Fig 2: Flow Diagram of Approach and Headline ResultsThe overall analysis and main results from this study. Whole-exome sequence data from SCOOP and INTERVAL participants were analyzed as single-variant, gene-base, and gene set analyses. Overall, 48 variants from single-variant and gene-based analysis and 9 genes from gene-based analysis were identified and taken forward to stage 2 validation, by genotyping or targeted-sequenced. Combined stage 1 and stage 2 significant results were found in three genes (PHIP, DGKI, and ZMYM4), which were taken for functional follow-up. In parallel, gene set analysis performed on five primary gene sets (obesity, syndromic obesity, DDG2P, Constrained [pLI > 0.9], Unconstrained [pLI = 0.9], and Genes Mapping to known Obesity or BMI GWAS loci) is represented. The strongest enrichment, which was detected in loss-of-function constrained genes (pLI > 0.9) mapping to obesity or BMI-associated GWAS loci, is shown (Genes in GWAS loci, pLI > 0.9). Related to Figure S1.

Fig 3: Functional Characterization of Obesity-Associated Variants in PHIP(A) Mapping of human PHIP coding and splice-site variants identified in cases only (red), controls only (blue), and in cases and controls (green) on the full-length PHIP isoform. One variant was found in two unrelated probands (underlined); some variants were found in probands who also had developmental delay (*).(B) Representative confocal fluorescence microscopy images showing PHIP localization in the nucleus and the cytoplasm of COS7 cells. Blue, nuclei; green, antibody against endogenous PHIP. Scale bar, 100 µm.(C) Overexpression of WT PHIP potentiates POMC transcription in basal and leptin-stimulated conditions. n = 15, mean + SEM, two-tailed unpaired t test with Welch’s correction, *p < 0.05, **p < 0.01, ***p < 0.001.(D) Effect of human PHIP mutants on POMC transcription compared to WT in basal conditions. n = 4–6, mean + SEM, ordinary one-way ANOVA with multiple comparisons to WT, *p < 0.05, **p < 0.01, ***p < 0.001.(E) Dominant-negative effect of human PHIP mutants in POMC transcription in basal conditions. n = 3, mean + SEM, ordinary one-way ANOVA with multiple comparisons to 0 ng, *p < 0.05, **p < 0.01, ***p < 0.001.(F) Representative confocal fluorescence microscopy images showing cytoplasmic protein localization of PHIP mutants in basal conditions.(G) Nuclei to cytoplasm ratio of PHIP subcellular localization. n = 3, mean + SEM, two-tailed unpaired t test with Welch’s correction, *p < 0.05, **p < 0.01, ***p < 0.001.(H) Nuclear to cytoplasmic ratio in variants found only in cases in comparison to controls. Lines represent medians and crosses represent means. Dashed lines span from the minimum to maximum values. Wilcoxon rank-sum test, **p = 0.004. Related to Figures S4 and S5.