Ribosomal protein haploinsufficiency results in developmental delays and variable body size without affecting lifespan in C. elegans. (A) The development of single copy large subunit ribosomal protein (RP) mutants alongside their wild-type counterparts after 96 h of incubation from embryo at 16°C. Images show ∼1 day of growth delay (left) and differences in vulval development (right), with a scale bar of 50 µm. Differential interference contrast images were taken with a 20× objective. (B) The developmental stage of animals after 72 and 96 h of incubation from embryo at 16°C were determined and plotted, with a stacked bar chart indicating the relative proportion of larvae at each stage. (C) Log₂ fold-change estimates of ribosomal protein levels in haploinsufficient RP mutants compared to stage-matched control animals are shown. Relative protein amounts were predicted by Differential Expression Proteomics (DEP package, R) based on three replicates of semi-quantitative proteomics analysis. (D) Body area measurements of stage-matched animals (at late L4 stage), normalized to the median body area of each wild type/balancer group are shown. Differences in the average normalized body area of mutant and control animals were determined using a two-tailed Welch’s two-sample t test. The central line represents the median and the bars represent interquartile distribution. (E) Brood size of each haploinsufficient RP mutant, normalized to the median of their respective wild-type controls is shown. Each dot represents the number of progeny per individual animal. Differences in the mean normalized brood size of each mutant, compared to its respective control, were analyzed using an independent two-tailed Student’s t test. Data distribution was assumed to be normal but this was not formally tested. (F) Lifespan of small subunit RP mutants (top) and large subunit RP mutants (bottom) alongside their respective wild-type controls are shown. Lifespan data are presented in a survival plot, analyzed using the Log-rank test and Bonferroni correction for multiple comparisons. Superscript numbers denote the specific wild type balancer chromosomes and are used to compare between an RP mutant and the wild-type counterpart. Balancer chromosomes are denoted as follows: +¹ = tmC20, +² = tmC5, +³ = mIn1. All experiments were performed in at least three biological replicates, and the animals were grown at 16°C. In panels C–F, RP mutants are color-coded for clarity: rpl-33(0)/+³ in green, rpl-5(0)/+³ in orange, rps-10(0)/+¹ in blue, and rps-23(0)/+² in red. The “n=” on the graphs indicates the total number of animals analyzed.