Biology subjects

Mooney, R.

Publications and source records attributed to Mooney, R..

13 recordsLinked to original sources

bioRxiv2026

RNAP-seq: in vitro genome-scale transcription reveals preferential RNA polymerase pausing on Clostridioides difficile antisense DNA

Mechanistic dissection of gene transcription and its regulation in diverse organisms remains a central challenge for molecular biology. In vivo analyses using high-throughput DNA sequencing readouts (e.g., RNA-seq, Term-seq, and NET-seq) have revealed that transcriptional pausing by RNA polymerase (RNAP) underpins the regulation of gene transcription. However, these in vivo methods are difficult to apply in non-model organisms and lack the power of bottom-up biochemical analysis that can deconvolute the confounding effects of the cellular environment. Here we report a method for unbiased transcription of entire genomes in vitro called RNAP-seq, which is capable of defining pause, arrest, and termination sites with single-nucleotide resolution and of defining the regulatory contributions of cis-acting RNA and DNA sequences and dissociable transcription factors with complete control over transcription conditions. Using RNAP-seq to compare transcription by recombinant RNAPs from Clostridioides difficile (Cdf) and Escherichia coli (Eco), we discovered that CdfRNAP exhibits pausing signatures distinct from EcoRNAP. CdfRNAP pauses more frequently and more strongly at T-rich sequences, particularly on antisense regions of genes. We also defined the action of CdfNusG at genome scale, revealing its preferential action in riboswitch control regions. These findings demonstrate lineage-specific pausing features and suggest that genome composition and RNAP co-evolve to shape gene expression. RNAP-seq is broadly adaptable to diverse bacterial species and offers a powerful framework for uncovering fundamental principles of transcription that govern gene expression. HighlightsO_LIRNAP-seq enables unbiased transcription of every base-pair in an organisms genome C_LIO_LIRNAP-seq reveals distinct E. coli (Eco) and C. difficile (Cdf) pause sequences C_LIO_LICdfNusG broadly enhances pausing and termination, notably in riboswitch-encoding DNA C_LIO_LICdfRNAP but not EcoRNAP pauses more on the T-rich antisense strand of Cdf genes C_LI

genomics↗

bioRxiv2026

Brain-wide mapping of neuroanatomical connections to the auditory cortex of hearing and deaf mice

Remarkable therapeutic innovations have made it possible to establish hearing in congenitally deaf subjects. Despite these advances, a potential obstacle to restoring auditory function is that the absence of auditory experience alters the connectivity of the auditory cortex, a region that contributes to auditory perception and cognition. Here we used an intersectional genetic approach to map the brainwide inputs to the primary auditory cortex of congenitally deaf mice and their hearing littermates. We found that deaf mice displayed a significant reduction in afferents arising from the basomedial amygdala, the core of the medial geniculate nucleus, and anterior auditory thalamic nuclei. Nonetheless, major aspects of auditory cortical connectivity, including input from other thalamic nuclei and from non-auditory regions of the cortex, were unaffected by deafness. These findings highlight altered and preserved connectivity of the auditory cortex in the absence of auditory experience, which may inform therapies designed to establish hearing in congenitally deaf subjects.

neuroscience↗

bioRxiv2026

A synaptic locus of song learning

Learning by imitation is the foundation for verbal and musical expression, but its underlying neural basis remains obscure. A juvenile male zebra finch imitates the multisyllabic song of an adult tutor in a process that depends on a song-specialized cortico-basal ganglia circuit, affording a powerful system to identify the synaptic substrates of imitative motor learning. Plasticity at a particular set of cortico-basal ganglia synapses is hypothesized to drive rapid learning-related changes in song before these changes are subsequently consolidated in downstream circuits. Nevertheless, this hypothesis is untested and the synaptic locus where learning initially occurs is unknown. By combining a computational framework to quantify song learning with synapse-specific optogenetic and chemogenetic manipulations within and directly downstream of the cortico-basal ganglia circuit, we identified the specific cortico-basal ganglia synapses that drive the acquisition and expression of rapid vocal changes during juvenile song learning and characterized the hours-long timescale over which these changes consolidate. Furthermore, transiently augmenting postsynaptic activity in the basal ganglia briefly accelerates learning rates and persistently alters song, demonstrating a direct link between basal ganglia activity and rapid learning. These results localize the specific cortico-basal ganglia synapses that enable a juvenile songbird to learn to sing and reveal the circuit logic and behavioral timescales of this imitative learning paradigm.

