Transcriptome profiles of distinct mechanoreceptive somatosensory neuron subtypes

The objective of the study was to identify molecules that are selectively expressed in proprioceptive sensory neurons (pSNs) in DRG. To try to achieve this goal, we performed RNAseq on genetically identified and FACS purified cohorts that either contained i) pSNs and rapidly-adapting low threshold mechanoreceptors (RA-LTMRs), iI) pSNs and slowly-adapting (SA)-LTMRs, or RA-LTMRs only. Differential expression analysis between these different sensory neuron cohorts identified multiple proprioceptor enriched transcripts. We conducted a molecular analysis of pSN, RA-LTMR and SA-LTMR cohorts on the basis of their differential PV and RX3 expression pattern. To do so, we took advantage of a previously generated Runx3 mutant allele, a PV:Cre driver, and the Cre-dependent Ai9:tdTomato (tdT) reporter (Taniuchi et al., 2002; Kramer et al., 2006; Madison et al., 2010), and separately crossed PV:Cre and Ai9:tdT alleles into the Rx3 mutant background. Subsequent intercrosses between these two strains resulted in some wild type Rx3 PV:Cre;Ai9:tdT animals, and some Rx3 mutant PV:Cre;Ai9:tdT animals (hereafter referred to as Rx3+/+;PV:tdT and Rx3-/-;PV:tdT respectively). In Rx3 mutants, pSNs fail to survive, and all remaining PVon neurons correspond to RA-LTMRs sensory neurons (Kramer et al., 2006; de Nooij et al., 2013). We isolated neonatal PV:tdT+ sensory neurons from dissociated Rx3+/+ (comprising pSNs and RA-LTMRs) and Rx3-/- DRG (comprising only RA-LTMRs), using fluorescent activated cell sorting (FACS). In addition, we used TrkC:GFP BAC transgenic mice, in which the GFP reporter is selectively expressed in Rx3onTrkCon neurons but not in Rx3offTrkCon neurons, to isolate a mixed population of neonatal pSNs and SA-LTMRs (Lee et al., 2012; de Nooij et al., 2013). We performed global transcriptome analysis (RNAseq) on each of the three isolated mechanoreceptor populations: Rx3+/+;PV:tdT+, Rx3-/-;PV:tdT+, and TrkC:GFP+.