Supplementary Information – Postema Et Al., the Genetics of Situs Inversus Totalis Without Primary Ciliary Dyskinesia

Supplementary information – Postema et al., The genetics of situs inversus totalis without primary ciliary dyskinesia

Table of Contents: Supplementary Methods 2 Supplementary Results 5 Supplementary References 6 Supplementary Tables and Figures  Table S1. Subject characteristics 9  Table S2. Inbreeding coefficients per subject 10  Figure S1. Multidimensional scaling to capture overall genomic diversity 11 among the 30 study samples  Table S3. Significantly enriched gene-sets under a recessive mutation model 12  Table S4. Broader list of candidate genes, and the sources that led to their 13 inclusion  Table S5. Potential recessive and X-linked mutations in the unsolved cases 15  Table S6. Potential mutations in the unsolved cases, dominant model 22

1.0 Supplementary Methods

1.1 Participants Fifteen people with radiologically documented SIT, including nine without PCD and six with Kartagener syndrome, and 15 healthy controls matched for age, sex, education and handedness, were recruited from Ghent University Hospital and Middelheim Hospital Antwerp. Details about the recruitment and selection procedure have been described elsewhere (1). Briefly, among the 15 people with radiologically documented SIT, those who had symptoms reminiscent of PCD, or who were formally diagnosed with PCD according to their medical record, were categorized as having Kartagener syndrome. Those who had no reported symptoms or formal diagnosis of PCD were assigned to the non-PCD SIT group. Handedness was assessed using the Edinburgh Handedness Inventory (EHI) (2). Tables 1 and S1 give overviews of the participants and their characteristics. Note that one non-PCD SIT subject reported being forced to switch from left- to right-handedness in childhood, in which case five out of nine of the non-PCD SIT cases are naturally left-handed (Table 1, Table S1). One of the six Kartagener cases also reported forced left-to-right switching, otherwise the rest were right- handed (Table 1, Table S1). All participants gave informed consent for DNA sample collection and genomic analysis in relation to body, brain and behavioural laterality.

1.2 Whole Genome Sequencing (WGS) and Pre-processing DNA was extracted from saliva samples using the Oragene kit (Oragene). WGS was performed by Novogene (Hong Kong) using Illumina high throughput sequencing (HiSeq-PE150), creating paired end reads with a length of 150 base pairs (bp). Raw reads, stored in BAM files, were aligned to the human reference genome (the extended “decoy” version of b37) using Burrows-Wheeler Aligner (BWA) software (3), and sorted and reordered using SAMtools (v1.3.1) (4). PCR duplicates, which could arise during cluster amplification, were marked using Picard (v2.9.0). Genome Analysis Toolkit (GATK v3.7) (5) software was used to realign reads around insertions/deletions (indels) and to recalibrate base quality scores per sample.

1.3 Variant Calling and Quality Control Indels (insertion/deletions) and single nucleotide polymorphisms (SNPs) were called as recommended by the GATK Best Practices workflow. Variant Quality Score Recalibration (VQSR) was performed, and variants with a high probability of being false positive were flagged according to their sensitivity tranche (90, 99, 99.9 and 100). All SNPs and indels within a VQSR tranche of 99.9% or higher were discarded. Variants with a quality depth ≤ 9 or a call rate ≤ 0.8 were also removed. Vcftools v1.13 was used to create a summary table in the Variant Call Format (vcf). A total of 13,989,941 SNPs and indels were identified across the 30 subjects, with an average number of 5,186,055 (min= 5,053,188, max = 5,272,561) alternative alleles per subject (i.e. different to the reference genome, build37 decoy version).

2.4 Genomic-level evaluation For overall descriptive analysis of the participant genomes, a subset of 40,387 variants distributed genome-wide was used that had known Minor Allele Frequencies (MAFs) > 0.1 and were the result of pruning to be in low linkage disequilibrium with one another (for this, the flag --indep-pairwise function in Plink (v.1.90b3w) (6) was applied with a pairwise linkage disequilibrium r2 greater than 0.2, based on a SNP-SNP correlation matrix of 1500 by 150 in window size). The resulting data were then used as the basis for inferring pairwise identity by descent (IBD) sharing between subjects (i.e. genetic relatedness), inbreeding, and 3 inconsistencies with reported sex, using the Plink operations --genome, --ibc/het, and --check- sex. A different subset of 61,467 independent variants distributed genome-wide was used for visualizing population structure through Multi-Dimensional Scaling (MDS) analysis with respect to the 1000 genomes reference dataset (v37) (7) (downloaded from: ftp://climb.genomics.cn/pub/10.5524/100001_101000/100116/1kg_phase1_all.tar.gz) using the Plink operation –mds-plot.