neuroscience↗

bioRxiv2025

Correctness is its own reward: bootstrapping error signals in self-guided reinforcement learning

Reinforcement learning (RL) offers a compelling account of how agents learn complex behaviors by trial and error, yet RL is predicated on the existence of a reward function provided by the agents environment. By contrast, many skills are learned without external guidance, posing a challenge to RLs ability to account for self-directed learning. For instance, juvenile male zebra finches first memorize and then train themselves to reproduce the song of an adult male tutor through extensive practice. This process is believed to be guided by an internally computed assessment of performance quality, though the mechanism and development of this signal remain unknown. Here, we propose that, contrary to prevailing assumptions, tutor song memorization and performance assessment are subserved by the same neural circuit, one trained to predictively cancel tutor song. To test this hypothesis, we built models of a local forebrain circuit that uses contextual premotor signals to cancel tutor song auditory input via synaptic plasticity. After learning, excitatory projection neurons signaled mismatches between the tutor song and birds own performance, best matching experimental data when learning involved anti-Hebbian plasticity in recurrent interneurons. We also found that learning proceeds by both sharpening error sensitivity and minimizing responses to the tutor song. Finally, the error signals produced by this model can train a simple RL agent to replicate the spectrograms of adult bird songs. These results suggest that local learning via predictive cancellation suffices for bootstrapping error signals capable of guiding self-directed learning of natural behaviors. SignificanceMany species, including humans, learn new skills by trial and error. But in the absence of external rewards and punishments, individuals need a means of assessing the quality of their performance. Here, we model the process by which juvenile male zebra finches memorize the song of an adult tutor, showing how local changes in brain circuits responsible for hearing can give rise during this process to an internal error signal that later allows for self-guided learning.

neuroscience↗

bioRxiv2024

A Cortical Site that Encodes Vocal Expression and Reception

Socially effective vocal communication requires brain regions that encode expressive and receptive aspects of vocal communication in a social context-dependent manner. Here, we combined a novel behavioral assay with microendoscopy to interrogate neuronal activity in the posterior insula (pIns) in socially interacting mice as they switched rapidly between states of vocal expression and reception. We found that distinct but spatially intermingled subsets of pIns neurons were active during vocal expression and reception. Notably, pIns activity during vocal expression increased prior to vocal onset and was also detected in congenitally deaf mice, pointing to a motor signal. Furthermore, receptive pIns activity depended strongly on social cues, including female odorants. Lastly, tracing experiments reveal that deep layer neurons in the pIns directly bridge the auditory thalamus to a midbrain vocal gating region. Therefore, the pIns is a site that encodes vocal expression and reception in a manner that depends on social context.

neuroscience↗

bioRxiv2022

Nested circuits mediate the decision to vocalize

Vocalizations facilitate mating and social affiliation, but may also inadvertently alert predators and rivals. Consequently, the decision to vocalize depends on brain circuits that can weigh and compare these potential benefits and risks. Male mice produce ultrasonic vocalizations (USVs) during courtship to facilitate mating and female mice produce USVs to promote social affiliation with each other. Previously we showed that a specialized set of neurons in the midbrain periaqueductal gray (PAG) are an obligatory gate for USV production in both male and female mice, and that both PAG-USV neurons and USVs can be switched on by their inputs from the preoptic area (POA) of the hypothalamus and switched off by their inputs from neurons on the border between the central and medial amygdala (AmgC/M-PAG neurons) (Michael et al., 2020). Here we show that the USV-suppressing AmgC/M-PAG neurons are strongly activated by predator cues or during social contexts that suppress USV production in male and female mice. Furthermore, a subset of USV-promoting POA neurons that provide input to the PAG-USV region also extend axon collaterals to the amygdala, where they directly inhibit AmgC/M-PAG neurons. Accordingly, AmgC/M-PAG neurons, along with POAPAG and PAG-USV neurons, form a nested hierarchical circuit in which environmental and social information converges to influence the decision to vocalize.

neuroscience↗

bioRxiv2022

Divergent projections from Locus Coeruleus to the cortico-basal ganglia system and ventral tegmental area of the zebra finch.

