Highlights d In somatosensory ganglia, Prdm12 is specific to the nociceptive lineage d Prdm12 is necessary for the survival of developing nociceptors d Prdm12 initiates and maintains the expression of TrkA in developing nociceptors d Prdm12 acts in conjunction with bHLH proteins Ngn1/2 to promote a nociceptor fate

The blood-brain barrier (BBB) protects the central nervous system (CNS) from harmful blood-borne factors. Although BBB dysfunction is a hallmark of several neurological disorders, therapies to restore BBB function are lacking. An attractive strategy is to repurpose developmental BBB regulators, such as Wnt7a, into BBB-protective agents. However, safe therapeutic use of Wnt ligands is complicated by their pleiotropic Frizzled signaling activities. Taking advantage of the Wnt7a/b-specific Gpr124/Reck co-receptor complex, we genetically engineered Wnt7a ligands into BBB-specific Wnt activators. In a “hit-and-run” adeno-associated virus–assisted CNS gene delivery setting, these new Gpr124/Reck-specific agonists protected BBB function, thereby mitigating glioblastoma expansion and ischemic stroke infarction. This work reveals that the signaling specificity of Wnt ligands is adjustable and defines a modality to treat CNS disorders by normalizing the BBB.

These authors contributed equally to this work.Keywords: Hamlet, nociception, PRDM12, sensory neurons, TRHDE Abbreviation: PRDM12, PR homology domain-containing member 12; HSAN, hereditary and sensory autonomic neuropathy; TRHDE, tyrotropin-releasing hormone degrading enzyme; RA, retinoic acid; Brn3d, brain 3d; Tlx3, T-cell leukemia homeobox 3; Hmx3, H6 family homeobox 3; Drgx, dorsal root ganglia homeobox; Sncg, Synuclein Gamma (Breast Cancer-Specific Protein 1); En1, engrailed-1; RT-qPCR, real-time quantitative polymerase chain reaction; MO, morpholino oligonucleotide; ChIP, chromatin immunoprecipitation; S1PR1, Sphi8ngosine-1-phosphate receptor 1; CGNL1, cyclin L1; IL1R1, interleukin 1 receptor type 1; TRH(DE), tryrotropin-releasing hormone degrading enzyme; pCMV6, plasmid cytomegalovirus; DDK, DYKDDDDK epitope; GAPDH, glyceraldehyde 3-phospate dehydrogenase; NBT/BCIP, nitro blue tetrazolium / 5-bromo-4-chloro-3-indolyl-phosphate; SET, Su(var)3-9 and 'Enhancer of zeste'; PDB, protein data base; HEK293, human embryonic kidney cell 293; PBS, phosphate buffered saline; DAPI, 4 0 ,6-diamidino-2-phenylindole; GFP, green fluorescent protein; BSA, bovine serum albumin; HRP, horseraddish peroxidase; ECL, enhanced chemiluminescence; FPKM, fragments per kilobase exon; FDR, false discovery rate; GEO, gene expression omnibus; PMID, pubmed identification.PR homology domain-containing member 12 (PRDM12) belongs to a family of conserved transcription factors implicated in cell fate decisions. Here we show that PRDM12 is a key regulator of sensory neuronal specification in Xenopus. Modeling of human PRDM12 mutations that cause hereditary sensory and autonomic neuropathy (HSAN) revealed remarkable conservation of the mutated residues in evolution. Expression of wild-type human PRDM12 in Xenopus induced the expression of sensory neuronal markers, which was reduced using various human PRDM12 mutants. In Drosophila, we identified Hamlet as the functional PRDM12 homolog that controls nociceptive behavior in sensory neurons. Furthermore, expression analysis of human patient fibroblasts with PRDM12 mutations uncovered possible downstream target genes. Knockdown of several of these target genes including thyrotropin-releasing hormone degrading enzyme (TRHDE) in Drosophila sensory neurons resulted in altered cellular morphology and impaired nociception. These data show that PRDM12 and its functional fly homolog Hamlet are evolutionary conserved master regulators of sensory neuronal specification and play a critical role in pain perception. Our data also uncover novel pathways in multiple species that regulate evolutionary conserved nociception.

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Prdm12 is a conserved epigenetic transcriptional regulator that displays restricted expression in nociceptors of the developing peripheral nervous system. In mice, Prdm12 is required for the development of the entire nociceptive lineage. In humans, PRDM12 mutations cause congenital insensitivity to pain, likely because of the loss of nociceptors. Prdm12 expression is maintained in mature nociceptors suggesting a yet-to-be explored functional role in adults. Using Prdm12 inducible conditional knockout mouse models, we report that in adult nociceptors Prdm12 is no longer required for cell survival but continues to play a role in the transcriptional control of a network of genes, many of them encoding ion channels and receptors. We found that disruption of Prdm12 alters the excitability of dorsal root ganglion neurons in culture. Phenotypically, we observed that mice lacking Prdm12 exhibit normal responses to thermal and mechanical nociceptive stimuli but a reduced response to capsaicin and hypersensitivity to formalin-induced inflammatory pain. Together, our data indicate that Prdm12 regulates pain-related behavior in a complex way by modulating gene expression in adult nociceptors and controlling their excitability. The results encourage further studies to assess the potential of Prdm12 as a target for analgesic development.

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