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2529 Janelia Publications
Showing 1371-1380 of 2529 resultsAlthough myosin II filaments are known to exist in non-muscle cells, their dynamics and organization are incompletely understood. Here, we combined structured illumination microscopy with pharmacological and genetic perturbations, to study the process of actomyosin cytoskeleton self-organization into arcs and stress fibres. A striking feature of the myosin II filament organization was their 'registered' alignment into stacks, spanning up to several micrometres in the direction orthogonal to the parallel actin bundles. While turnover of individual myosin II filaments was fast (characteristic half-life time 60 s) and independent of actin filament turnover, the process of stack formation lasted a longer time (in the range of several minutes) and required myosin II contractility, as well as actin filament assembly/disassembly and crosslinking (dependent on formin Fmnl3, cofilin1 and α-actinin-4). Furthermore, myosin filament stack formation involved long-range movements of individual myosin filaments towards each other suggesting the existence of attractive forces between myosin II filaments. These forces, possibly transmitted via mechanical deformations of the intervening actin filament network, may in turn remodel the actomyosin cytoskeleton and drive its self-organization.
During their lifetime, animals must adapt their behavior to survive in changing environments. This ability requires the nervous system to adjust through dynamic expression of neurotransmitters and receptors but also through growth, spatial reorganization and connectivity while integrating external stimuli. For instance, despite having a fixed neuronal cell lineage, the nematode Caenorhabditis elegans’ nervous system remains plastic throughout its development. Here, we focus on a specific example of nervous system plasticity, the C. elegans dauer exit decision. Under unfavorable conditions, larvae will enter the non-feeding and non-reproductive dauer stage and adapt their behavior to cope with a new environment. Upon improved conditions, this stress resistant developmental stage is actively reversed to resume reproductive development. However, how different environmental stimuli regulate the exit decision mechanism and thereby drive the larva’s behavioral change is unknown. To fill this gap, we developed a new open hardware method for long-term imaging (12h) of C. elegans larvae. We identified dauer-specific behavioral motifs and characterized the behavioral trajectory of dauer exit in different environments to identify key decision points. Combining long-term behavioral imaging with transcriptomics, we find that bacterial ingestion triggers a change in neuropeptide gene expression to establish post-dauer behavior. Taken together, we show how a developing nervous system can robustly integrate environmental changes, activate a developmental switch and adapt the organism’s behavior to a new environment.
Starting a new research campus is a leap of faith. Only later, in the full measure of time, is it possible to take stock of what has worked and what could have been done better or differently. The Janelia Research Campus opened its doors 12 years ago. What has it achieved? What has it taught us? And where does Janelia go from here?
How do evolved genetic changes alter the nervous system to produce different patterns of behavior? We address this question using Drosophila male courtship behavior, which is innate, stereotyped, and evolves rapidly between species. D. melanogaster male courtship requires the male-specific isoforms of two transcription factors, fruitless and doublesex. These genes underlie genetic switches between female and male behaviors, making them excellent candidate genes for courtship behavior evolution. We tested their role in courtship evolution by transferring the entire locus for each gene from divergent species to D. melanogaster. We found that despite differences in Fru+ and Dsx+ cell numbers in wild-type species, cross-species transgenes rescued D. melanogaster courtship behavior and no species-specific behaviors were conferred. Therefore, fru and dsx are not a significant source of evolutionary variation in courtship behavior.
A low-contrast spot that activates just one ganglion cell in the retina is detected in the spike train of the cell with about the same sensitivity as it is detected behaviorally. This is consistent with Barlow’s proposal that the ganglion cell and later stages of spiking neurons transfer information essentially without loss. Yet, when losses of sensitivity by all preneural factors are accounted for, predicted sensitivity near threshold is considerably greater than behavioral sensitivity, implying that somewhere in the brain information is lost. We hypothesized that the losses occur mainly in the retina, where graded signals are processed by analog circuits that transfer information at high rates and low metabolic cost. To test this, we constructed a model that included all preneural losses for an in vitro mammalian retina, and evaluated the model to predict sensitivity at the cone output. Recording graded responses postsynaptic to the cones (from the type A horizontal cell) and comparing to predicted preneural sensitivity, we found substantial loss of sensitivity (4.2-fold) across the first visual synapse. Recording spike responses from brisk-transient ganglion cells stimulated with the same spot, we found a similar loss (3.5-fold) across the second synapse. The total retinal loss approximated the known overall loss, supporting the hypothesis that from stimulus to perception, most loss near threshold is retinal.