The locus coeruleus (LC) is a small brainstem nucleus that is the primary source of noradrenaline (NA) throughout the vertebrate brain. Understanding the anatomical organization of the LC is important because NA plays a central role in regulating arousal, attention, and performance. In the mammalian brain, individual LC neurons make highly divergent axonal projections to different brain regions, which are distinguished in part by which NA receptor subtypes they express. Here we sought to determine whether similar organizational features - divergent LC projections acting regionally through different receptor subtypes - characterize cortico-basal ganglia (CBG) circuitry important to birdsong learning and performance. Single and dual retrograde tracer injections reveal that single LC-NA neurons make divergent projections to both cortical and basal ganglia components of this circuit, as well as to dopaminergic nuclei that innervate this circuit. Moreover, in situ hybridization revealed that differential expression of the 2A and 2C adrenoreceptor distinguish LC-recipient song nuclei. Therefore, LC - NA signaling in the songbird CBG circuit could employ a similar strategy as in mammals, which allows a relatively small number of LC neurons to exert widespread yet distinct effects across multiple brain regions. Key PointsO_LIThe locus coeruleus projects to most of the song system (HVC, RA, LMAN, Area X, DLM). C_LIO_LINoradrenergic receptors are regionally specialized in BG despite divergent connectivity of LC neurons. C_LIO_LINoradrenergic projections to dopaminergic nuclei could influence vocal variability and learning. C_LI

neuroscience↗

bioRxiv2021

A neural hub that coordinates learned and innate courtship behaviors

Holistic behaviors often require the coordination of innate and learned movements. The neural circuits that enable such coordination remain unknown. Here we identify a midbrain cell group (A11) that enables male zebra finches to coordinate their learned songs with various innate behaviors, including female-directed calling, orientation and pursuit. Anatomical mapping reveals that A11 is at the center of a complex network including the song premotor nucleus HVC as well as brainstem regions crucial to innate calling and locomotion. Notably, lesioning A11 terminals in HVC blocked female-directed singing, but did not interfere with female-directed calling, orientation or pursuit. In contrast, lesioning A11 cell bodies abolished all female-directed courtship behaviors. However, males with either type of lesion still produced songs when in social isolation. Lastly, monitoring A11 terminals in HVC showed that during courtship A11 inputs to the song premotor cortex signal the transition from innate to learned vocalizations. These results show how a brain region important to reproduction in both birds and mammals coordinates learned vocalizations with innate, ancestral courtship behaviors.

neuroscience↗

bioRxiv2019

A novel ribosomal protein 20 variant in a family with unexplained colorectal cancer and polyposis

Colorectal cancer (CRC) has a large hereditary component, which is only partially explained by known genetic causes. Recently, variants in ribosomal protein S20 (RPS20, [OMIM: 603682]) were identified in a family with familial CRC type X and in a CRC cancer case-control screen. This study describes a novel splice donor variant in RPS20, NM_001023.3:c.177+1G>A. It segregates with CRC [OMIM: 114500] and polyposis [HP: 0200063] within the probands family. Reverse transcription-polymerase chain reaction (RT-PCR) confirms the variant results in two aberrantly-spliced transcripts that are absent in controls. The location of the novel RPS20 variant is near two previously-reported truncating RPS20 variants associated with CRC. DNA from colon adenocarcinoma showed no evidence of loss-of-heterozygosity, supporting a haploinsufficiency or dominant negative disease mechanism. These findings support designation of RPS20 as a CRC predisposition gene, and expand the phenotypic spectrum of RPS20 truncating variants to include polyposis.

genetics↗

bioRxiv2019

Circuit and synaptic organization of forebrain-to-midbrain pathways that promote and suppress vocalization

Animals vocalize only in certain behavioral contexts, but the circuits and synapses through which forebrain neurons trigger or suppress vocalization remain unknown. Here we used transsynaptic tracing to identify two populations of inhibitory neurons that lie upstream of neurons in the periaqueductal gray that gate the production of ultrasonic vocalizations in mice (i.e., PAG-USV neurons). Activating PAG-projecting neurons in the preoptic hypothalamus (POAPAG neurons) elicited USV production in the absence of social cues. In contrast, activating PAG-projecting neurons in the central-medial boundary zone of the amygdala (AmgC/M-PAG neurons) transiently suppressed USV production without disrupting non-vocal social behavior. Optogenetics-assisted circuit mapping in brain slices revealed that POAPAG neurons directly inhibit PAG interneurons, which in turn inhibit PAG-USV neurons, whereas AmgC/M-PAG neurons directly inhibit PAG-USV neurons. These experiments identify two major forebrain inputs to the PAG that trigger and suppress vocalization, respectively, while also establishing the synaptic mechanisms through which these neurons exert opposing behavioral effects.

