Biology subjects

Chen, K.-F.

Publications and source records attributed to Chen, K.-F..

11 recordsLinked to original sources

bioRxiv2026

Drosophila orthologues of oculocutaneous albinism-associated genes regulate sleep and circadian rhythm via visual neurotransmission

Melanin is a pigment found in the skin and cuticle of animals. Oculocutaneous albinism (OCA) is a group of autosomal recessive disorders defined by reduced melanin in skin and eyes, and is associated with visual defects such as foveal hypoplasia and infantile nystagmus. Sleep disturbance has been documented in children with OCA, and oca2 loss-of-function in cavefish causes constitutive sleep loss, indicating a sleep regulatory function of OCA-associated genes in the visual system. To test potential roles of OCA-associated genes in regulating sleep and vision through evolutionarily conserved mechanisms, we used Drosophila melanogaster, a high-throughput phenotyping system to screen sleep and visual phenotypes for genetic mutants of OCA-associated genes. Among the OCA-associated genes, bidirectional DRSC Integrative Ortholog Prediction Tool identified Drosophila orthologues for OCA2, SLC45A2, SLC24A5 and LRMDA. RNAi-mediated knockdown in developing Drosophila eye tissue identified the OCA2 orthologues hoe1, hoe2, and the SLC45A2 orthologue lovit as candidate genes required for normal sleep. Moreover, hoe1, hoe2 and lovit1 null alleles reduced sleep and circadian rhythmicity, and showed altered photoreceptor neurotransmission. Consistently, the enhancer-trap reporter assay and scRNA dataset revealed that these OCA orthologues express in brain regions critical for sleep and vision regulation. Collectively the data indicate evolutionarily conserved neuronal function of OCA2 and SLC45A2 orthologues that regulate sleep and photoreceptor neurotransmission. Significance StatementOculocutaneous albinism (OCA) is a pigmentation disorder of the skin and eyes accompanied by reduced visual acuity. Children with albinism also report sleep disturbance, and the mechanism is unclear. We tested whether genes mutated in albinism regulate sleep in the fruit fly Drosophila melanogaster, an organism that has a divergent melanin synthesis pathway and lacks tyrosinase, the principal pigmentation enzyme in mammals. We found Drosophila OCA orthologues, hoe1, hoe2 and lovit express widely in the nervous system and control sleep, circadian rhythmicity and neural activity of photoreceptors. Importantly no pigmentation defects were detected in aforementioned mutants, our finding is consistent with the proposal that pleiotropic neuronal function of OCA2 and SLC45A2 underlies the sleep and vision symptoms in albinism.

neuroscience↗

bioRxiv2025

Nighttime sleep modulates mechanosensory habituation in Drosophila

Sleep is a highly conserved behaviour that facilitates episodic, procedural, and/or associative modes of memory across species 1-3, a process hypothesised to involve structural downscaling of synapses potentiated during waking episodes 4-6. Habituation, an ancient form of non-associative memory in which repetition of a stimulus leads to diminishing behavioural responses, is considered a prerequisite for such complex forms of learning 7,8. Yet whether sleep also influences the capacity of organisms to habituate is poorly understood. Here we examine this question using the fruit fly, Drosophila. We describe an automated system that yields long-term analogue measurements of habituation to mechanosensory stimuli in adult flies. Using this platform, we find that Drosophila lacking neuronal calcium sensor Neurocalcin - which exhibit reduced sleep specifically during the night 9 - display impaired mechanosensory habituation during the day. Mimicking a method used to treat insomnia 10, we show that compressing nighttime duration both restores consolidated night sleep and renormalises habituation in Neurocalcin mutants. Conversely, in wild type flies without sleep deficits, decreasing night length reduces total night sleep and disrupts daytime habituation. In wild type flies, exposure to longer nights - which increases overall night sleep - enhances synaptic downscaling in sensory circuits. Strikingly, this effect is abolished in Neurocalcin mutants, which do not exhibit increased sleep under these conditions. Collectively, our findings reveal a role for nighttime sleep in facilitating habituation and suggest that this may occur by promoting synaptic downscaling.

