Biology subjects

Holt, L.

Publications and source records attributed to Holt, L..

12 recordsLinked to original sources

bioRxiv2026

Quiescence improves Candida albicans survival of fungicidal drug exposure

Quiescence, defined as the reversible exit from mitotic division and proliferative growth, is the predominant state of all microbes. Despite its prevalence, the properties and consequences of quiescence in Candida albicans, an opportunistic fungal pathogen, remain largely unexplored. In this study, we characterized the morphological, molecular, and biophysical properties of quiescent C. albicans cells and assessed the effects of quiescence on antifungal drug efficacy. Quiescent cells that were induced via carbon starvation in rich and minimal media underwent distinct morphological changes upon entry into quiescence; this included an increase in cell buoyant density, altered fluidity of the cytoplasm and nucleus, and remodeling of mitochondria. Most C. albicans cells arrested in an unbudded G1/G0 state, although a significant fraction of cells had budded morphologies and 4N DNA content, indicating that they arrested at other cell cycle phases. Both budded and unbudded quiescent cells efficiently re-entered the cell cycle upon nutrient replenishment, with time-to-quiescence exit varying depending on the total nutritional quality of the medium. Quiescence was associated with large-scale gene expression remodeling, including downregulation of ribosomal biogenesis genes and upregulation of autophagy and stress response pathways. Notably, a greater proportion of quiescent cells than proliferative cells survived exposure to the commonly used antifungal drugs micafungin, caspofungin, and amphotericin B in genetically diverse strains. Thus, quiescence is a distinct cellular state with important implications for antifungal drug efficacy in C. albicans. Author SummaryWe show that Candida albicans, a common fungal pathogen, can enter a reversible, non-dividing state when starved of carbon. Starved cells become smaller and denser, reorganize their mitochondria, change how densely packed the inside of the cell and its nucleus are, and switch on stress-protection and internal recycling programs while reducing protein synthesis activity. Most cells have ceased to actively divide, but many retained budded shapes and could restart growth when nutrients returned; the timing of recovery depended on the nutritional environment in which quiescence was initiated. Critically, quiescent cells from laboratory and clinical strains exhibited greater survival than proliferative cells when exposed to widely used fungicidal drugs including micafungin, caspofungin, and amphotericin B. These findings indicate that quiescence is an active, adaptive physiological state that helps Candida albicans survive hostile environmental conditions such as temperature stress and drug exposure. Accounting for the metabolic state of fungal cells in diagnostics and drug development may improve treatment outcomes.

microbiology↗

bioRxiv2026

Cortical astrocytes control stress resilience

BackgroundChronic stress exposure is a risk factor for several psychiatric disorders, including post-traumatic stress disorder (PTSD) and major depression (MDD), with the prefrontal cortex (PFC) playing a key role in mediating this stress susceptibility. However, most individuals who are exposed to chronic stress are resilient and do not develop psychopathology. Recent evidence suggests that glial cells, especially astrocytes, play an important role in controlling stress-induced anxiety- and depression-like behavior, yet their role in contributing to stress resilience is not understood. MethodsUsing fiber photometry, chemogenetics, and RNA-sequencing in male mice, we establish a role for PFC astrocytes in stress resilience. ResultsWe demonstrate that stress-induced increases in astrocytic calcium activity are both necessary and sufficient for resilience. Bioinformatic analysis reveals robust transcriptional responses in PFC astrocytes that differ between susceptible vs. resilient mice and are unique when compared to astrocytic transcriptional changes in other limbic regions. Comparison with human RNA-sequencing data indicates that molecular changes observed in PFC astrocytes from susceptible mice converge with gene expression changes observed in MDD patients. ConclusionsTogether, these data support targeting astrocytes as a potential therapy for negative behavioral consequences following stress exposure and reveal potential molecular mechanisms within PFC astrocytes that could contribute to depressive-like behaviors.

neuroscience↗

bioRxiv2025

An α-1,3-fucosyltransferase influences thermal nocifensive escape behaviors in Drosophila larvae

When exposed to noxious thermal or mechanical stimuli, Drosophila melanogaster larvae will aZempt to escape by performing a stereotypical nocifensive rolling behavior. Here, we report the identification of a mutation in Drosophila that specifically alters the thermal nocifensive escape behaviors of larvae. We provide genetic, molecular, and histological evidence that this mutation maps to the FucTA gene, which encodes an -1,3-fucosyltransferase. Our results suggest that fucosylation is important for the thermal nociception rolling behavior of Drosophila larvae.