neuroscience↗

bioRxiv2019

Inferring low-dimensional latent descriptions of animal vocalizations

Increases in the scale and complexity of behavioral data pose an increasing challenge for data analysis. A common strategy involves replacing entire behaviors with small numbers of handpicked, domain-specific features, but this approach suffers from several crucial limitations. For example, handpicked features may miss important dimensions of variability, and correlations among them complicate statistical testing. Here, by contrast, we apply the variational autoencoder (VAE), an unsupervised learning method, to learn features directly from data and quantify the vocal behavior of two model species: the laboratory mouse and the zebra finch. The VAE converges on a parsimonious representation that outperforms handpicked features on a variety of common analysis tasks, enables the measurement of moment-by-moment vocal variability on the timescale of tens of milliseconds in the zebra finch, provides strong evidence that mouse ultrasonic vocalizations do not cluster as is commonly believed, and captures the similarity of tutor and pupil birdsong with qualitatively higher fidelity than previous approaches. In all, we demonstrate the utility of modern unsupervised learning approaches to the quantification of complex and high-dimensional vocal behavior.

animal behavior and cognition↗

bioRxiv2018

MIN1PIPE: A Miniscope 1-photon-based Calcium Imaging Signal Extraction Pipeline

In vivo calcium imaging using 1-photon based miniscope and microendoscopic lens enables studies of neural activities in freely behaving animals. However, the high and fluctuating background, the inevitable movements and distortions of imaging field, and the extensive spatial overlaps of fluorescent signals emitted from imaged neurons inherent in this 1-photon imaging method present major challenges for extracting neuronal signals reliably and automatically from the raw imaging data. Here we develop a unifying algorithm called MINiscope 1-photon imaging PIPEline (MIN1PIPE) that contains several standalone modules and can handle a wide range of imaging conditions and qualities with minimal parameter tuning, and automatically and accurately isolate spatially localized neural signals. We quantitatively compare MIN1PIPE with other existing partial methods using both synthetic and real datasets obtained from different animal models, and show that MIN1PIPE has a superior performance both in terms of efficiency and precision in analyzing noisy miniscope calcium imaging data.

neuroscience↗

bioRxiv2018

Identification of midbrain neurons essential for vocal communication

Vocalizations are an essential medium for communication and courtship in numerous mammalian species ranging from mice to humans. In mammals, the midbrain PAG serves as an obligatory node in a vocalization-related network that spans the forebrain and brainstem1-3, as bilateral lesions of the PAG result in mutism2-5. Despite the PAGs importance for vocal production, the identity, function, and connectivity of PAG neurons involved in vocalization has remained elusive, in part because the PAG is a functionally and anatomically heterogeneous structure that serves myriad roles including nociception, defensive behaviors, and autonomic regulation6-9. Here we used a viral genetic \"tagging\" method10,11 to identify a distinct subset of PAG neurons in the male mouse that are selectively activated during the production of ultrasonic vocalizations (USVs) elicited by female cues. Silencing these PAG-USV neurons rendered males mute without affecting their other courtship behaviors and also impaired their ability to attract female mice in a social choice assay. Activating these neurons using chemogenetic or optogenetic methods strongly elevated USV production, even in the absence of female cues. Notably, the timing of individual USVs was entrained to the expiratory phase of breathing but not to the pattern of optogenetic stimulation, suggesting that PAG-USV neural activity initiates and sets the duration of vocal bouts and recruits downstream premotor circuits that precisely pattern vocal output. Consistent with this idea, we found that PAG-USV neurons extend axons into pontine and medullary regions that are speculated to contain premotor central pattern generators important for vocalization3,12,13. These experiments establish the identity of the PAG neurons selectively required for USV production in mice, map their efferent connections, and demonstrate the communicative salience of male USVs in promoting female social affiliation.

neuroscience↗