neuroscience↗

bioRxiv2025

A sleep promoting role of phototransduction in Drosophila melanogaster

The daily sleep-wake cycle is a conserved behaviour defined by locomotion quiescence and enhanced responsive threshold to sensory stimuli. Both the circadian clock and sleep-homeostasis determine the daily sleep profile. Environmental light is a major sensory input and also regulates circadian clock and the balance between sleep and wakefulness. In Drosophila, the cellular mechanism and neural circuitry underlying light-mediated circadian synchronization are well-established, yet the direct relationship between light/visual input and sleep remains unclear. To address this knowledge gap, we measured sleep behaviour in Drosophila with mutations in genes involved in phototransduction and downstream neural transmission. We observed consistent day sleep fragmentation in flies with mutations in multiple phototransduction components. We also found that mutation that led to hyperpolarised Drosophila photoreceptors resulted in shorter day sleep. We found a severe reduction in locomotor speed in several visual mutants during normal waking time preventing assessment of their sleep-linked immobility. Taken together, our rigorous quantification of sleep in phototransduction genetic mutants reveals the key role of visual input in promoting sleep.

neuroscience↗

bioRxiv2022

Antigenic characterization and pandemic risk assessment of North American H1 influenza A viruses circulating in swine

The first pandemic of the 21st century was caused by an H1N1 influenza A virus (IAV) introduced from pigs into humans, highlighting the importance of swine as reservoirs for pandemic viruses. Two major lineages of swine H1 circulate in North America: the 1A classical swine lineage (including the 2009 pandemic H1N1) and 1B human seasonal-like lineage. Here, we investigated the evolution of these H1 IAV lineages in North American swine and their potential pandemic risk. We assessed the antigenic distance between the HA of representative swine H1 and human seasonal vaccine strains (1978-2015) in hemagglutination inhibition (HI) assays using a panel of monovalent anti-sera raised in pigs. Antigenic cross-reactivity varied by strain but was associated with genetic distance. Generally, swine 1A lineage viruses that seeded the 2009 H1 pandemic were antigenically most similar to H1 pandemic vaccine strains, with the exception of viruses in the genetic clade 1A.1.1.3 that had a two-amino acid deletion mutation near the receptor-binding site, dramatically reducing antibody recognition. The swine 1B lineage strains, which arose from previously circulating (pre-2009 pandemic) human seasonal viruses, were more antigenically similar to pre-2009 human seasonal H1 vaccine viruses than post-2009 strains. Human population immunity was measured by cross-reactivity in HI assays to representative swine H1 strains. There was a broad range of titers against each swine strain that was not associated with age, sex, or location. However, there was almost no cross-reactivity in human sera to the 1A.1.1.3 and 1B.2.1 genetic clades of swine viruses, and the 1A.1.1.3 and 1B.2.1 clades were also the most antigenically distant from all human vaccine strains. Our data demonstrate that antigenic distances of representative swine strains from human vaccine strains represent a rational assessment of swine IAV for zoonotic risk research and pandemic preparedness prioritization. ImportanceHuman H1 influenza A viruses (IAV) spread to pigs in North America, resulting in sustained circulation of two major groups of H1 viruses in swine. We quantified the genetic diversity of H1 in swine and measured antigenic phenotypes. We demonstrated that swine H1 lineages were significantly different from human vaccine strains and this antigenic dissimilarity increased over time as the viruses evolved in swine. Pandemic preparedness vaccine strains for human vaccines also demonstrated a loss in similarity with contemporary swine strains. Human sera revealed a range of responses to swine IAV, including two groups of viruses with little to no immunity. Surveillance and risk assessment of IAV diversity in pig populations are essential to detect strains with reduced immunity in humans, providing critical information for pandemic preparedness.

microbiology↗

bioRxiv2022

Quantum CART (qCART), a piggyBac-based system for development and production of virus-free multiplex CAR-T cell therapy

Chimeric antigen receptor T (CAR-T) cell therapy has the potential to transform cancer treatment. However, CAR-T therapy application is currently limited to certain types of relapsed/refractory B cell lymphomas. To unlock the full potential of CAR-T therapy, technologic breakthroughs will be needed in multiple areas, including optimization of autologous CAR-T development, shortening the innovation cycle, and further manufacturing advancement of next-generation CAR-T therapies. Here, we established a simple and robust virus-free multiplex Quantum CART system that seamlessly and synergistically integrates four platforms: 1. GTailor for rapid identification of lead CAR construct design, 2. Quantum Nufect for effective but gentle electroporation-based gene delivery, 3. Quantum pBac, featuring a virus-free transposon-based vector with large payload capacity and integration profile similar to retrovirus, and 4. iCellar for robust and high-quality CAR+ T memory stem cell expansion. This robust, virus-free multiplex Quantum CART system is expected to unleash the full potential of CAR-T therapy for treating diseases.

cancer biology↗

bioRxiv2021

Nuclear Export of Drosophila PERIOD contributes to temperature compensation of the circadian clock