In animals, Hox transcription factors define regional identity in distinct anatomical domains. How Hox genes encode this specificity is a paradox, because different Hox proteins bind with high affinity in vitro to similar DNA sequences. Here, we demonstrate that the Hox protein Ultrabithorax (Ubx) in complex with its cofactor Extradenticle (Exd) bound specifically to clusters of very low affinity sites in enhancers of the shavenbaby gene of Drosophila. These low affinity sites conferred specificity for Ubx binding in vivo, but multiple clustered sites were required for robust expression when embryos developed in variable environments. Although most individual Ubx binding sites are not evolutionarily conserved, the overall enhancer architecture-clusters of low affinity binding sites-is maintained and required for enhancer function. Natural selection therefore works at the level of the enhancer, requiring a particular density of low affinity Ubx sites to confer both specific and robust expression.
Control of metabolism by compartmentation is a widespread feature of higher cells. Recent studies have focused on dynamic intracellular bodies such as stress granules, P-bodies, nucleoli, and metabolic puncta. These bodies appear as separate phases, some containing reversible, amyloid-like fibrils formed by interactions of low-complexity protein domains. Here we report five atomic structures of segments of low-complexity domains from granule-forming proteins, one determined to 1.1 Å resolution by micro-electron diffraction. Four of these interacting protein segments show common characteristics, all in contrast to pathogenic amyloid: kinked peptide backbones, small surface areas of interaction, and predominate attractions between aromatic side-chains. By computationally threading the human proteome on three of our kinked structures, we identified hundreds of low-complexity segments potentially capable of forming such reversible interactions. These segments are found in proteins as diverse as RNA binders, nuclear pore proteins, keratins, and cornified envelope proteins, consistent with the capacity of cells to form a wide variety of dynamic intracellular bodies.
Neurons in multiple brain regions fire trains of action potentials anticipating specific movements, but this 'preparatory activity' has not been systematically compared across behavioral tasks. We compared preparatory activity in auditory and tactile delayed-response tasks in male mice. Skilled, directional licking was the motor output. The anterior lateral motor cortex (ALM) is necessary for motor planning in both tasks. Multiple features of ALM preparatory activity during the delay epoch were similar across tasks. First, majority of neurons showed direction-selective activity and spatially intermingled neurons were selective for either movement direction. Second, many cells showed mixed coding of sensory stimulus and licking direction, with a bias toward licking direction. Third, delay activity was monotonic and low-dimensional. Fourth, pairs of neurons with similar direction selectivity showed high spike-count correlations. Our study forms the foundation to analyze the neural circuit mechanisms underlying preparatory activity in a genetically tractable model organism.Short-term memories link events separated in time. Neurons in frontal cortex fire trains of action potentials anticipating specific movements, often seconds before the movement. This 'preparatory activity' has been observed in multiple brain regions, but has rarely been compared systematically across behavioral tasks in the same brain region. To identify common features of preparatory activity, we developed and compared preparatory activity in auditory and tactile delayed-response tasks in mice. The same cortical area is necessary for both tasks. Multiple features of preparatory activity, measured with high-density silicon probes, were similar across tasks. We find that preparatory activity is low-dimensional and monotonic. Our study forms the foundation to analyze the circuit mechanisms underlying preparatory activity in a genetically tractable model organism.
Real-time neural signal processing is essential for brain-machine interfaces and closed-loop neuronal perturbations. However, most existing applications sacrifice cell-specific identity and temporal spiking information for speed. We developed a hybrid hardware-software system that utilizes a Field Programmable Gate Array (FPGA) chip to acquire and process data in parallel, enabling individual spikes from many simultaneously recorded neurons to be assigned single-neuron identities with 1-millisecond latency. The FPGA assigns labels, validated with ground-truth data, by comparing multichannel spike waveforms from tetrode or silicon probe recordings to a spike-sorted model generated offline in software. This platform allowed us to rapidly inactivate a region in vivo based on spikes from an upstream neuron before these spikes could excite the downstream region. Furthermore, we could decode animal location within 3 ms using data from a population of individual hippocampal neurons. These results demonstrate our system’s suitability for a broad spectrum of research and clinical applications.
Cortical spike trains often appear noisy, with the timing and number of spikes varying across repetitions of stimuli. Spiking variability can arise from internal (behavioral state, unreliable neurons, or chaotic dynamics in neural circuits) and external (uncontrolled behavior or sensory stimuli) sources. The amount of irreducible internal noise in spike trains, an important constraint on models of cortical networks, has been difficult to estimate, since behavior and brain state must be precisely controlled or tracked. We recorded from excitatory barrel cortex neurons in layer 4 during active behavior, where mice control tactile input through learned whisker movements. Touch was the dominant sensorimotor feature, with >70% spikes occurring in millisecond timescale epochs after touch onset. The variance of touch responses was smaller than expected from Poisson processes, often reaching the theoretical minimum. Layer 4 spike trains thus reflect the millisecond-timescale structure of tactile input with little noise.