Hernán López-Schier

The histone variant macroH2A is an epigenetic regulator of key developmental genes

The histone variants macroH2A1 and macroH2A2 are associated with X chromosome inactivation in female mammals. However, the physiological function of macroH2A proteins on autosomes is poorly understood. Microarray-based analysis in human male pluripotent cells uncovered occupancy of both macroH2A variants at many genes encoding key regulators of development and cell fate decisions. On these genes, the presence of macroH2A1+2 is a repressive mark that overlaps locally and functionally with Polycomb repressive complex 2. We demonstrate that macroH2A1+2 contribute to the fine-tuning of temporal activation of HOXA cluster genes during neuronal differentiation. Furthermore, elimination of macroH2A2 function in zebrafish embryos produced severe but specific phenotypes. Taken together, our data demonstrate that macroH2A variants constitute an important epigenetic mark involved in the concerted regulation of gene expression programs during cellular differentiation and vertebrate development.

A two-step mechanism underlies the planar polarization of regenerating sensory hair cells

The restoration of planar cell polarity is an essential but poorly understood step toward physiological recovery during sensory-organ regeneration. Investigating this issue in the lateral line of the zebrafish, we found that hair cells regenerate in pairs along a single axis established by the restricted localization and oriented division of their progenitors. By analyzing mutants lacking the planar-polarity determinant Vangl2, we ascertained that the uniaxial production of hair cells and the subsequent orientation of their hair bundles are controlled by distinct pathways, whose combination underlies the establishment of hair-cell orientation during development and regeneration. This mechanism may represent a general principle governing the long-term maintenance of planar cell polarity in remodeling epithelia.

Afferent Neurons of the Zebrafish Lateral Line Are Strict Selectors of Hair-Cell Orientation

Hair cells in the inner ear display a characteristic polarization of their apical stereocilia across the plane of the sensory epithelium. This planar orientation allows coherent transduction of mechanical stimuli because the axis of morphological polarity of the stereocilia corresponds to the direction of excitability of the hair cells. Neuromasts of the lateral line in fishes and amphibians form two intermingled populations of hair cells oriented at 180° relative to each other, however, creating a stimulus-polarity ambiguity. Therefore, it is unknown how these animals resolve the vectorial component of a mechanical stimulus. Using genetic mosaics and live imaging in transgenic zebrafish to visualize hair cells and neurons at single-cell resolution, we show that lateral-line afferents can recognize the planar polarization of hair cells. Each neuron forms synapses with hair cells of identical orientation to divide the neuromast into functional planar-polarity compartments. We also show that afferent neurons are strict selectors of polarity that can re-establish synapses with identically oriented targets during hair-cell regeneration. Our results provide the anatomical bases for the physiological models of signal-polarity resolution by the lateral line.

The transmembrane inner ear (Tmie) protein is essential for normal hearing and balance in the zebrafish

Little is known about the proteins that mediate mechanoelectrical transduction, the process by which acoustic and accelerational stimuli are transformed by hair cells of the inner ear into electrical signals. In our search for molecules involved in mechanotransduction, we discovered a line of deaf and uncoordinated zebrafish with defective hair-cell function. The hair cells of mutant larvae fail to incorporate fluorophores that normally traverse the transduction channels and their ears lack microphonic potentials in response to vibratory stimuli. Hair cells in the posterior lateral lines of mutants contain numerous lysosomes and have short, disordered hair bundles. Their stereocilia lack two components of the transduction apparatus, tip links and insertional plaques. Positional cloning revealed an early frameshift mutation in tmie, the zebrafish ortholog of the mammalian gene transmembrane inner ear. The mutant line therefore affords us an opportunity to investigate the role of the corresponding protein in mechanoelectrical transduction.

Neuronal Birth Order Identifies a Dimorphic Sensorineural Map

Spatially distributed sensory information is topographically mapped in the brain by point-to-point correspondence of connections between peripheral receptors and central target neurons. In fishes, for example, the axonal projections from the mechanosensory lateral line organize a somatotopic neural map. The lateral line provides hydrodynamic information for intricate behaviors such as navigation and prey detection. It also mediates fast startle reactions triggered by the Mauthner cell. However, it is not known how the lateralis neural map is built to subserve these contrasting behaviors. Here we reveal that birth order diversifies lateralis afferent neurons in the zebrafish. We demonstrate that early- and late-born lateralis afferents diverge along the main axes of the hindbrain to synapse with hundreds of second-order targets. However, early-born afferents projecting from primary neuromasts also assemble a separate map by converging on the lateral dendrite of the Mauthner cell, whereas projections from secondary neuromasts never make physical contact with the Mauthner cell. We also show that neuronal diversity and map topology occur normally in animals permanently deprived of mechanosensory activity. We conclude that neuronal birth order correlates with the assembly of neural submaps, whose combination is likely to govern appropriate behavioral reactions to the sensory context.

High-resolution live imaging reveals axon-glia interactions during peripheral nerve injury and repair in zebrafish.