Circadian clocks are self-sustained molecular oscillators controlling daily changes of behavioral activity and physiology. For functional reliability and precision the frequency of these molecular oscillations must be stable at different environmental temperatures, known as temperature compensation. Despite being an intrinsic property of all circadian clocks, this phenomenon is not well understood at the molecular level. Here we use behavioral and molecular approaches to characterize a novel mutation in the period (per) clock gene of Drosophila melanogaster, which alters a predicted nuclear export sequence (NES) of the PER protein. We show that this new perI530A allele leads to progressively longer behavioral periods and clock oscillations with increasing temperature in both clock neurons and peripheral clock cells. While the mutant PERI530A protein shows normal circadian fluctuations and post-translational modifications at cool temperatures, increasing temperatures lead to both, severe amplitude dampening and hypophosphorylation of PERI530A. We further show that PERI530A displays reduced repressor activity at warmer temperatures, presumably because it cannot inactivate the transcription factor CLOCK (CLK). With increasing temperatures nuclear accumulation of PERI530A within clock neurons is increased, suggesting that PER is normally exported out of the nucleus at warm temperatures. Consequently, downregulating the nuclear export factor CRM1 also leads to temperature-dependent changes of behavioral rhythms. In summary, our results suggest that the PER NES and the nuclear export of clock proteins play an important role in temperature compensation of the Drosophila circadian clock.

neuroscience↗

bioRxiv2020

CRYPTOCHROMES confer robustness, not rhythmicity, to circadian timekeeping

SummaryCircadian (approximately daily) rhythms are a pervasive property of mammalian cells, tissues, and behaviour, ensuring physiological and metabolic adaptation to solar time. Models of daily cellular timekeeping revolve around transcriptional feedback repression, whereby CLOCK and BMAL1 activate the expression of clock proteins PERIOD (PER) and CRYPTOCHROME (CRY), which in turn repress CLOCK/BMAL1 activity. CRY proteins are thus considered essential negative regulators of the oscillation; a function supported by behavioural arrhythmicity of CRY-deficient mice when kept under constant conditions. Challenging this interpretation, however, we find evidence for persistent circadian rhythms in mouse behaviour and cellular PER2 levels when CRY is absent. CRY-less oscillations are variable in their expression and have a shorter period than wild type controls. Importantly, we find classic circadian hallmarks such as temperature compensation and determination of period by casein kinase 1{delta}/{varepsilon} activity to be maintained. In the absence of CRY-mediated transcriptional feedback repression and rhythmic Per2 transcription, PER2 protein rhythms are sustained for several cycles, accompanied by circadian variation in protein stability. We suggest that, whereas circadian transcriptional feedback imparts robustness and functionality onto biological clocks, the core timekeeping mechanism is post-translational. Our findings suggest that PER proteins normally act as signalling hubs that transduce timing information to the nucleus, imparting daily rhythms upon the activity of transcriptional effectors. Highlights[tpltrtarr] PER/CRY-mediated negative feedback is dispensable for mammalian circadian timekeeping [tpltrtarr]Circadian variation in PER2 levels persists in the absence of rhythmic Per2 transcription [tpltrtarr]CK1 and GSK3 are plausible mechanistic components of a cytoscillator mechanism [tpltrtarr]CRY-mediated feedback repression imparts robustness to biological timekeeping In briefCircadian turnover of mammalian clock protein PERIOD2 persists in the absence of canonical transcriptional feedback repression and rhythmic clock gene activity, demanding a re-evaluation of cellular clock function and evolution.

cell biology↗

bioRxiv2020

Human Influenza A Virus H1N1pdm Full-length Genome Sequencing in USA and Taiwan in 2015-16 Flu Season and Disease Correlation

During the 2015-16 winter, the US experienced a relatively mild influenza season compared to Taiwan which had a higher number of total and severe cases. While H1N1pdm viruses dominated global surveillance efforts that season, the global distribution of genetic variants and their contributions to disease severity have not been investigated. Samples collected from influenza A positive patients by the Johns Hopkins Center of Excellence for Influenza Research and Surveillance (JH-CEIRS) active surveillance in the emergency rooms in Baltimore, Maryland, USA and northern Taiwan between November 2015 and April 2016, were processed for influenza A virus whole genome sequencing. In Baltimore, the majority of the viruses were the H1N1pdm clade 6B.1 and no H1N1pdm clade 6B.2 viruses were detected. In northern Taiwan, more than half of the H1N1pdm viruses were clade 6B.1 and 38% were clade 6B.2, consistent with the global observation that most 6B.2 viruses circulated in Asia and not North America. Whole virus genome sequence analysis identified two genetic subgroups present in each of the 6B.1 and 6B.2 clades and one 6B.1 intraclade reassortant virus. Clinical data showed 6B.2 patients had more disease symptoms including higher crude and inverse probability weighted odds of pneumonia than 6B.1 patients, suggesting 6B.2 circulation may contribute to the severe flu season in Taiwan. Local surveillance efforts linking H1N1pdm virus sequences to patient clinical and demographic data improve our understanding of influenza circulation and disease potential.

