Biology subjects

Prichard, A.

Publications and source records attributed to Prichard, A..

12 recordsLinked to original sources

bioRxiv2024

Histone H3 tail modifications required for meiosis in Saccharomyces cerevisiae

Histone tail phosphorylation has diverse effects on a myriad of cellular processes, including cell division, and is highly conserved throughout eukaryotes. Histone H3 phosphorylation at threonine 3 (H3T3) during mitosis occurs at the inner centromeres and is required for proper biorientation of chromosomes on the mitotic spindle. While H3T3 is also phosphorylated during meiosis, a possible role for this modification has not been tested. Here, we asked if H3T3 phosphorylation (H3T3ph) is important for meiotic division by quantifying sporulation efficiency and spore viability in Saccharomyces cerevisiae mutants with a T3A amino acid substitution. The T3A substitution resulted in greatly reduced sporulation efficiency and reduced spore viability. Analysis of two other H3 tail mutants, K4A and S10A, revealed different effects on sporulation efficiency and spore viability compared to the T3A mutant, suggesting that these phenotypes are due to failures in distinct functions. To determine if the spindle checkpoint promotes spore viability of the T3A mutant, the MAD2 gene required for the spindle assembly checkpoint was deleted to abolish spindle assembly checkpoint function. This resulted in a severe reduction in spore viability following meiosis. Altogether, the data reveal a critical function for histone H3 threonine 3 that requires monitoring by the spindle checkpoint to ensure successful completion of meiosis.

genetics↗

bioRxiv2024

Asesino: a nucleus-forming phage that lacks PhuZ

As nucleus-forming phages become better characterized, understanding their unifying similarities and unique differences will help us understand how they occupy varied niches and infect diverse hosts. All identified nucleus-forming phages fall within the proposed Chimalliviridae family and share a core genome of 68 unique genes including chimallin, the major nuclear shell protein. A well-studied but non-essential protein encoded by many nucleus-forming phages is PhuZ, a tubulin homolog which aids in capsid migration, nucleus rotation, and nucleus positioning. One clade that represents 24% of all currently known chimalliviruses lacks a PhuZ homolog. Here we show that Erwinia phage Asesino, one member of this PhuZ-less clade, shares a common overall replication mechanism with other characterized nucleus-forming phages despite lacking PhuZ. We show that Asesino replicates via a phage nucleus that encloses phage DNA and partitions proteins in the nuclear compartment and cytoplasm in a manner similar to previously characterized nucleus-forming phages. Consistent with a lack of PhuZ, however, we did not observe active positioning or rotation of the phage nucleus within infected cells. These data show that some nucleus-forming phages have evolved to replicate efficiently without PhuZ, providing an example of a unique variation in the nucleus-based replication pathway.

microbiology↗

bioRxiv2024

An essential and highly selective protein import pathway encoded by nucleus-forming phage

Targeting proteins to specific subcellular destinations is essential in prokaryotes, eukaryotes, and the viruses that infect them. Chimalliviridae phages encapsulate their genomes in a nucleus-like replication compartment composed of the protein chimallin (ChmA) that excludes ribosomes and decouples transcription from translation. These phages selectively partition proteins between the phage nucleus and the bacterial cytoplasm. Currently, the genes and signals that govern selective protein import into the phage nucleus are unknown. Here we identify two components of this novel protein import pathway: a species-specific surface-exposed region of a phage intranuclear protein required for nuclear entry and a conserved protein, PicA, that facilitates cargo protein trafficking across the phage nuclear shell. We also identify a defective cargo protein that is targeted to PicA on the nuclear periphery but fails to enter the nucleus, providing insight into the mechanism of nuclear protein trafficking. Using CRISPRi-ART protein expression knockdown of PicA, we show that PicA is essential early in the chimallivirus replication cycle. Together our results allow us to propose a multistep model for the Protein Import Chimallivirus (PIC) pathway, where proteins are targeted to PicA by amino acids on their surface, and then licensed by PicA for nuclear entry. The divergence in the selectivity of this pathway between closely-related chimalliviruses implicates its role as a key player in the evolutionary arms race between competing phages and their hosts. Significance StatementThe phage nucleus is an enclosed replication compartment built by Chimalliviridae phages that, similar to the eukaryotic nucleus, separates transcription from translation and selectively imports certain proteins. This allows the phage to concentrate proteins required for DNA replication and transcription while excluding DNA-targeting host defense proteins. However, the mechanism of selective trafficking into the phage nucleus is currently unknown. Here we determine the region of a phage nuclear protein that targets it for nuclear import and identify a conserved, essential nuclear shell-associated protein that plays a key role in this process. This work provides the first mechanistic model of selective import into the phage nucleus.

