Translational regulation ensures ribosome balance despite haploinsufficiency. (A) Genes were filtered based on log₂ fold-change estimates from RNA-seq, Ribo-seq, and translation efficiency (TE), using the following criteria: (1) over or under expression at both the RNA and TE levels across all mutants, and (2) expression changes in opposite directions at the RNA or TE levels, indicating potential buffering effects. Significant gene ontology (GO) enrichment was performed for each of these categories (upper black panels: RNA and TE over, RNA under TE over, RNA over TE under, RNA and TE under). Log₂ fold-change distributions from edgeR predictions were plotted for selected significant GO categories, showing RNA-seq levels (blue), Ribo-seq levels (Ribo, cyan), and TE levels (TE, red). Significant GO annotations are provided below (X/N), where X denotes the number of significantly expressed genes and N represents the total number of genes in the GO category. Each dot in the plot represents a gene differentially expressed in the significant GO category annotated below. (B) Density plots of log₂ fold change estimates (RP mutant/control) in RNA, TE, and protein expression for all detected ribosomal protein (RP) genes (large or small subunit) in response to the single-copy loss of each RP haploinsufficient mutant, relative to stage-matched control animals generated. The density lines represent different data types: blue for RNA, red for TE and purple for protein levels. Each RP haploinsufficient mutant genotype is labeled at the top of the plots. The y-axis represents large subunit RP genes in the top row and small subunit RP genes in the bottom row. The x-axis displays the log₂ fold-change estimate distributions for RP haploinsufficient/control comparisons. ROAST multivariate gene expression analysis was conducted to calculate P values for RNA and TE level expression differences in small and large subunit RP gene expression. P values provided on the plots (blue and red values represent RNA TE level differences respectively) and indicates significant overexpression of both small and large subunit RP genes at the RNA level in all mutants, while showing significantly decreased TE levels for both subunits across all RP haploinsufficient mutants, except for rps-10(0)/+¹. Log₂ fold changes in rps-10(0)/+¹ are smaller in magnitude, likely due to the relatively modest decrease in RPS-10 protein levels (∼10% decrease, Fig. 1 C).