microbiology↗

bioRxiv2020

A knock-in Drosophila model supports a conserved link between potassium channelopathy and involuntary movement

BackgroundGenetic and in vitro studies have linked a heterozygous gain-of-function mutation (D434G) in the hSlo1 BK (Big potassium) channel to paroxysmal dyskinesia. However, support for this linkage from in vivo models has been lacking. ObjectivesWe aimed to re-create the equivalent mutation to hSlo1 D434G in the fruit fly, Drosophila, and examine how this mutation altered movement and action potential waveforms. MethodsWe generated a knock-in Drosophila model of hSlo1 D434G. We used video-tracking and infra-red beam-break systems to test whether locomotion was altered in this model, and patch-clamp electrophysiology to determine how the mutation affected action potential waveforms. ResultsWe identified profound motor dysfunction and sporadic leg twitches, as well as a reduced width and an enhancement of the afterhyperpolarization phase of action potentials, in the model background. ConclusionOur results support a conserved relationship between enhanced BK channel function and disrupted motor control across distantly related species.

neuroscience↗

bioRxiv2017

Neurocalcin acts in a clock- and light-modulated dopaminergic pathway to promote night sleep in Drosophila

Primary dystonia is a hyperkinetic movement disorder linked to altered dopaminergic signaling and synaptic plasticity in regions of the brain involved in motor control. Mutations in HPCA, encoding the neuronal calcium sensor Hippocalcin, are associated with primary dystonia, suggesting a function for Hippocalcin in regulating the initiation and/or maintenance of activity. However, such a role for Hippocalcin or Hippocalcin homologs has yet to be demonstrated in vivo. Here we investigate the cellular and organismal functions of the Drosophila Hippocalcin homolog Neurocalcin (NCA), and define a role for NCA in promoting sleep by suppressing nighttime hyperactivity. We show that NCA acts in a common pathway with the D1-type Dop1R1 dopamine receptor and facilitates sleep by inhibiting neurotransmitter release from a multi-component activity-promoting circuit. Our results suggest conserved roles for Hippocalcin homologs in modulating motor control through dopaminergic pathways, suppressing aberrant movements in humans and inappropriate nighttime locomotion in Drosophila.

neuroscience↗

bioRxiv2017

Mupirocin-associated temporal changes in the nasal microbiota and host’s antimicrobial responses: A pilot study in healthy staphylococcal carriers

BackgroundHow mupirocin affects the human nasal microbiota over time remains uncharacterized.\n\nMethodsWe repeatedly sampled the anterior nares of four healthy staphylococcal carriers before and after mupirocin use. By sequencing bacterial 16S ribosomal cDNA, we characterized sequential changes in the carriage status, the nasal microbiota, and the hosts antimicrobial peptide expression up to 90 days after decolonization.\n\nResultsBefore mupirocin use, the nasal microbiota differed by the initial, culture-based staphylococcal carriage status, with Firmicutes (54.1%) being the most predominant in carriers and Proteobacteria (75.8%) in the only noncarrier. The nasal microbiota became less diverse (Shannon diversity: 1.33, 95% confidence interval [CI]: 1.06-1.54) immediately after decolonisation than that before decolonisation (1.78, 95%CI: 0.58-1.93). Based on results of differential abundance analysis, Firmicutes were significantly enriched (log2 fold changes [≥] 4, Benjamini-Hochberg adjusted P < .01) while Actinobacteria, particularly Corynbebacterium, were relatively depleted in samples from staphylococcal carriers. Results of nonmetric multidimensional scaling (NMDS) and constrained correspondence analysis (CCA) also suggested that the initial staphylococcal carriage status, human neutrophil peptide 1 levels, and sampling times were major contributors to the between-community dissimilarities (P for marginal permutation test: .014) though the significance attenuated when within-group correlation was considered (P for blocked permutation test: .047).\n\nConclusionThese findings suggest that large-scale investigations on antibiotic effects on the human nasal microbiota are warranted.

microbiology↗