microbiology↗

bioRxiv2023

A mobile intron facilitates interference competition between co-infecting viruses

Mobile introns containing homing endonucleases are widespread in nature and have long been assumed to be selfish elements that provide no benefit to the host organism. These genetic elements are common in viruses, but whether they confer a selective advantage is unclear. Here we studied a mobile intron in bacteriophage {Phi}PA3 and found its homing endonuclease gp210 contributes to viral competition by interfering with the virogenesis of co-infecting phage {Phi}KZ. We show that gp210 targets a specific sequence in its competitor {Phi}KZ, preventing the assembly of progeny viruses. This work reports the first demonstration of how a mobile intron can be deployed to engage in interference competition and provide a reproductive advantage. Given the ubiquity of introns, this selective advantage likely has widespread evolutionary implications in nature.

microbiology↗

bioRxiv2023

Brain rhythms control microglial response and cytokine expression via NFkappaB signaling

Microglia, the brains primary immune cells, transform in response to changes in sensory or neural activity, like sensory deprivation. However, little is known about how specific frequencies of neural activity, or brain rhythms, impact microglia and cytokine signaling. Using visual noninvasive flickering sensory stimulation (flicker) to induce electrical neural activity at different frequencies, 40Hz, within the gamma band and 20Hz, within the beta band, we discovered these brain rhythms differentially affect microglial morphology and cytokine expression in healthy animals. We found that flicker induced expression of certain cytokines, including IL-10 and M-CSF, that was independent of microglia. Because NF{kappa}B is activated by synaptic activity and regulates cytokines, we hypothesized this pathway plays a causal role in frequency-specific cytokine and microglial responses. Indeed, we found that after flicker, phospho-NF{kappa}B co-labeled with neurons more than microglia. Furthermore, inhibition of NF{kappa}B signaling by a small molecule inhibitor down-regulated flicker-induced cytokine expression and attenuated flicker-induced changes in microglia morphology. These results reveal a new mechanism through which brain rhythms affect brain function by altering microglia morphology and cytokines via NF{kappa}B. TeaserFrequency-specific brain rhythms regulate cytokine expression, microglia morphology, and microglia-independent expression of M-CSF and IL10 via NF{kappa}B.

neuroscience↗

bioRxiv2023

Identifying the core genome of the nucleus-forming bacteriophage family and characterization of Erwinia phage RAY

We recently discovered that some bacteriophages establish a nucleus-like replication compartment (phage nucleus), but the core genes that define nucleus-based phage replication and their phylogenetic distribution were unknown. By studying phages that encode the major phage nucleus protein chimallin, including previously sequenced yet uncharacterized phages, we discovered that chimallin-encoding phages share a set of 72 highly conserved genes encoded within seven distinct gene blocks. Of these, 21 core genes are unique to this group, and all but one of these unique genes encode proteins of unknown function. We propose that phages with this core genome comprise a novel viral family we term Chimalliviridae. Fluorescence microscopy and cryo-electron tomography studies of Erwinia phage vB_EamM_RAY confirm that many of the key steps of nucleus-based replication encoded in the core genome are conserved among diverse chimalliviruses, and reveal that non-core components can confer intriguing variations on this replication mechanism. For instance, unlike previously studied nucleus-forming phages, RAY doesnt degrade the host genome, and its PhuZ homolog appears to form a five-stranded filament with a lumen. This work expands our understanding of phage nucleus and PhuZ spindle diversity and function, providing a roadmap for identifying key mechanisms underlying nucleus-based phage replication.