genetics↗

bioRxiv2024

Sex-Specific Regulation of Stress Susceptibility by the Astrocytic Gene Htra1

Major depressive disorder (MDD) is linked to impaired structural and synaptic plasticity in limbic brain regions. Astrocytes, which regulate synapses and are influenced by chronic stress, likely contribute to these changes. We analyzed astrocyte gene profiles in the nucleus accumbens (NAc) of humans with MDD and mice exposed to chronic stress. Htra1, which encodes an astrocyte-secreted protease targeting the extracellular matrix (ECM), was significantly downregulated in the NAc of males but upregulated in females in both species. Manipulating Htra1 in mouse NAc astrocytes bidirectionally controlled stress susceptibility in a sex-specific manner. Such Htra1manipulations also altered neuronal signaling and ECM structural integrity in NAc. These findings highlight astroglia and the brains ECM as key mediators of sex-specific stress vulnerability, offering new approaches for MDD therapies.

neuroscience↗

bioRxiv2024

From small to tall: breed-varied household pet dogs can be trained to detect Parkinson's Disease

ObjectiveParkinsons Disease (PD) is a clinically diagnosed disease that carries a reported misdiagnosis rate of 10-20%. Recent scientific discoveries have provided evidence of volatile organic compounds in sebum that are unique to patients with PD. The purpose of this study was to determine if companion dogs could be trained to distinguish between sebum samples provided by PD-positive patients and PD-negative human controls. MethodsThis was a randomized, handler-blind, controlled study. Twenty-three canines of varying breeds, ages, and environmental backgrounds were included. The two-year study period reported here was the final two years of a seven-year program which started in January of 2016. This study encompassed 200 total working session days from 2021 and 2022. ResultsWhen averaged as a group over two years, the 23 dogs were 89% sensitive and 87% specific to an olfactory distinction between PD-positive and PD-negative human donor samples. Ten of the twenty-three dogs averaged 90% or higher in both sensitivity and specificity. In 161 separate instances, dogs were presented with both unique PD-positive and PD-negative samples (the dogs had not previously encountered any of the samples presented). For these first-time exposures, the 23 dogs collectively averaged 86% sensitivity and 89% specificity. When comparing the sensitivity and specificity of PD-positive samples from donors who reported levodopa usage to PD-positive samples from donors who reported no levodopa usage, the dogs showed no statistical difference in sensitivity or specificity at the 95% significance level, indicating levodopa usage is not a factor in PD canine detection. Other factors investigated as part of this study included sample donor gender, canine breed, age, duration of time in the program, and training. ConclusionsCompanion dogs can be trained with reward-biased detection methodologies to distinguish between PD-positive and PD-negative donor sebum samples in a controlled setting. This study provides further evidence of one or more volatile organic compounds in the sebum of PD-positive patients that can be detected by canines. Summarily, study findings support the application of trained companion dogs for the screening of PD-positive and PD-negative samples in which numbers of samples are limited and the dogs are worked in short intervals, followed by recovery training.

animal behavior and cognition↗

bioRxiv2021

Control of nuclear size by osmotic forces in Schizosaccharomyces pombe

The size of the nucleus scales robustly with cell size so that the nuclear-to-cell volume ratio (N/C ratio) is maintained during cell growth in many cell types. The mechanism responsible for this scaling remains mysterious. Previous studies have established that the N/C ratio is not determined by DNA amount but is instead influenced by factors such as nuclear envelope mechanics and nuclear transport. Here, we developed a quantitative model for nuclear size control based upon colloid osmotic pressure and tested key predictions in the fission yeast Schizosaccharomyces pombe. This model posits that the N/C ratio is determined by the numbers of macromolecules in the nucleoplasm and cytoplasm. Osmotic shift experiments showed that the fission yeast nucleus behaves as an ideal osmometer whose volume is primarily dictated by osmotic forces. Inhibition of nuclear export caused accumulation of macromolecules and an increase in crowding in the nucleoplasm, leading to nuclear swelling. We further demonstrated that the N/C ratio is maintained by a homeostasis mechanism based upon synthesis of macromolecules during growth. These studies demonstrate the functions of colloid osmotic pressure in intracellular organization and size control.

