Six1 is a key regulator of the developmental and evolutionary architecture of sensory neurons in craniates

Abstract

BackgroundVarious senses and sensory nerve architectures of animals have evolved during adaptation to exploit diverse environments. In craniates, the trunk sensory system has evolved from simple mechanosensory neurons inside the spinal cord (intramedullary), called Rohon-Beard (RB) cells, to multimodal sensory neurons of dorsal root ganglia (DRG) outside the spinal cord (extramedullary). The fish and amphibian trunk sensory systems switch from RB cells to DRG during development, while amniotes rely exclusively on the DRG system. The mechanisms underlying the ontogenic switching and its link to phylogenetic transition remain unknown.ResultsIn Xenopus, Six1 overexpression promoted precocious apoptosis of RB cells and emergence of extramedullary sensory neurons, whereas Six1 knockdown delayed the reduction in RB cell number. Genetic ablation of Six1 and Six4 in mice led to the appearance of intramedullary sensory neuron-like cells as a result of medial migration of neural crest cells into the spinal cord and production of immature DRG neurons and fused DRG. Restoration of SIX1 expression in the neural crest-linage partially rescued the phenotype, indicating the cell autonomous requirements of SIX1 for normal extramedullary sensory neurogenesis. Mouse Six1 enhancer that mediates the expression in DRG neurons activated transcription in Xenopus RB cells earlier than endogenous six1 expression, suggesting earlier onset of mouse SIX1 expression than Xenopus during sensory development.ConclusionsThe results indicated the critical role of Six1 in transition of RB cells to DRG neurons during Xenopus development and establishment of exclusive DRG system of mice. The study provided evidence that early appearance of SIX1 expression, which correlated with mouse Six1 enhancer, is essential for the formation of DRG-dominant system in mice, suggesting that heterochronic changes in Six1 enhancer sequence play an important role in alteration of trunk sensory architecture and contribute to the evolution of the trunk sensory system.

Regulation of continuous but complex expression pattern of Six1 during early sensory development

“…A). We reported recently the mSix1‐8‐NLSCre line, in which Cre is expressed under the control of Six1‐8, a major sensory neuron‐specific enhancer of Six1 (Yajima et al, ; Sato et al, ). In the otic vesicle of mSix1‐8‐NLSCre/R26R‐LacZ embryos (Fig.…”

“…Immunofluorescence was carried out as described in detail previously (Sato et al, ; Sato et al, ; Sato et al, ). The following primary antibodies were used: rabbit anti‐ß‐Gal (dilution, 1:5,000, Covance, Berkeley, CA), chicken anti‐ß‐Gal (dilution, 1:1,000–2,000, Abcam, Cambridge, UK), mouse anti‐ISL1/2 (dilution, 1:150, mixture of hybridoma supernatants, 39.4D5 and 40.2D6, Developmental Studies Hybridoma Bank), guinea pig anti‐SIX1 (dilution, 1:5,000) (Ikeda et al, ), rat anti‐SIX1 (dilution, 1:2,000) (Konishi et al, ), guinea pig anti‐SOX10 (dilution, 1:5,000) (Yajima et al, ), mouse anti‐TUBB3 (tubulin, beta 3 class III, clone TuJ1, dilution, 1:5,000, Covance, Berkeley, CA), goat anti‐SOX2 (dilution 1:3,000, Santa Cruz Biotechnology, CA) and rabbit anti‐MYO7A (dilution 1:3,000, Proteus BioSciences, CA) antibodies. The secondary antibodies were fluorophore (Alexa Fluor 488, 546, 555, and 633)‐labeled species‐specific antibodies (dilution, 1:1,000–2,000) (Molecular Probes/Invitrogen and Amersham Biosciences).…”

“…Finally, the transgene mSix1-21-tkintron-NLSCre-ins (3.7 kb) was excised from the ASSinsBBins construct by cutting with the flanking KpnI (5 0 -end) and SacII (3 0 -end) sites, run on an agarose gel and purified using QIAEX II gel extraction kit (Qiagen). 40.2D6, Developmental Studies Hybridoma Bank), guinea pig anti-SIX1 (dilution, 1:5,000) (Ikeda et al, 2007), rat anti-SIX1 (dilution, 1:2,000) (Konishi et al, 2006), guinea pig anti-SOX10 (dilution, 1:5,000) (Yajima et al, 2014), mouse anti-TUBB3 (tubulin, beta 3 class III, clone TuJ1, dilution, 1:5,000, Covance, Berkeley, CA), goat anti-SOX2 (dilution 1:3,000, Santa Cruz Biotechnology, CA) and rabbit anti-MYO7A (dilution 1:3,000, Proteus BioSciences, CA) antibodies. The secondary antibodies were fluorophore (Alexa Fluor 488, 546, 555, and 633)-labeled speciesspecific antibodies (dilution, 1:1,000-2,000) (Molecular Probes/ Invitrogen and Amersham Biosciences).…”

Low Six4 and Six5 gene dosage improves dystrophic phenotype and prolongs life span of mdx mice

“…Previous studies investigated the roles of Six family genes in the developmental processes of various primordia and organs that belong to the peripheral and central nervous systems (Li et al 2002;Lagutin et al 2003;Zheng et al 2003;Ozaki et al 2004;Konishi et al 2006;Liu et al 2006;Ikeda et al 2007Ikeda et al , 2010Suzuki et al 2010;Yajima et al 2014), musculoskeletal system (Ozaki et al 2001;Laclef et al 2003a,b) and excretory system (Laclef et al 2003b;Xu et al 2003;Self et al 2006;Kobayashi et al 2007). In contrast, the functions of Six genes in tissue regeneration and homeostasis have rarely been examined in adult animals.…”

Tbx3 represses bmp4 expression and, with Pax6, is required and sufficient for retina formation

“…5J). Islet-1/2 is expressed in the dorsal Rohon-Beard cells and ventral motor neurons (MNs) (Diez del Corral and Storey, 2001;Olesnicky et al, 2010;Yajima et al, 2014) ( Fig. 5J; Fig.…”