microbiology↗

bioRxiv2020

Isolation and characterization of Streptomyces bacteriophages and Streptomyces strains encoding biosynthetic arsenals

The threat to public health posed by drug-resistant bacteria is rapidly increasing, as some of healthcares most potent antibiotics are becoming obsolete. Approximately two-thirds of the worlds antibiotics are derived from natural products produced by Streptomyces encoded biosynthetic gene clusters. Thus, in order to identify novel gene clusters, we sequenced the genomes of four bioactive Streptomyces strains isolated from the soil in San Diego County and used Bacterial Cytological Profiling adapted for agar plate culturing in order to examine the mechanisms of bacterial inhibition exhibited by these strains. In the four strains, we identified 101 biosynthetic gene clusters. Some of these clusters were predicted to produce previously studied antibiotics; however, the known mechanisms of these molecules could not fully account for the antibacterial activity exhibited by the strains, suggesting that novel clusters might encode antibiotics. When assessed for their ability to inhibit the growth of clinically isolated pathogens, three Streptomyces strains demonstrated activity against methicillin-resistant Staphylococcus aureus. Additionally, due to the utility of bacteriophages for genetically manipulating bacterial strains via transduction, we also isolated four new phages (BartholomewSD, IceWarrior, Shawty, and TrvxScott) against S. platensis. A genomic analysis of our phages revealed nearly 200 uncharacterized proteins, including a new site-specific serine integrase that could prove to be a useful genetic tool. Sequence analysis of the Streptomyces strains identified CRISPR-Cas systems and specific spacer sequences that allowed us to predict phage host ranges. Ultimately, this study identified Streptomyces strains with the potential to produce novel chemical matter as well as integrase-encoding phages that could potentially be used to manipulate these strains.

microbiology↗

bioRxiv2020

2D or Not 2D? An FMRI Study of How Dogs Visually Process Objects

ABSTRACTGiven humans’ habitual use of screens, they rarely consider potential differences when viewing two dimensional (2D) stimuli and real-world versions of dimensional stimuli. Dogs also have access to many forms of screens and touch pads, with owners even subscribing to dog-directed content. Humans understand that 2D stimuli are representations of real-world objects, but do dogs? In canine cognition studies, 2D stimuli are almost always used to study what is normally 3D, like faces, and may assume that both 2D and 3D stimuli are represented in the brain the same way. Here, we used awake fMRI of 15 dogs to examine the neural mechanisms underlying dogs’ perception of two- and three-dimensional objects after the dogs were trained on either a two- or three-dimensional version of the objects. Activation within reward processing regions and parietal cortex of the dog brain to 2D and 3D versions of objects was determined by their training experience, as dogs trained on one dimensionality showed greater activation to the dimension on which they were trained. These results show that dogs do not automatically generalize between two- and three-dimensional stimuli and caution against implicit assumptions when using pictures or videos with dogs.Competing Interest StatementG.B. & M.S. own equity in Dog Star Technologies and developed technology used in some of the research described in this paper. The terms of this arrangement have been reviewed and approved by Emory University in accordance with its conflict of interest policies.View Full Text

animal behavior and cognition↗

bioRxiv2019

Canine sense of quantity: evidence for numerical ratio-dependent activation in parietotemporal cortex

The approximate number system, which supports the rapid estimation of quantity, emerges early in human development and is widespread across species. Neural evidence from both human and non-human primates suggests the parietal cortex as a primary locus of numerical estimation, but it is unclear whether the numerical competencies observed across non-primate species are subserved by similar neural mechanisms. Moreover, because studies with non-human animals typically involve extensive training, little is known about the spontaneous numerical capacities of non-human animals. To address these questions, we examined the neural underpinnings of number perception using awake canine functional magnetic resonance imaging. Dogs passively viewed dot arrays that varied in ratio and, critically, received no task-relevant training or exposure prior to testing. We found evidence of ratio-dependent activation, which is a key feature of the approximate number system, in canine parietotemporal cortex in the majority of dogs tested. This finding is suggestive of a neural mechanism for quantity perception that has been conserved across mammalian evolution.

