Time-lapse imaging and cell-specific expression profiling reveal dynamic branching and molecular determinants of a multi-dendritic nociceptor in C. elegans

Abstract

Nociceptive neurons innervate the skin with complex dendritic arbors that respond to pain-evoking stimuli such as harsh mechanical force or extreme temperatures. Here we describe the structure and development of a model nociceptor, the PVD neuron of C. elegans, and identify transcription factors that control morphogenesis of the PVD dendritic arbor. The two PVD neuron cell bodies occupy positions on either the right (PVDR) or left (PVDL) sides of the animal in posterior lateral locations. Imaging with a GFP reporter revealed a single axon projecting from the PVD soma to the ventral cord and an elaborate, highly-branched arbor of dendritic processes that envelop the animal with a web-like array directly beneath the skin. Dendritic branches emerge in a step-wise fashion during larval development and may use an existing network of peripheral nerve cords as guideposts for key branching decisions. Time-lapse imaging revealed that branching is highly dynamic with active extension and withdrawal and that PVD branch overlap is prevented by a contact-dependent self-avoidance, a mechanism that is also employed by sensory neurons in other organisms. With the goal of identifying genes that regulate dendritic morphogenesis, we used the mRNA tagging method to produce a gene expression profile of PVD during late larval development. This microarray experiment identified > 2,000 genes that are 1.5 X elevated relative to all larval cells. The enriched transcripts encode a wide range of proteins with potential roles in PVD function (e.g., DEG/ENaC and Trp channels) or development (e.g., UNC-5 and LIN-17/frizzled receptors). We used RNAi and genetic tests to screen 86 transcription factors from this list and identified eleven genes that specify PVD dendritic structure. These transcription factors appear to control discrete steps in PVD morphogenesis and may either promote or limit PVD branching at specific developmental stages. For example, time-lapse imaging revealed that the MEC-3 (LIM homeodomain) is required for branch initiation in early larval development whereas EGL-44 (TEAD domain) prevents ectopic PVD branching in the adult. A comparison of PVD-enriched transcripts to a microarray profile of mammalian nociceptors revealed homologous genes with potentially shared nociceptive functions. We conclude that PVD neurons display striking structural, functional and molecular similarities to nociceptive neurons from more complex organisms and can thus provide a useful model system in which to identify evolutionarily conserved determinants of nociceptor fate.

AFF-1 fusogen can rejuvenate the regenerative potential of adult dendritic trees via self-fusion

“…The elaborate PVD structure is highly dynamic in young animals, exhibiting constant growth and retraction of individual high-order branches (Oren-Suissa et al, 2010;Smith et al, 2010Smith et al, , 2012. To further understand the dynamics of the age-dependent dendritic tree remodeling, we followed individual animals over time and analyzed time-lapse movies.…”

“…We have been studying the role of fusion proteins in the PVD neuron, a polymodal nociceptor that senses harsh touch to the body, cold temperature and posture (Chatzigeorgiou et al, 2010;OrenSuissa et al, 2010;Smith et al, 2010;Tsalik et al, 2003). The PVD Fig.…”

“…1A) (Oren-Suissa et al, 2010). These dendritic trees arise and are dynamically maintained through several intrinsic and extrinsic genetic pathways (Chatzigeorgiou et al, 2010;Cohen et al, 2014;Dong et al, 2013Dong et al, , 2015Husson et al, 2012;Liu and Shen, 2011;Maniar et al, 2012;Oren-Suissa et al, 2010;Salzberg et al, 2013;Smith et al, 2010;Wei et al, 2015).…”

C. elegans bicd-1, homolog of theDrosophiladynein accessory factorBicaudal D, regulates the branching of PVD sensory neuron dendrites

“…PVD neurons are bilaterally symmetric nociceptors that respond to harsh mechanical stimuli and cold temperatures (Chatzigeorgiou et al, 2010;Way and Chalfie, 1989). Importantly, these neurons elaborate large and complex dendritic arbors which envelop the body of late-larval and adult-staged animals (OrenSuissa et al, 2010;Smith et al, 2010;Tsalik et al, 2003;Yassin et al, 2001), and provide an ideal system for studies aimed at identifying novel regulators of dendrite morphogenesis.…”

Dendritic diversification through transcription factor-mediated suppression of alternative morphologies

“…Sd and Vg therefore operate in several contexts to regulate Cut levels and promote cell diversification, and our results identify roles in differentiating neurons in the control of dendrite morphology. Similarly, in C. elegans, the sd homolog egl-44 suppresses late stage ectopic branching in multidendritic PVD neurons (Smith et al, 2010). Given these roles in dendritic branching in invertebrate neurons, and the broad expression of Tead1 (the mouse homolog of Sd) in the mouse brain (Lein et al, 2007), it will be interesting to examine whether TEAD transcription factors also regulate dendritic branching in vertebrate neurons.…”