Biology subjects

Bracht, J. R.

Publications and source records attributed to Bracht, J. R..

6 recordsLinked to original sources

bioRxiv2023

Evolution of a biological thermocouple by adaptation of cytochrome c oxidase in asubterrestrial metazoan

In this study we report a naturally evolved temperature-sensing electrical regulator in the cytochrome c oxidase of the Devil Worm, Halicephalobus mephisto. This extremophile metazoan was isolated 1.3 km underground in a South African goldmine, where it adapted to heat and potentially to hypoxia, making its mitochondrial sequence a likely target of adaptational change. We obtained the full mitochondrial genome sequence of this organism, and show through dN/dS analysis statistically robust evidence of positive selection in H. mephisto cytochrome c oxidase subunits. Seventeen of these positively-selected amino acid substitutions were localized in proximity to the H- and K-pathway proton channels of the complex. Surprisingly, the H. mephisto cytochrome c oxidase proton pump completely shuts down at low temperatures (20{degrees}C) leading to approximately a 4.8-fold reduction in the transmembrane proton gradient voltage ({Delta}{Psi}m) compared to optimal temperature (37{degrees}C). Direct measurement of oxygen consumption found a corresponding 4.7-fold drop at 20{degrees}C compared to 37{degrees}C. Correspondingly, the lifecycle of H. mephisto takes four-fold longer at the low temperature compared to higher. This elegant evolutionary adaptation creates a finely-tuned mitochondrial temperature sensor, allowing this ectothermic organism to maximize its reproductive success in varying environmental temperatures. Our study shows that evolutionary innovation may remodel core metabolism to make it more accurately map onto environmental variation.

evolutionary biology↗

bioRxiv2022

Surface detection of SARS-CoV-2 by lateral flow LAMP

Slowing the transmission of SARS-CoV-2 requires rapid and accurate diagnostic testing. Toward this end, loop-mediated isothermal amplification (LAMP), an isothermal genomic detection method, offers great promise but the readout tends to be difficult because it does not generate linear DNA products. Rapid antigen tests are coupled to lateral flow strips, with one (negative) or two (positive) bands providing simple rapid readout, but are less sensitive than genomic amplification methods. To address the need for a genomic amplification method that can be visualized on a lateral flow strip, we developed a novel strand-displacement probe. In this work we validate this pipeline for purified RNA, intact virus, and even virus deposited onto a surface. We demonstrate robust sensitivity (100 genomic copies) and and we demonstrate the utility of our assay as a surveillance system, with the capability to detect viral particles from surfaces, even after a week of complete dry-down. Our innovation couples the diagnostic advantages of a nucleic acid amplification test (NAAT) with the simplicity of lateral-flow readouts.

genetics↗

bioRxiv2020

Epigenetic regulation of Neuregulin-1 tunes white adipose stem cell differentiation

Expansion of subcutaneous adipose tissue by differentiation of new adipocytes has been linked to improvements in metabolic health. However, an expandability limit has been observed wherein new adipocytes cannot be produced, the existing adipocytes become enlarged (hypertrophic) and lipids spill over into ectopic sites. Inappropriate ectopic storage of these surplus lipids in liver, muscle, and visceral depots has been linked with metabolic dysfunction. Here we show that Neuregulin-1 (NRG1) serves as a regulator of adipogenic differentiation in subcutaneous primary human stem cells. We further demonstrate that DNA methylation modulates NRG1 expression in these cells, and a 3-day exposure of stem cells to a recombinant NRG1 peptide fragment is sufficient to reprogram adipogenic cellular differentiation to higher levels. These results define a novel molecular adipogenic rheostat with potential implications for the expansion of adipose tissue in vivo.

cell biology↗

bioRxiv2019

The genome of a subterrestrial nematode reveals an evolutionary strategy for adaptation to heat

The nematode Halicephalobus mephisto was originally discovered inhabiting a deep terrestrial aquifer 1.3 km underground. H. mephisto can thrive under conditions of abiotic stress including heat and minimal oxygen, where it feeds on a community of both chemolithotrophic and heterotrophic prokaryotes in an unusual ecosystem isolated from the surface biosphere. Here we report the comprehensive genome and transcriptome of this organism, identifying a signature of adaptation: an expanded repertoire of 70 kilodalton heat-shock proteins (Hsp70) and avrRpt2 induced gene 1 (AIG1) proteins. We find that positive selection has driven the expansion of Hsp70 genes, which are also transcriptionally induced upon growth under heat stress. We further show that AIG1 may have been acquired by horizontal gene transfer (HGT) from a rhizobial fungus. Over one-third of the genes of H. mephisto are novel, highlighting the divergence of this nematode from other sequenced organisms. This work sheds light on the genomic strategies of adaptation to heat in the first complete subterrestrial eukaryotic genome.

