Biology subjects

Mullen, S.

Publications and source records attributed to Mullen, S..

10 recordsLinked to original sources

bioRxiv2026

Establishment of microbial strains from the river, groundwater, and soil in the hyporheic zone is limited despite connectivity

Microbial diversity across ecosystems may be impacted by dispersal pathways and selection due to physiochemical conditions, and so it is hard to predict the extent of strain sharing across heterogeneous environmental compartments. Evaluation of patterns of strain-level overlap requires genome-level comparisons using extensive datasets collected over large spatial scales. Here, we sampled microbiomes beneath (hyporheic zone) and within the East River (Colorado, USA) and found little or no strain overlap at sites along the river corridor, despite connection by river flow, suggesting selection due to the local environment. Comparisons involving microbiomes from hillslopes and riparian zone soil yielded essentially no strain-level overlap with the microbiomes of river and hyporheic zones. We also sampled a nearby groundwater well and found near-perfect genotypic overlap with strains in one hyporheic zone location that exhibited active groundwater upwelling. Given an absence of direct connectivity between these two locations via discrete hydrologic flow paths, we conclude that groundwater strains are widely dispersed in the aquifer. As hyporheic zone microbiomes in zones with low river flow share no strains with groundwater microbiomes, we infer that strains introduced by groundwater mixing in the hyporheic zone are transitory. We conclude that despite evidence for mixing of river-and-hyporheic zone water, and river-and-groundwater on short time scales, establishment of transported strains in the hyporheic zone is minimal.

microbiology↗

bioRxiv2026

Increasing absolute prey community density protects aposematic models and their imperfect Batesian mimics: Evidence from Neotropical Adelpha butterflies

Batesian mimicry is a defensive adaptation where predators learn to avoid aposematic prey and generalize their warning signals to phenotypically similar mimics. The phenotypic accuracy needed for mimics to benefit from this adaptation depends on the relative densities of models and mimics and the models unpalatability. As aposematic models become more unpalatable or more common relative to their mimics, warning signals become stronger, allowing even poor mimics to benefit. However, few studies have disentangled the importance of relative frequencies of models and mimics from absolute density of the prey community (both models and mimics) in driving relaxed selection on imperfect mimics. Here, we test the hypothesis that increasing model unpalatability and absolute prey community density accelerates predator avoidance learning and enhances protection for imperfect mimics. Using replicas of the model Adelpha iphiclus (Linnaeus), its imperfect mimic Adelpha serpa (Boisduval), and the palatable control Junonia evarete (Cramer), we conducted field experiments that enhanced model unpalatability and doubled absolute prey density while maintaining a constant ratio of model, mimic, and control phenotypes. We found that enhanced model unpalatability and increased absolute density significantly reduced predation on all species, highlighting absolute community density as an underappreciated mechanism shaping selection on imperfect Batesian mimics.

ecology↗

bioRxiv2023

Giant genes are rare but implicated in cell wall degradation by predatory bacteria

Across the tree of life, gene lengths vary, but most are no more than a few thousand base pairs in length. The largest protein often reported is the [~]40,000 aa eukaryotic Titin. Even larger proteins may occur in the rapidly expanding set of metagenome-derived sequences, but their existence may be obscured by assembly fragmentation. Here, we leverage genome curation to complete metagenome-derived sequences that encode predicted proteins of up to 85,804 aa. Overall, the findings illuminate a huge knowledge gap related to giant proteins. Although predicted proteins of >30,000 aa occur in bacterial phyla such as Firmicutes and Actinobacteria, they are most common in ca. Omnitrophota, ultra small bacteria that adopt predatory lifestyles. All full length giant genes encode numerous transmembrane regions and most encode divergent secA DEAD helicase domains. In silico structural prediction of protein subregions was required to identify domains in unannotated protein segments, and revealed putative domains implicated in attachment and carbohydrate degradation. Many giant genes in new complete and near-complete Omnitrophota genomes occur in close proximity to genes homologous to type II secretion systems as well as carbohydrate import systems. This, in combination with the domain content, suggests that many bacterial giant proteins enable prey adhesion and cell wall digestion during bacterial predation.

microbiology↗

bioRxiv2023

Searching for Structure: Characterizing the Protein Conformational Landscape with Clustering-based Algorithms

