Photographs of patients #7 and #29 (<i>MED13L</i> mutation).

<p>The patient (A-G) at 3 months of age (A,B,D) and at 2 years and 2 months of age (C) as well as patient 47961 (H-J) at 4 years and 2 months of age. Both patients were diagnosed with MED13L syndrome. Note long eyelashes, broad nasal tip and open mouth appearance as well as preauricular tags in both patients.</p

Sequence coverage and occurrence of <i>ADAMTSL2</i> variants.

<p>The data is shown in the UCSC genome browser “multi region view” (<a href="http://genome.ucsc.edu/" target="_blank">http://genome.ucsc.edu</a>), which displays exons in full length (dark blue boxes), flanked by 50 bp of intronic sequence (dark blue vertical line). The scale on top refers to the condensed sequence shown here. The full <i>ADAMTSL2</i> gene comprises 40.6 kb of genomic DNA (chr9:136399975–136440641, hg19). Green: read coverage, target position and variants identified in this cohort; black: corresponding data in gnomAD.</p

Photographs of patient #15 (<i>MECP2</i> mutation).

<p>The patient at 3 years and 6 months. The patient was diagnosed with a de novo mutation p.(Arg133Cys) in <i>MECP2</i>. Note arched eyebrows with slight synophrys, short anteverted nose, thin upper lip and smooth long philtrum.</p

Summary of clinical information and mutation detection rates for subgroups of the cohort.

<p>Summary of clinical information and mutation detection rates for subgroups of the cohort.</p

<i>ERCC2</i> allele frequencies (%) in BC/OC patients and corresponding control cohorts.

<p>The allele frequency is counted on the basis of sample size (in brackets) and number of observed cases (see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1006248#pgen.1006248.t001" target="_blank">Table 1</a>) with hetero- and homozygosity.</p

<i>ERCC2</i> frameshift mutation c.1703_1704delTT (p.Phe568fs) in familial breast and ovarian cancer pedigrees.

<p>Individuals with breast cancer (BC), ovarian cancer (OC) or both (BC, OC) are shown as circles filled in black. Individuals tested positive for the familial mutation are indicated in detail; those with WT (wild-type) have been tested negative. All affected individuals with BC or OC not tested for germline mutations in ERCC2 were either deceased or refused testing. (A) German, (B) Lithuanian and (C-E) Czech pedigrees.</p

Nucleotide excision repair (NER) capacity and Transcriptional activity of breast cancer associated XPD/ERCC2 variants.

<p>(A) Several XPD/ERCC2 variants cloned into an expression vector were analyzed regarding to complementation of <i>ERCC2</i>-defective XP6BE cells overexpressing the NER-deficient R601W XPD mutant [<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1006248#pgen.1006248.ref015" target="_blank">15</a>] (normalization for overexpression artifacts). Black bars indicate the mean relative repair capacity (in %, WT-XPD was set to 100%) of an UV irradiated firefly luciferase reporter gene plasmid (UVC 1000 J/m²) obtained by host cell reactivation (n>6 in triplicates). Red lines mark the range between DNA-repair levels of empty vector, i.e. residual repair activity of the cells, and WT-XPD, i.e. 100% repair capacity. (B) Dominant modulation of firefly luciferase reporter gene expression (without irradiation) via overexpression of XPD/ERCC2 BC/OC-associated variants was estimated in the transcriptionally-proficient but repair-deficient XPD/ERCC2-defective XP6BE cells. Black bars indicate the mean relative reporter gene expression (in %, empty vector control was set to 100%), obtained by CMV-promotor driven basal transcription (n>6 in triplicates). Error bars indicate the standard error of the mean. Significance levels were calculated, after pairwise testing for normal distribution of the values, using appropriate statistical tests for comparison of two groups (T-Test or U-Test, # = reference group, *** = p<0.001, ** = p<0.01, * = p<0.05, n.s. = not significant). (C) Additional characteristics of the mutations tested for repair efficiency and transcriptional activity.</p

Domain structure and modeling of the ERCC2 mutations.

<p>(A) Mutations in the XPD/ERCC2 protein domains. The diagram shows the ERCC2 protein with the four XPD domains shown as HD1 (blue), HD2 (green), FeS (Orange) and Arch (purple). The human enzyme has a C-terminal (grey) extension (CTE) that probably forms an interaction surface with the p44 protein. Disease-relevant <i>ERCC2</i> mutation sites are indicated in boxes (blue or red frame: missense or truncating mutation, respectively; fillings: light-gray, cases with breast cancer (BC); pink, case with ovarian cancer only (OC); dark-gray: cases with either breast- or ovarian cancer (BC or OC); dark-green, patients with both breast- and ovarian cancer (BC + OC)). Numbers in brackets indicate recurrent mutations. (B) Structural placement of mutations on a C-alpha trace model of human ERCC2. The residues targeted by HBOC-causing mutations are represented as space-filled red spheres. Xeroderma pigmentosum (XP) and trichothiodystrophy (TTD) disease causing mutations sites as reported in ClinVar are shown in yellow and black spheres. Missense variants at residue position 423, 461, 487, 568, 461 and 722 have been found in both BC/OC as well as XP (red-yellow spheres) and TTD (red-black spheres) patients.</p