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. Author manuscript; available in PMC: 2023 Mar 16.
Published in final edited form as: Neuron. 2022 Jan 18;110(6):992–1008.e11. doi: 10.1016/j.neuron.2021.12.019

Figure 3. Transcriptomics supports the functional importance of RefMap genes in motor neurons and in ALS.

Figure 3.

(A) Comparative gene expression analysis of RefMap genes in iPSC-derived MNs from neurologically normal individuals (n=3). For a fair comparison, we only considered those genes with evidence for MN expression (TPM>1). (B) Comparative gene expression analysis of RefMap genes in post-mortem CNS tissues from C9orf72-ALS (n=8) and sporadic ALS (n=10) patients versus neurologically normal controls (n=17). FC, frontal cortex; CB, cerebellum. (C) Comparative gene expression analysis of RefMap genes in iPSC-derived MNs from ALS patients (n=55) versus neurologically normal controls (n=15). All comparisons in A-C were performed using the one-sided Wilcoxon rank-sum test, and the Benjamini-Hochberg (BH) correction was carried out in B. In A-C, the bottom and top of the boxes indicate the first and third quartiles, respectively, where the black line in between indicates the median. Whiskers denote the minimal value within 1.5 IQR of the lower quartile and the maximum value within 1.5 IQR of the upper quartile. Red symbols denote outliers. In B and C, black dashed lines indicate the lower and upper limits of the regions with regular scale. Outliers beyond the black dashed lines are visualized with a compressed scale in the regions denoted by gray lines. (D) Heatmap showing hierarchical clustering of expression changes of RefMap genes during disease progression based on the SOD1-G93A mouse model. RefMap genes were mapped to their mouse homologs (n=510). Gene expression levels were estimated using the β scores calculated in (Maniatis et al., 2019), and were averaged across different sections of spinal cords at each time point. Time points p30, p70, p100, and p120 represent presymptomatic, onset, symptomatic, and end-stage, respectively. Difference of gene expression levels between SOD1-G93A and SOD1-WT mice at each time point was quantified by the difference in β (Δβ). Before clustering, Δβ were standardized across genes, and one minus correlation was used as the clustering distance. (E) Two distinct expression patterns (C1: 286 genes; C2: 224 genes) of RefMap genes were identified after clustering. The larger cluster C1 was progressively downregulated during ALS progression. Solid plot represents the mean of expression levels within each cluster, and the standard error is shown as shading. (F) Gene ontology analysis of C1, showing that C1 is enriched with functions related to the MN distal axon and synapse. GO, gene ontology; GOBP, gene ontology biological process; GOCC, gene ontology cellular compartment. Black vertical line represents P=0.05. See also Table S4.