The identification and characterization of the main conformations from a protein population is a challenging, inherently high-dimensional problem. We introduce the Secondary sTructural Ensembles with machine LeArning (StELa) double clustering method, which clusters protein structures based on the underlying Ramachandran plot. Our approach takes advantage of the relationship between the phi and psi dihedral angles in a protein backbone and the secondary structure of the protein. The classification of states as vectors composed of the clusters indices arising naturally from the Ramachandran plot, followed by the hierarchical clustering of the vectors, enables the identification of the minima from the corresponding free energy landscape (FEL) by lifting the high structure degeneracy found with existing approaches such as the RMSD-based clustering GROMOS. We compare the performance of StELa with not only GROMOS but also with CATS, the combinatorial averaged transient structure clustering method based on distributions of the phi and psi dihedral angle coordinates. Using ensembles of conformations from molecular dynamics (MD) simulations of either intrinsically disordered proteins (IDPs) of various lengths (tau protein fragments) or from local structures from a globular protein, we show that StELa is the only clustering method that identifies nearly all the minima from the corresponding FELs. In contrast, GROMOS yields a large number of clusters that cover the entire FEL and CATS, even with an additional clustering step, is unable to sample well the FEL for long IDPs and for fragments from globular proteins as it misses important minima. TOC Graphic O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=111 SRC="FIGDIR/small/557631v1_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@634f90org.highwire.dtl.DTLVardef@1fca966org.highwire.dtl.DTLVardef@d59bd9org.highwire.dtl.DTLVardef@1eadb94_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2023

Distinctive in vitro phenotypes in iPSC-derived neurons from patients with gain- and loss-of-function SCN2A developmental and epileptic encephalopathy

SCN2A encodes NaV1.2, an excitatory neuron voltage-gated sodium channel and major monogenic cause of neurodevelopmental disorders, including developmental and epileptic encephalopathies (DEE) and autism. Clinical presentation and pharmocosensitivity vary with nature of SCN2A variant dysfunction with gain-of-function (GoF) cases presenting with pre- or peri-natal seizures and loss-of-function (LoF) patients typically having infantile spasms after 6 months of age. Here, we established and assessed patient induced pluripotent stem cell (iPSC) - derived neuronal models for two recurrent SCN2A DEE variants with GoF R1882Q and LoF R853Q associated with early- and late-onset DEE, respectively. Patient-derived iPSC lines were differentiated using a Neurogenin-2 overexpression yielding populations of cortical-like glutamatergic neurons. Electrophysiological and transcriptomic profiles were assessed after 2-4 weeks in culture. Increased neuronal activity at both cellular and network level was observed for R1882Q iPSC-derived neurons at three weeks of differentiation. In contrast, R853Q neurons showed only subtle changes in excitability after four weeks in vitro. In alignment with the reported efficacy in some GoF SCN2A patients, phenytoin (sodium channel blocker) reduced excitability of neurons to the control levels in R1882Q neuronal cultures. Transcriptomic alterations in neurons were detected for each variant and convergent pathways pointed at the shared mechanisms underlying SCN2A DEE.

neuroscience↗

bioRxiv2022

Microtubule severing enzymes oligomerization and allostery: a tale of two domains

Severing proteins are nanomachines from the AAA+ (ATPases associated with various cellular activities) superfamily whose function is to remodel the largest cellular filaments, microtubules. The standard AAA+ machines adopt hexameric ring structures for functional reasons, while being primarily monomeric in the absence of the nucleotide. Both major severing proteins, katanin and spastin, are believed to follow this trend. However, studies proposed that they populate lower-order oligomers in the presence of co-factors, which are functionally relevant. Our simulations show that the preferred oligomeric assembly is dependent on the binding partners, and on the type of severing protein. Essential dynamics analysis predicts that the stability of an oligomer is dependent on the strength of the interface between the helical bundle domain (HBD) of a monomer and the convex face of the nucleotide binding domain (NBD) of a neighboring monomer. Hot spots analysis found that the region consisting of the HBD tip and the C-terminal (CT) helix is the only common element between the allosteric networks responding to nucleotide, substrate, and inter-monomer binding. Clustering analysis indicates the existence of multiple pathways for the transition between the secondary structure of the HBD tip in monomers and the structure(s) it adopts in oligomers.

biophysics↗

bioRxiv2022

Exploring the effect of mechanical anisotropy of protein structures in the unfoldase mechanism of AAA+ molecular machines

