Abstract
Promoter-based genetic recombination (via, e.g., Cre-lox) is most useful when all cells of interest express a particular gene. The discovery that the actin-binding protein advillin is expressed in all somatic sensory neurons has been exploited repeatedly to drive DNA recombination therein, yet specificity of expression has not been well demonstrated. Here, we characterize advillin expression amongst sensory neurons and in several other neural and non-neural tissues. We first validate an advillin antibody against advillin knock-out tissue, advillin promoter-driven EGFP, and advillin mRNA expression. In the dorsal root ganglion (DRG), advillin is enriched in non-peptidergic nociceptors. We also show that advillin expression, and advillin promotor-driven EGFP and Cre-recombinase expression, occurs in multiple tissues including the dorsal habenula of the epithalamus, endocrine cells of the gut, Merkel cells in the skin, and most strikingly, throughout the autonomic nervous system (sympathetic, parasympathetic, and enteric neurons) in mice, rats, and non-human primates. In the mouse pelvic ganglion, advillin immunoreactivity is most intense in pairs of small neurons, and concentrated in spine-like structures on the axon initial segment contacted by sympathetic preganglionic axons. In autonomic targets (iris and blood vessels), advillin is distributed along cholinergic parasympathetic axons and in sympathetic varicosities. Developmentally, advillin expression is absent from sympathetics at postnatal day 4 but begins to emerge by day 7, accounting for previous reports (based on embryonic expression) of advillin’s specificity to sensory neurons. These results indicate that caution is warranted in interpreting previous studies in which advillin-driven genomic editing is either constitutive or performed after postnatal day 4.
Highlights
The ability to manipulate gene expression within specific cells using recombinase driver (e.g., Cre, Dre, Flp) mice has proved a useful tool in studying gene function
We examined the relationship between EGFP and advillin expression in the dorsal root ganglion (DRG) of advillinEGFP mice
Advillin promoter activity, and advillin mRNA is expressed in the brain in mesencephalic trigeminal nucleus sensory neurons, and in the dorsal habenula (Fig. 1D)
Summary
The ability to manipulate gene expression within specific cells using recombinase driver (e.g., Cre, Dre, Flp) mice has proved a useful tool in studying gene function. Identifying appropriate promoters, expressed exclusively and ubiquitously within a cell population of interest, to drive recombinase expression is a crucial and challenging first step This is true for primary sensory neurons of the dorsal root ganglion (DRG), which represent an extremely heterogeneous population that have significant overlap in gene expression with other peripheral neurons (Usoskin et al, 2015). Other sensory neuron populations can be targeted with other subtype-specific promoters (e.g., parvalbumin for proprioceptors, Ret for low-threshold cutaneous mechanoreceptors; Niu et al, 2013; Chiu et al, 2014). These promoters target only some DRG neurons and their expression in other tissues reduces their overall utility. The discovery of the expression patterns of the advillin gene generated significant interest
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