Figure 4. SAMD9L R1281K mutation in MLSM7 family 5.

(A) Pedigree showing the germline SAMD9L p.R1281K mutation (in red) in the healthy carrier (5M) and all 8 children. Squares indicate male family members and circles female members. Arrow indicates the proband. Unaffected individuals are indicated with open symbols, mutation carriers are denoted with black dots, individuals with a clinical history of BM Mo7 are shown with filled symbols, hatched symbol indicates CN-LOH involving chromosome 7q, carriers with dysplastic BM changes are shown with symbols filled with gray in addition to a black dot. (B) Sequencing results of individual PCR products from the amplified genomic DNA of patient 5A at the age of 15. The green boxes indicate individual missense mutations and the white boxes depict the normal sequence. Each column is an individual clone. Note that the p.D1171N mutation occur in cis with the germline p.R1281K mutation, while the p.T1053I does not. (C) Clonal evolution of BM cells from patient 5A (proband) over 18 years. Purple arrows indicate the time points at which cytogenetics (Cyto), SNP array (SNP), or SAMD9L sequencing (Seq) was performed. (D) EdU cell cycle in 293T cells transfected with vectors encoding GFP (empty vector), wild-type SAMD9L, SAMD9L p.R1281K, SAMD9L p.R1281K+p.D1171N, or SAMD9L p.T1053I. Percentages of cells in the S phase of the cell division cycle were compared using 1-way ANOVAs with repeated measures followed by Tukey’s post hoc multiple-comparisons test; significance was based on α = 0.05 (n = 3). ****P < 0.0001. n.s., not significant. Individual data points, and mean ± SD are shown. Data representative of 2 experiments completed in triplicate.