neuroscience↗

bioRxiv2019

Decoding Odor Mixtures in the Dog Brain: An Awake fMRI Study

In working and practical contexts, dogs rely upon their ability to discriminate a target odor from distracting odors and other sensory stimuli. Few studies have examined odor discrimination using non-behavioral methods or have approached odor discrimination from the dogs perspective. Using awake fMRI in 18 dogs, we examined the neural mechanisms underlying odor discrimination between two odors and a mixture of the odors. Neural activation was measured during the presentation of a target odor (A) associated with a food reward, a distractor odor (B) associated with nothing, and a mixture of the two odors (A+B). Changes in neural activation during the presentations of the odor stimuli in individual dogs were measured over time within three regions known to be involved with odor processing: the caudate nucleus, the amygdala, and the olfactory bulbs. Average activation within the amygdala showed that dogs maximally differentiated between odor stimuli based on the stimulus-reward associations by the first run, while activation to the mixture (A+B) was most similar to the no-reward (B) stimulus. To identify the neural representation of odor mixtures in the dog brain, we used a random forest classifier to compare multilabel (elemental) vs. multiclass (configural) models. The multiclass model performed much better than the multilabel (weighted-F1 0.44 vs. 0.14), suggesting the odor mixture was processed configurally. Analysis of the subset of high-performing dogs based on their brain classification metrics revealed a network of olfactory information-carrying brain regions that included the amygdala, piriform cortex, and posterior cingulate. These results add further evidence for the configural processing of odor mixtures in dogs and suggest a novel way to identify high-performers based on brain classification metrics.

neuroscience↗

bioRxiv2017

Awake fMRI Reveals Covert Arousal in Aggressive Dogs Under Social Resource Threat

Domestic dogs are highly social, and have been shown sensitive not only to the actions of humans and other dogs but to the interactions between them. To examine the canine neurobiological response to observed interactions between a human and another dog, we collected fMRI data from dogs while they watched their owner feed a realistic fake dog or deposit food in a bucket. Given the likelyihood that arousal and affective state may contribute to responses to observed social situations, we examined the relationship between amygdala activation in these two conditions and an independent measure of aggressive temperament from the C-BARQ scale. Dogs rated more aggressive showed significantly higher activation in the fake-dog versus bucket condition. This finding suggests a neurobiological mechanism mediated by the amygdala for dog-directed aggression, especially when their owner interacts with another dog. Such a mechanism may have some parallels to human jealousy. Further, it adds to a growing body of evidence that specific neurobiological responses correlate with canine temperament and can be a predictor of future behavior. We also found evidence that the amygdala response habituates with repeated observed interactions. This suggests value in exposure-based interventions for potentially aggressive dogs.

neuroscience↗

bioRxiv2017

Awake fMRI Reveals Mechanisms of Language Comprehension in Dogs

How do dogs understand human words? At a basic level, understanding would require the discrimination of words from non-words. To determine the mechanisms of such a discrimination, we trained 12 dogs to retrieve two objects based on object names, then probed the neural basis for these auditory discriminations using awake-fMRI. We compared the neural response to these trained words relative to \"oddball\" pseudowords the dogs had not heard before. Consistent with novelty detection, we found greater activation for pseudowords relative to trained words bilaterally in the parietotemporal cortex. To probe the neural basis for representations of trained words, searchlight multivoxel pattern analysis (MVPA) revealed that a subset of dogs had clusters of informative voxels that discriminated between the two trained words. These clusters included the left temporal cortex and amygdala, left caudate nucleus, and thalamus. These results demonstrate that dogs processing of human words utilizes basic processes like novelty detection, and for some dogs, may also include auditory and hedonic representations.

neuroscience↗