Sustained Hox5 gene activity is required for respiratory motor neuron development

Abstract

Respiration in mammals relies on the rhythmic firing of neurons within the Phrenic Motor Column (PMC), a motor neuron group that provides the sole source of diaphragm innervation. Despite their essential role in breathing, the specific determinants of PMC identity and patterns of connectivity are largely unknown. We show that two Hox genes, Hoxa5 and Hoxc5, control diverse aspects of PMC development including their clustering, intramuscular branching, and survival. In mice lacking Hox5 genes in motor neurons, axons extend to the diaphragm but fail to arborize, leading to respiratory failure. Genetic rescue of cell death fails to restore columnar organization and branching patterns, indicating these defects are independent of neuronal loss. Unexpectedly, late Hox5 removal preserves columnar organization but depletes PMC number and branches, demonstrating a continuous requirement for Hox function in motor neurons. These findings indicate that Hox5 genes orchestrate PMC development through deployment of temporally distinct wiring programs.

How to make spinal motor neurons

“…Experimental manipulation of the Hox code in mouse and chick embryos can alter MN subtype and projection pattern (Tiret et al, 1998 Misra et al, 2009;Jung et al, 2010;Philippidou et al, 2012;Lacombe et al, 2013). As a particularly dramatic example, mutation of Hoxc9 in the mouse causes loss of thoracic preganglionic column (PGC) and hypaxial motor column (HMC) MNs and expansion of the brachial lateral motor column (LMC) domain (Jung et al, 2010).…”

The role of Sema3–Npn-1 signaling during diaphragm innervation and muscle development

“…Similar phrenic nerve guidance defects into the limb or epiaxial mesenchyme in Npn-1 Sema−/− and Npn-1 cond−/− ;Olig2-Cre + mutants cannot be completely excluded, and might be uncovered by the combination of retrograde tracings of limb or epaxial projecting axons and specific neuronal markers of the PMC. One mechanism that might govern correct early fasciculation of the phrenic nerves is the interaction of the transmembrane glycoprotein activated leukocyte cell adhesion molecule (ALCAM) with its membrane-bound binding partner CD6, as it is specifically expressed at brachial levels in motor neurons of the PMC or all somatic motor neurons, respectively, and has been shown to be involved in axon fasciculation (Philippidou et al, 2012;Pourquié et al, 1990;Weiner et al, 2004).…”

“…The substantial incidence of neonatal lethality might indicate that the degree of defasciculation and therewith myofiber innervation might have an effect on neonatal survival. For example, in animals where Hox5 and Unc5c were mutated, a profound reduction of costal muscle innervation of the diaphragm, which leads to neonatal cyanosis, is observed (Burgess et al, 2006;Philippidou et al, 2012). Furthermore, systemic ablation of erbB2 results in a similar defasciculation of the phrenic nerves during innervation of both diaphragm muscles.…”

“…Therefore, faithful innervation by the phrenic nerves is a requirement for the functionality of the highly specialized diaphragm and thus, ultimately, for the viability of the entire organism. Retrograde labeling experiments have revealed that phrenic motor neurons within the cervical spinal cord are demarcated by the expression of several transcription factors, most notably Hoxa5, Hoxc5, Isl1 and Scip (also known as Pou3f1), whereas they exclude the transcription factor FoxP1, which labels limb-innervating motor neurons (Bermingham et al, 1996;Dasen et al, 2008;Philippidou et al, 2012). Indeed, mice in which Hoxa5 and Hoxc5 are ablated show a severe loss of Scip-positive phrenic motor neurons, whereas genetic elimination of FoxP1 induces a fate change of brachial motor neurons to acquire phrenic motor neuron characteristics (Dasen et al, 2008;Philippidou et al, 2012).…”

Neural development and regeneration: it's all in your spinal cord

“…Another important factor is Hoxa5, investigated by Polyxeni Philippidou (Dasen lab, NYU Medical Center, USA) to understand the control of phrenic MN (PMN) identity in the development of respiratory circuits (Philippidou et al, 2012). Hoxa5 conditional knockout mice, which have fewer PMNs and thinner phrenic nerves, show dramatic changes in the dendritic patterns of PMNs and increased frequency of respiratory bursts, pointing to defects in the integration of PMNs into respiratory circuitry.…”