genomics↗

bioRxiv2018

Reconstitution of eukaryotic chromosomes and manipulation of DNA N6-methyladenine alters chromatin and gene expression

DNA N6-adenine methylation (6mA) has recently been reported in diverse eukaryotes, spanning unicellular organisms to metazoans. Yet the functional significance of 6mA remains elusive due to its low abundance, difficulty of manipulation within native DNA, and lack of understanding of eukaryotic 6mA writers. Here, we report a novel DNA 6mA methyltransferase in ciliates, termed MTA1. The enzyme contains an MT-A70 domain but is phylogenetically distinct from all known RNA and DNA methyltransferases. Disruption of MTA1 in vivo leads to the genome-wide loss of 6mA in asexually growing cells and abolishment of the consensus ApT dimethylated motif. Genes exhibit subtle changes in chromatin organization or RNA expression upon loss of 6mA, depending on their starting methylation level. Mutants fail to complete the sexual cycle, which normally coincides with a peak of MTA1 expression. Thus, MTA1 functions in a developmental stage-specific manner. We determine the impact of 6mA on chromatin organization in vitro by reconstructing complete, full-length ciliate chromosomes harboring 6mA in native or ectopic positions. Using these synthetic chromosomes, we show that 6mA directly disfavors nucleosomes in vitro in a local, quantitative manner, independent of DNA sequence. Furthermore, the chromatin remodeler ACF can overcome this effect. Our study identifies a novel MT-A70 protein necessary for eukaryotic 6mA methylation and defines the impact of 6mA on chromatin organization using epigenetically defined synthetic chromosomes.\n\nHighlightsO_LIThe MT-A70 protein MTA1 mediates DNA N6-adenine methylation in Oxytricha\nC_LIO_LIMTA1 mutants exhibit subtle changes in nucleosome organization and transcription in vivo\nC_LIO_LI6mA directly disfavors nucleosome occupancy in natural and synthetic chromosomes in vitro\nC_LIO_LIDe novo synthesis of complete, epigenetically defined Oxytricha chromosomes\nC_LI

biochemistry↗

bioRxiv2017

Synthesis of a eukaryotic chromosome reveals a role for N6-methyladenine in nucleosome organization

Biochemical studies of chromatin have typically used either artificial DNA templates with unnaturally high affinity for histones, or small genomic DNA fragments deprived of their cognate physical environment. It has thus been difficult to dissect chromatin structure and function within fully native DNA substrates. Here, we circumvent these limitations by exploiting the minimalist genome of the eukaryote Oxytricha trifallax, whose notably small ~3kb chromosomes mainly encode single genes. Guided by high-resolution epigenomic maps of nucleosome organization, transcription, and DNA N6-methyladenine (m6dA) locations, we reconstruct full-length Oxytricha chromosomes in vitro and use these synthetic facsimiles to dissect the influence of m6dA and histone post-translational modifications on nucleosome organization. We show that m6dA directly disfavors nucleosomes in a quantitative manner, leading to local decreases in nucleosome occupancy that are synergistic with histone acetylation. The effect of m6dA can be partially reversed by the action of an ATP-dependent chromatin remodeler. Furthermore, erasing m6dA marks from Oxytricha chromosomes leads to proportional increases in nucleosome occupancy across the genome. This work showcases Oxytricha chromosomes as powerful yet practical models for studying eukaryotic chromatin and transcription in the context of biologically relevant DNA substrates.\n\nHighlightsO_LIDe novo synthesis of complete, epigenetically defined Oxytricha chromosomes\nC_LIO_LIEpigenomic profiles of chromatin organization in Oxytrichas miniature chromosomes\nC_LIO_LIm6dA directly disfavors nucleosome occupancy in natural and synthetic chromosomes\nC_LIO_LIHistone acetylation and chromatin remodelers temper the impact of m6dA on chromatin\nC_LI

biochemistry↗