Down syndrome (DS; trisomy 21) confers a ~100-fold increased risk of Hirschsprung disease (HSCR), yet the causal contributions of specific chromosome 21 genes remain unresolved. Here we show that increased dosage of SOD1 alone is sufficient to perturb enteric nervous system (ENS) development. We engineered a humanized SOD1 trisomic mouse line by inserting a 28 kb human SOD1 locus into ROSA26 and genetically profiled the distal colon at postnatal day 0 using single-cell RNA-seq and immunofluorescence. Sod1/SOD1 was elevated ~1.5X in the ENS but varied by cell type, with transcriptionally active progenitors showing the greatest increase. Cell composition shifted toward transcriptionally active cells and glia, with concomitant loss of excitatory and inhibitory motor neurons and interneurons. Genetically, Sod1/SOD1 trisomy downregulated synaptic and neuronal communication programs but upregulated DNA replication/cell-cycle and genome maintenance pathways, especially within glia. Consequently, key HSCR genes were dysregulated: Ret, Ednrb, and Sema3c were decreased, while Sema3a ,a negative guidance cue, was increased. Ret was selectively reduced in inhibitory and excitatory motor neurons and progenitors, unchanged in glia, and reduced at the protein level in vivo. Within glia, Sod1/SOD1 was particularly elevated in proliferating/active glia with a glia-specific bias toward endogenous mouse Sod1 expression. Taken together, these data support a dual mechanism whereby increased Sod1/SOD1 dosage suppresses RET-dependent neurogenesis while independently promoting reactive/proliferative glial states. Thus, SOD1 is sufficient to alter ENS development significantly and provide the susceptibility substrate for HSCR with further reductions in RET gene expression leading to aganglionosis.
Grullon et al. (Wed,) studied this question.