RNA-seq analysis of Rrp40 mutant flies identifies key neuronal targets that are altered.

(A) Principal component analysis (PCA) analysis of RNA-seq data from adult Drosophila heads from Rrp40-WT and Rrp40 mutant flies (Rrp40^G11A and Rrp40^D87A) shows that results from the three independent experiments cluster based on the genotype and that mutants are more distinct from control flies than from one another. (B,C) Volcano plots show transcripts differentially expressed in each Rrp40 mutant [(B) G11A/G11A (2,351) and (C) D87A/D87A (3,502)] compared to Rrp40-WT (q<0.05, DEseq). The number of transcripts that show ≥ 5-fold change in steady-state levels (Down or Up) are indicated to the side in each plot. Representative regulators of neuronal function in flies (Arc1 and Or82a) are highlighted. (D) A correlation curve comparing the changes in gene expression relative to wildtype for each Rrp40 allele (G11A/G11A and D87A/D87A) was produced by plotting on a logarithmic scale. This analysis shows that the changes in transcript levels are highly correlated (R = 0.85) with respect to both magnitude and direction. (E) A pie chart illustrates the class of RNAs affected in each Rrp40 mutant by showing total RNAs increased at least 5-fold in each Rrp40 mutant corresponding to coding (mRNA) and non-coding RNA (ncRNA). (F) A pie chart illustrates the classes of ncRNAs increased at least 5-fold in each Rrp40 mutant (lncRNAs, long noncoding RNAs; asRNAs, Antisense RNAs; tRNAs, transfer RNAs; snoRNAs, small nucleolar RNAs; sisRNAs, stable intronic sequence RNAs; scaRNAs, small Cajal body RNAs). (G) The enriched pathways from FlyEnrichr database for Human Disease from Flybase are shown for transcripts that are increased at least 1.2-fold in each Rrp40 mutant (left, G11A/G11A; right, D87A/D87A) as compared to Rrp40-WT. The bars shown in green correspond to significant enrichment (Adj. p-val<0.05).