biophysics↗

bioRxiv2021

Categorical encoding of voice in human superior temporal cortex

The ability to recognize abstract features of voice during auditory perception is a complex, yet poorly understood, feat of human audition. For the listener, this occurs in near-automatic fasion to seamlessly extract complex cues from a highly variable auditory signal. Voice perception depends on specialized regions of auditory cortex, including superior temporal gyrus (STG) and superior temporal sulcus (STS). However, the nature of voice encoding at the cortical level remains poorly understoood. We leverage intracerebral recordings across human auditory cortex during presentation of voice and non-voice acoustic stimuli to examine voice encoding in auditory cortex, in eight patient-participants undergoing epilepsy surgery evaluation. We show that voice-selectivity increases along the auditory hierarchy from supratemporal plane (STP) to the STG and STS. Results show accurate decoding of vocalizations from human auditory cortical activity even in the complete absence of linguistic content. These findings show an early, less-selective temporal window of neural activity in the STG and STS followed by a sustained, strongly voice-selective window. We then developed encoding models that demonstrate divergence in the encoding of acoustic features along the auditory hierarchy, wherein STG/STS responses were best explained by voice category as opposed to the acoustic features of voice stimuli. This is in contrast to neural activity recorded from STP, in which responses were accounted for by acoustic features. These findings support a model of voice perception that engages categorical encoding mechanisms within STG and STS. Significance StatementVoice perception occurs via specialized networks in higher order auditory cortex, yet how voice features are encoded remains a central unanswered question. With human intracerebral recordings of auditory cortex, we provide evidence for categorical encoding of voice in STG and STS and occurs in the absence of linguistic content. This selectivity strengthens after an initial onset response and cannot be explained by simple acoustic features. Together, these data support the existence of sites within STG and STS that are specialized for voice perception.

neuroscience↗

bioRxiv2021

Prosodic categories in speech are acoustically multidimensional: evidence from dimension-based statistical learning

Segmental speech units such as phonemes are cued by multiple acoustic dimensions (e.g. F0 and duration), but dimensions do not carry equal perceptual weight. The relative perceptual weights of acoustic speech dimensions are not fixed but vary with context. For example, when speech is altered to create an accent in which two acoustic dimensions are correlated in a manner opposite that of long-term experience, the dimension that carries less perceptual weight is down-weighted to contribute less in category decisions. It remains unclear, however, whether this short-term reweighting is limited to segmental categorization, or if it extends to categorization of suprasegmental features which span multiple phonemes, syllables, or words, which would suggest that such "dimension-based statistical learning" is a widespread phenomenon in speech perception. Here we investigated the relative contribution of two acoustic dimensions to word emphasis. Participants categorized instances of a two-word phrase pronounced with typical covariation of fundamental frequency (F0) and duration, and in the context of an artificial accent in which F0 and duration (established in prior research on English speech as primary and secondary dimensions, respectively) covaried atypically. When categorizing accented speech, listeners rapidly down-weighted the secondary dimension (duration) while continuing to rely on the primary dimension (F0). This result indicates that listeners continually track short-term regularities across speech input and dynamically adjust the weight of acoustic evidence for suprasegmental categories. Thus, dimension-based statistical learning appears to be a widespread phenomenon in speech perception extending to both segmental and suprasegmental categorization.

neuroscience↗

bioRxiv2018

Synthetic-evolution reveals that phosphoregulation of the mitotic kinesin-5 Cin8 is constrained

Cdk1 has been found to phosphorylate the majority of its substrates in disordered regions. These phosphorylation sites do not appear to require precise positioning for their function. The mitotic kinesin-5 Cin8 was shown to be phosphoregulated at three Cdk1 sites in disordered loops within its catalytic motor domain. Here, we examined the flexibility of Cin8 phosphoregulation by analyzing the phenotypes of synthetic Cdk1-sites that were systematically generated by single amino-acid substitutions, starting from a phosphodeficient variant of Cin8. Out of 29 synthetic Cdk1 sites that we created, eight were non-functional; 19 were neutral, similar to the phosphodeficient variant; and two gave rise to phosphorylation-dependent spindle phenotypes. Of these two, one site resulted in novel phosphoregulation, and only one site, immediately adjacent to a native Cdk1 site, produced phosphoregulation similar to wild-type. This study shows that, while the gain of a single phosphorylation site can confer regulation and modulate the dynamics of the spindle, to achieve optimal regulation of a mitotic kinesin-5 motor protein, phosphoregulation has to be site-specific and precise.

cell biology↗

bioRxiv2017

The SMuSh pathway is essential for survival during growth-induced compressive mechanical stress

