The precision in which neural circuits are assembled during embryonic development plays a major role in defining the spectrum of behaviors present in the mature organism. The ability to move is an essential behavior displayed by all animals, and many of the neural circuits controlling vertebrate locomotor behaviors are composed of discrete groups of neurons residing within the spinal cord. Our research focuses on defining the genetic programs controlling the specificity of synaptic connections between sensory neurons, motor neurons, and limb muscles. We study a large family of genes encoding Hox transcription factors and their roles in controlling identity and connectivity of the diverse neurons that comprise the circuits required for coordinated movement.
One aspect of our work is aimed at understanding how developing spinal motor neurons acquire intrinsic properties that allow them to connect selectively to specific muscle groups and neurons. The combinatorial activities of Hox proteins appear to determine the selectivity of connections between motor neurons and muscle targets, but the downstream mechanisms that determine this specificity are not known. A potential output of Hox function is to control, directly or indirectly, the expression of surface molecules that define the specificity of target selection. Through genetic and microarray analysis of Hox mutants, we are attempting to identify and characterize the molecular effectors of Hox genes that control motor axon guidance in the limb and synaptic specificity at their target sites.
We have found that, within motor neurons, Hox proteins display distinct transcriptional activator and repressor functions in the control of downstream gene programs. Hox repressor function establishes mutually exclusive domains of Hox gene expression along the rostrocaudal axis, and within a given segment a Hox "repressillator" transcriptional network controls motor neuron fates. Once the initial Hox patterns are established, Hox activator function positively regulates motor neuron–specific genes and orchestrates the downstream programs required for synaptic specificity. How these diverse functions are encoded within Hox proteins is not known. To explore the molecular mechanisms that control the distinct transcriptional outputs of Hox proteins, we are identifying additional Hox cofactors in motor neurons and analyzing the regulation of target genes.
Hox proteins are widely expressed in developing embryos, suggesting the requirement for transcriptional cofactors to restrict the activities of Hox proteins in specific cell types. In addition to their distinct activator and repressor functions, Hox proteins also have activities that are specifically directed to motor neuron cell types. Our studies indicate that a vertebrate forkhead transcription factor, FoxP1, is a contextual cofactor for Hox proteins in the specification of motor neuron identities. In the absence of FoxP1 in mice, motor neurons fail to form appropriate connections with muscles, and the motor system appears to revert to an ancestral state. We are exploring how Hox-FoxP1 interactions control the large repertoire of motor neuron gene targets.
In addition to motor neurons Hox proteins localize to two other major classes of neurons in the spinal cord: sensory neurons and interneurons. We have found that expression of Hox factors in sensory neurons and interneurons parallels the segment-specific Hox patterns in motor neurons, suggesting that Hox factors are involved in the molecular matching of motor neurons with their synaptic partners in the formation of locomotor circuits. To explore a role for Hox factors in sensory-motor connectivity, we are using gene-targeting strategies in mouse to generate motor and sensory neurons that lack specific Hox proteins. Altering Hox expression may lead to the failure of these neurons to innervate their appropriate targets. To test this hypothesis, we are using anatomical assays to trace the projections of sensory and motor axons and to define the molecular and functional properties of their monosynaptic connections.
