFDC domains. This genomic organisation appears to be restricted to the Muridae and Cricetidae families of rodents. Similar to the situation in man, mouse wfdc2 can be alternatively spliced to yield a number of distinct transcripts that have the potential to generate a repertoire of distinct protein isoforms. We used immunohistochemistry to localise the proteins to tissues of the respiratory tract and head and neck regions. Although the protein was limited to epithelial cells of the respiratory tract and nasal and oral cavities, it was expressed in different cells in different regions suggesting expression is governed by a unique regulatory mechanism. Recombinant WFDC2 did not possess antiproteinase activity against trypsin or elastase and had no clear antimicrobial activity.

genomics↗

bioRxiv2020

DNA methylation covariation in human whole blood and sperm: implications for studies of intergenerational epigenetic effects

BackgroundEpidemiological studies suggest that paternal obesity may increase the risk of fathering small for gestational age offspring. Studies in non-human mammals suggest that such associations could be mediated by DNA methylation changes in spermatozoa that influence offspring development in utero. Human obesity is associated with differential DNA methylation in peripheral blood. It is unclear, however, whether this differential DNA methylation is reflected in spermatozoa. We profiled genome-wide DNA methylation using the Illumina MethylationEPIC array in matched human blood and sperm from lean (discovery n=47; replication n=21) and obese (n=22) males to analyse tissue covariation of DNA methylation, and identify whether this covariation is influenced by obesity. ResultsDNA methylation signatures of human blood and spermatozoa are highly discordant, and methylation levels are correlated at only a minority of CpG sites ([~]1%). While at the majority of these sites, DNA methylation appears to be influenced by genetic variation, obesity-associated DNA methylation in blood was not generally reflected in spermatozoa, and obesity did not influence covariation patterns. However, one cross-tissue obesity-specific hypermethylated site (cg19357369; chr4:2429884; P=8.95 x 10-8; beta=0.02) was identified, warranting replication and further investigation. When compared to a wide range of human somatic tissue samples (n=5,917), spermatozoa displayed differential DNA methylation in pathways enriched in transcriptional regulation. ConclusionsHuman sperm displays a unique DNA methylation profile that is highly discordant to, and practically uncorrelated with, that of matched peripheral blood. Obesity only nominally influences sperm DNA methylation, making it an unlikely mediator of intergenerational effects of metabolic traits.

genomics↗

bioRxiv2019

A new family of Type VI secretion system-delivered effector proteins displays ion-selective pore-forming activity

Type VI secretion systems (T6SSs) are nanomachines widely used by bacteria to compete with rivals. T6SSs deliver multiple toxic effector proteins directly into neighbouring cells and play key roles in shaping diverse polymicrobial communities. A number of families of T6SS-dependent anti-bacterial effectors have been characterised, however the mode of action of others remains unknown. Here we report that Ssp6, an anti-bacterial effector delivered by the Serratia marcescens T6SS, is an ion-selective pore-forming toxin. In vivo, Ssp6 inhibits growth by causing depolarisation of the inner membrane of intoxicated cells and also leads to increased outer membrane permeability, whilst reconstruction of Ssp6 activity in vitro demonstrated that it forms cation-selective pores. A survey of bacterial genomes revealed that Ssp6-like effectors are widespread in Enterobacteriaceae and often linked with T6SS genes. We conclude that Ssp6 represents a new family of T6SS-delivered anti-bacterial effectors, further diversifying the portfolio of weapons available for deployment during inter-bacterial conflict.

microbiology↗