Fig. 2.

Haploinsufficiency at the RPSA locus. (A) RPSA cDNA was obtained from activated T cells of patients ICA-C-I.2, ICA-C-II.3 and ICA-C-II.4. Sequences of WT and mutant cDNA are shown. The deduced frequency of each mRNA is indicated in the diagram on the right. (B) Relative levels of RPSA mRNA in activated T cells from three patients, a healthy member of kindred C (ICA-C-I.1), and four unrelated healthy controls. PBMCs were activated with PHA for 5 days. A mean of four independent experiments is shown. Error bars indicate the SEM. ***, p<0.001. (C) Immunoblot showing the levels of the WT and mutant RPSA proteins following overproduction in HEK293T cells. GAPDH, loading control; GFP, transfection contol. The blot shown is representative of 4 independent experiments. Below: intensity of the bands corresponding to the FLAG antibody normalized with respect to the band from the GFP immunoblot. Error bars indicate the SEM. (D) Genome-wide distribution of the strength of purifying selection acting in 14,993 human genes. A low f estimate (13) indicates that the gene is particularly constrained.