Essential cellular processes of microtubule disassembly and protein degradation, which span lengths from tens of m to nm, are mediated by specialized molecular machines with similar hexameric structure and function. Our molecular simulations at atomistic and coarse-grained scales show that both the microtubule severing protein spastin and the caseinolytic protease ClpY, accomplish spectacular unfolding of their diverse substrates, a microtubule lattice and dihydrofolate reductase (DHFR), by taking advantage of mechanical anisotropy in these proteins. By considering wild-type and variants of DHFR, we found that optimal ClpY-mediated action probes favorable orientations of the substrate relative to the machine. Unfolding of wild-type DHFR involves strong mechanical interfaces near each terminal and occurs along branched pathways, whereas unfolding of DHFR variants involves softer mechanical interfaces and occurs through single pathways, but translocation hindrance can arise from internal mechanical resistance. For spastin, optimum severing action initiated by pulling on a tubulin subunit is achieved through the orientation of the machine versus the substrate (microtubule lattice). Moreover, changes in the strength of the interactions between spastin and a microtubule filament, which can be driven by the tubulin code, lead to drastically different outcomes for the integrity of the hexameric structure of the machine.

biophysics↗

bioRxiv2021

Borgs are giant extrachromosomal elements with the potential to augment methane oxidation

Anaerobic methane oxidation exerts a key control on greenhouse gas emissions 1, yet factors that modulate the activity of microorganisms performing this function remain little explored. In studying groundwater, sediments, and wetland soil where methane production and oxidation occur, we discovered extraordinarily large, diverse DNA sequences that primarily encode hypothetical proteins. Four curated, complete genomes are linear, up to ~1 Mbp in length and share genome organization, including replicore structure, long inverted terminal repeats, and genome-wide unique perfect tandem direct repeats that are intergenic or generate amino acid repeats. We infer that these are a new type of archaeal extrachromosomal element with a distinct evolutionary origin. Gene sequence similarity, phylogeny, and local divergence of sequence composition indicate that many of their genes were assimilated from methane-oxidizing Methanoperedens archaea. We refer to these elements as "Borgs". We identified at least 19 different Borg types coexisting with Methanoperedens in four distinct ecosystems. Borg genes expand redox and respiratory capacity (e.g., clusters of multiheme cytochromes), ability to respond to changing environmental conditions, and likely augment Methanoperedens capacity for methane oxidation (e.g., methyl coenzyme M reductase). By this process, Borgs could play a previously unrecognized role in controlling greenhouse gas emissions.

microbiology↗

bioRxiv2019

A rapid method to determine the bactericidal activity of compounds against non-replicating Mycobacterium tuberculosis at low pH

There is an urgent need for new anti-tubercular agents which can lead to a shortened treatment time by targeting persistent or non-replicating bacilli. In order to assess compound activity against non-replicating Mycobacterium tuberculosis, we developed a method to detect the bactericidal activity of novel compounds within 7 days. Our method uses incubation at low pH in order to induce a non-replicating state. We used a strain of M. tuberculosis expressing luciferase; we first confirmed the linear relationship between luminescence and viable bacteria (determined by colony forming units) under our assay conditions. We optimized the assay parameters in 96-well plates in order to achieve a reproducible assay. Our final assay used M. tuberculosis in phosphate-citrate buffer, pH 4.5 exposed to compounds for 7 days; viable bacteria were determined by luminescence. We recorded the minimum bactericidal concentration at pH 4.5 (MBC4.5) representing >2 logs of kill. We confirmed the utility of the assay with control compounds. The ionophores monensin, niclosamide, and carbonyl cyanide 3-chlorophenylhydrazone and the anit-tubercular drugs pretomanid and rifampicin were active, while several other drugs such as isoniazid, ethambutol, and linezolid were not.

microbiology↗

bioRxiv2017

Cell-cycle dependent organization and dynamics of RNA Polymerase I in live human cells

RNA Polymerase I (Pol I) is responsible for over 60% of transcriptional output in human cells, yet basic questions concerning the spatial and temporal organization of the polymerase remain unanswered. Here we investigate how mammalian cells rely on Pol I organization throughout the cell cycle to balance different needs, from complete transcription shut down to massive increase in protein synthesis (and thus ribosomal RNA synthesis) before cell division. In contrast to our previous reports on RNA Polymerase II, Pol I clusters are stable with active transcription, and the presence of transient Pol I clusters correlates with inactive ribosomal transcription. Our results suggest that both stable and transient populations Pol I clusters co-exist in individual living cells, and their relative fraction may directly reflect the global gene expression need of the cell.

biophysics↗