Cells that proliferate within a confined environment build up mechanical compressive stress. For example, mechanical pressure emerges in the naturally space-limited tumor environment. However, little is known about how cells sense and respond to mechanical compression. We developed microfluidic bioreactors to enable the investigation of the effects of compressive stress on the growth of the genetically tractable model organism Saccharomyces cerevisiae. We used this system to determine that compressive stress is partly partly sensed through a module consisting of the mucin Msb2, and the cell wall protein Sho1, which act together as a sensor module in one of the two major osmosensing pathways in budding yeast. This signal is transmitted via the MAPKKK kinase Ste11. Thus, we term this mechanosensitive pathway the SMuSh pathway, for Ste11 through Mucin / Sho1 pathway. The SMuSh pathway delays cells in the G1 phase of the cell cycle and improves cell survival in response to growth-induced pressure. We also found that the Cell Wall Integrity (CWI) pathway contributes to the response to mechanical compressive stress. These latter results are confirmed in complimentary experiments in the accompanying manuscript from Mishra et al. When both the SMuSh and the CWI pathways are deleted, cells fail to adapt to compressive stress and all cells lyse at relatively low pressure when grown in confinement. Thus, we define a network that is essential for cell survival during growth under pressure. We term this new mechanosensory system the SCWISh (Survival through the CWI and SMuSh) network.\n\nSignificance StatementGrowth in confined environments leads to the build up of compressive mechanical stresses, which are relevant to diverse fields, from cancer to microbiology. In contrast to tensile stress, little is known about the molecular integration of compressive stresses. In this study, we elucidate the SMuSh pathway, which, together with the Cell Wall Integrity pathway, is essential for viability of the budding yeast S. cerevisiae when growing under mechanical pressure. Pressure-sensing requires the transmembrane mucin, Msb2, which is linked to the actin cortex. Our result raises the intriguing question of whether mucins, widely conserved in eukaryotes and frequently misregulated in cancers, might sense compressive stresses in other organisms, including humans.

cell biology↗

bioRxiv2017

Extensive Tonotopic Mapping Across Auditory Cortex Is Recapitulated By Spectrally-Directed Attention, And Systematically Related To Cortical Myeloarchitecture

Auditory selective attention is vital in natural soundscapes. But, it is unclear how attentional focus on the primary dimension of auditory representation - acoustic frequency - might modulate basic auditory functional topography during active listening. In contrast to visual selective attention, which is supported by motor-mediated optimization of input across saccades and pupil dilation, the primate auditory system has fewer means of differentially sampling the world. This makes spectrally-directed endogenous attention a particularly crucial aspect of auditory attention. Using a novel functional paradigm combined with quantitative MRI, we establish that human frequency-band-selective attention drives activation in both myeloarchitectonically-estimated auditory core, and across the majority of tonotopically-mapped non-primary auditory cortex. The attentionally-driven best-frequency maps show strong concordance with sensory-driven maps in the same subjects across much of the temporal plane, with poor concordance in non-auditory areas. There is significantly greater activation across most of auditory cortex when best frequency is attended, versus ignored. Moreover, the single frequency bands that evoke the least activation and the frequency bands that elicit the least activation when attention is directed to them also correspond closely. Finally, the results demonstrate that there is spatial correspondence between the degree of myelination and the strength of the tonotopic signal across a number of regions in auditory cortex. Strong frequency preferences across tonotopically-mapped auditory cortex spatially correlate with R1-estimated myeloarchitecture, indicating shared functional and anatomical organization that may underlie intrinsic auditory regionalization.\n\nSignificancePerception is an active process especially sensitive to attentional state. Listeners direct auditory attention to track a violins melody within an ensemble performance, or to follow a voice in a crowded cafe. Although diverse pathologies reduce quality of life by impacting such spectrally-directed auditory attention, its neurobiological bases are unclear. We demonstrate that human primary and non-primary auditory cortical activation is modulated by spectrally-directed attention in a manner that recapitulates its tonotopic sensory organization. Further, the graded activation profiles evoked by single frequency bands are correlated with attentionally-driven activation when these bands are presented in complex soundscapes. Finally, we observe a strong concordance in the degree of cortical myelination and the strength of tonotopic activation across several auditory cortical regions.

neuroscience↗

bioRxiv2017

Structural and functional dissection of the interplay between lipid and Notch binding by human Notch ligands

AbstractRecent data have expanded our understanding of Notch signaling by identifying a C2 domain at the N-terminus of Notch ligands which has both lipid- and receptor-binding properties. We present novel structures of human ligands Jagged2 and DLL4 and human Notch-2, together with functional assays, which suggest that ligand-mediated coupling of membrane recognition and Notch binding is likely to be critical in establishing the optimal context for Notch signaling. Comparisons between the Jagged and Delta family show a huge diversity in the structures of the loops at the apex of the C2 domain implicated in membrane recognition and Jagged1 missense mutations which affect these loops and are associated with extrahepatic biliary atresia lead to a loss of membrane recognition, but do not alter Notch binding. Taken together, these data suggest that C2 domain binding to membranes is an important element in tuning ligand-dependent Notch signaling in different physiological contexts.

biochemistry↗