Recurrent mutations in KNSTRN encoding p.Ser24Phe in cutaneous SCC. (a) The percentages of somatic point mutations in 100 primary cutaneous SCCs that are transitions compared to transversions. (b) Characterization of SCC–matched normal pairs. Mutation frequency is shown in parentheses next to each gene name. FFPE, formalin fixed and paraffin embedded. (c) Distribution of the SCC-associated alterations identified in this study across the kinastrin coding sequence. SXLP, SXLP motif; CC, coiled-coil region. (d) Detection of KNSTRN mutation encoding p.Ser24Phe in SCC precursor actinic keratoses (AKs) as well as primary SCCs by allelic discrimination quantitative PCR. NL, freshly excised normal skin. Data shown represent technical triplicates. WT, wild type.

Ser24Phe kinastrin promotes aneuploidy and enhances tumorigenesis in vivo. (a) Disrupted sister chromatid cohesion in early-passage, primary human keratinocytes pooled from multiple donors and transduced to express wild-type or Ser24Phe kinastrin. Arrowheads mark unpaired chromatids. Scale bars, 5 µm. (b) Quantification of unpaired chromatids (n = 2 biological replicates). *P = 0.0002. (c) Percentage of the genome affected by chromosomal gains and losses in SCC tissues with either wild-type or Ser24Phe mutant kinastrin. Data shown represent means ± s.d.; n = 5 independent primary tumors per group. *P = 0.007. CNVs, copy number variants. (d) Weight of mouse xenograft tumors 23 d after injection. Primary human keratinocytes were transduced to express human Cdk4 and oncogenic Ras as well as LacZ (CTR), wild-type kinastrin or Ser24Phe kinastrin and were injected subcutaneously into NOD SCID mice. Data shown represent means ± s.d.; n = 4 tumors per group. *P = 0.03, **P = 0.04; NS, not significant. (e) Ki-67 immunohistochemistry of the tumors from d. Representative fields are shown. Scale bars, 30 µm. (f) Quantification of the mitotic index for the tumors from d. Two high-power fields each containing an average of 408 cells were quantified per tumor (n = 4 tumors per group). Error bars, s.d. *P = 0.019, **P = 0.002; NS, not significant.