ABSTRACT Neural damage is a devastating outcome of physical trauma. The glia are one of the main effectors of neuronal repair in the nervous system, but the dynamic interactions between peripheral neurons and Schwann cells during injury and regeneration remain incompletely characterized. Here, we combine laser microsurgery, genetic analysis, high-resolution intravital imaging and lattice light-sheet microscopy to study the interaction between Schwann cells and sensory neurons in a zebrafish model of neurotrauma. We found that chronic denervation by neuronal ablation leads to Schwann-cell death, whereas acute denervation by axonal severing does not affect the overall complexity and architecture of the glia. Neuronal-circuit regeneration begins when Schwann cells extend bridging processes to close the injury gap. Regenerating axons grow faster and directionally after the physiological clearing of distal debris by the Schwann cells. This might facilitate circuit repair by ensuring that axons are guided through unoccupied spaces within bands of Büngner towards their original peripheral target. Accordingly, in the absence of Schwann cells, regenerating axons are misrouted, impairing the re-innervation of sensory organs. Our results indicate that regenerating axons use haptotaxis as a directional cue during the reconstitution of a neural circuit. These findings have implications for therapies aimed at neurorepair, which will benefit from preserving the architecture of the peripheral glia during periods of denervation. Summary: Schwann cells are important components of the peripheral glia. We use microsurgery and high-resolution live imaging to show how Schwann cells control the regeneration of a sensorineural circuit.

Converging Axons Collectively Initiate and Maintain Synaptic Selectivity in a Constantly Remodeling Sensory Organ

Sensory receptors are the functional link between the environment and the brain. The repair of sensory organs enables animals to continuously detect environmental stimuli. However, receptor cell turnover can affect sensory acuity by changing neural connectivity patterns. In zebrafish, two to four postsynaptic lateralis afferent axons converge into individual peripheral mechanosensory organs called neuromasts, which contain hair cell receptors of opposing planar polarity. Yet, each axon exclusively synapses with hair cells of identical polarity during development and regeneration to transmit unidirectional mechanical signals to the brain. The mechanism that governs this exceptionally accurate and resilient synaptic selectivity remains unknown. We show here that converging axons are mutually dependent for polarity-selective connectivity. If rendered solitary, these axons establish simultaneous functional synapses with hair cells of opposing polarities to transmit bidirectional mechanical signals. Remarkably, nonselectivity by solitary axons can be corrected upon the reintroduction of additional axons. Collectively, our results suggest that lateralis synaptogenesis is intrinsically nonselective and that interaxonal interactions continuously rectify mismatched synapses. This dynamic organization of neural connectivity may represent a general solution to maintain coherent synaptic transmission from sensory organs undergoing frequent variations in the number and spatial distribution of receptor cells.

Developmental and architectural principles of the lateral-line neural map.

The transmission and central representation of sensory cues through the accurate construction of neural maps is essential for animals to react to environmental stimuli. Structural diversity of sensorineural maps along a continuum between discrete- and continuous-map architectures can influence behavior. The mechanosensory lateral line of fishes and amphibians, for example, detects complex hydrodynamics occurring around the animal body. It triggers innate fast escape reactions but also modulates complex navigation behaviors that require constant knowledge about the environment. The aim of this article is to summarize recent work in the zebrafish that has shed light on the development and structure of the lateralis neural map, which is helping to understand how individual sensory modalities generate appropriate behavioral responses to the sensory context.

Multispectral four-dimensional imaging reveals that evoked activity modulates peripheral arborization and the selection of plane-polarized targets by sensory neurons.

The polarity of apical stereocilia endows hair cells with directional excitability, which in turn enables animals to determine the vectorial component of a sound. Neuromasts of the lateral line of aquatic vertebrates harbor two populations of hair cells that are oriented at 180° relative to each other. The resulting sensory-vectorial ambiguity is solved by lateralis afferent neurons that discriminate between hair cells of opposite polarities to innervate only those with the same orientation. How neurons select identically oriented hair cells remains unknown. To gain insight into the mechanism that underlies this selection, we devised a simple method to gather dynamic morphometric information about axonal terminals in toto by four-dimensional imaging. Applying this strategy to the zebrafish allowed us to correlate hair cell orientation to single afferent neurons at subcellular resolution. Here we show that in zebrafish with absent hair cell mechanoreception, lateralis afferents arborize profusely in the periphery, display less stability, and make improper target selections. Central axons, however, show no dynamic changes and establish normal contacts with the Mauthner cell, a characteristic second-order target in the hindbrain. We propose that the hardwired developmental mechanisms that underlie peripheral arborization and target recognition are modulated by evoked hair cell activity. This interplay between intrinsic and extrinsic cues is essential for plane-polarized target selection by lateralis afferent neurons.

Delaying Gal4-Driven Gene Expression in the Zebrafish with Morpholinos and Gal80

The modular Gal4/UAS gene expression system has become an indispensable tool in modern biology. Several large-scale gene- and enhancer-trap screens in the zebrafish have generated hundreds of transgenic lines expressing Gal4 in unique patterns. However, the early embryonic expression of the Gal4 severely limits their use for studies on regeneration or behavior because UAS-driven effectors could disrupt normal organogenesis. To overcome this limitation, we explored the use of the Gal4 repressor Gal80 in transient assays and with stable transgenes to temporally control Gal4 activity. We also validated a strategy to delay Gal4-driven gene expression using a morpholino targeted to Gal4. The first approach is limited to transgenes expressing the native Gal4. The morphant approach can also be applied to transgenic lines expressing the Gal4-VP16 fusion protein. It promises to become a standard approach to delay Gal4-driven transgene expression and enhance the genetic toolkit for the zebrafish.