2.5 Annotation and Filtering of SNVs SNVs were normalized using the software tool vt normalize (v0.5772-60f436c3) (8), which ensures consistent representation of variants across the genome. Normalized SNVs were annotated using Annovar (9) and Variant Effect Predictor (v88) (10). Gemini (v 0.20.0) was used to initially select protein coding variants with ‘MEDIUM’ or ‘HIGH’ impact severity annotations, as well as non-coding variants with ‘HIGH’ impact severity annotations (in practice those altering splice donor or acceptor sites). Additional filtering was done in R and comprised the removal of synonymous variants, and of ‘MEDIUM’ variants with a PolyPhen (11) or Sift (12) prediction score of "benign" or "tolerant" respectively. Data were then processed and analyzed separately under recessive and dominant models: Recessive model: For the recessive model we further excluded variants with a known MAF > 0.005 in any of the following databases: GNOMAD (13), ESP (14), 1000 Genomes (7) and ExAC databases (13). The remaining variants were considered as putative mutations. Gene- level mutation counts per subject were then made, with a given gene being assigned as recessively mutated when it carried two copies of the same mutation (homozygous) or two different mutations (possible compound heterozygous). Integrative Genome Viewer (IGV v2.3.97) (15) was used to visualize the possible compound heterozygous mutations, and genes carrying these were discarded when both mutations were definitely present on the same allele (i.e. "in phase") on a given sequence read. Finally, genes recessively mutated according to these criteria in any of the fifteen healthy control subjects were excluded as being potentially causative in cases, and also removed for the purposes of Gene Set Enrichment Analysis (GSEA; see below): this step had the advantage of removing many false variants from potential technical artifacts, or variants which are common in the population but not annotated as such in on-line databases. Genes on chromosome X were processed as part of the recessive pipeline, such that females would need to carry two mutations in a given gene, and males one mutation. Dominant model: An upper MAF threshold of 5.0E-5 was applied in this case, and genes carrying at least one remaining variant according to this criterion were considered as potentially causative. Again, genes mutated according to this criterion in any of the fifteen healthy control subjects were excluded as being potentially causative in cases, and removed for the purposes of GSEA analysis (below).

2.6 Gene Set Enrichment Analysis To test whether a given list of mutated genes in a certain set of subjects (e.g. those recessively mutated in Kartagener subjects) contained functionally related genes across subjects, we performed Gene Set Enrichment Analysis (GSEA) using the g:Profiler R package (version 0.6.1) (16). The following settings were used: minimum set size = 15, maximum set size = 500, minimum intersect number = 2, hierarchical filtering = moderate. Gene classification schemes derived from Gene Ontology (GO) (17, 18), Kyoto Encyclopedia of Genes and Genomes (KEGG) (19) and the Human Phenotype (HP) (20) databases were considered. Since g:Profiler performs GSEA for each database separately, the number of genes in the default background, as well as the number of gene-sets tested, differed per analysis. That is to say, 6380 gene-sets were tested against a background of 19327 genes in the GO analysis, while the KEGG analysis 4 comprised 301 gene-sets and 7214 background genes, and the HP analysis 2541 gene-sets and 3682 background genes. P-values were corrected for testing of multiple gene-sets based on the gSCS correction method in g:Profiler, separately for each input gene list and database. This method of multiple testing correction was specifically designed for pathway enrichment analyses, as it takes into account the hierarchical structure of the pathways, and does not treat them independently of each other (in contrast to Bonferroni or Benjamini-Hochberg False Discovery Rate (FDR) corrections) (16). Mutations on chromosome X were combined with autosomal recessive mutations for the purposes of GSEA. GSEAs were performed for recessively mutated genes in Kartagener cases (n = 54 input genes), non-PCD SIT cases (n = 60 input genes), left handed non-PCD SIT cases (n = 42 input genes), and cases that were unsolved under the recessive model (n = 42 input genes) (See table 1 and main text for the numbers of subjects involved). For the dominant model, GSEA was only performed for mutated genes in cases that remained unsolved under the recessive model (n = 217 input genes, Table S6). In order to confirm that healthy controls showed no significant pathway enrichment among their mutated genes, a mirrored exclusion was performed whereby any genes mutated in cases were excluded from the control lists. This resulted in 56 genes exclusively mutated in controls for the recessive model, and 533 genes exclusively mutated in controls under the dominant model.

1.8 Candidate Gene Lists The Clinvar database (21) was used as the initial source for identifying genes already known to cause SIT, PCD, Kartagener, situs ambiguus (SA), or Heterotaxy (HTX) in humans. A broader candidate gene list was also created which included genes from the literature that were suggested to be associated with human laterality phenotypes and/or ciliopathies, but not (yet) recorded in the Clinvar database as such (see main text; the complete list is in Table S4). Specifically, a list of ciliopathy genes from a review of this topic (22) was searched for the terms: “PCD”, “heterotaxy”, “situs”, “left-right”, “asymmetry” or “laterality”, yielding 18 additional candidate genes. Additionally, a list of genes potentially associated with human laterality disorders, as compiled in a 2015 review (23), resulted in the addition of 25 candidate genes. Six additional genes were included, of which three were reported as potentially associated with PCD or heterotaxy (24), and three with potential associations with non- syndromic heterotaxy (25). Finally, 39 more genes – associated with the ‘situs inversus’ phenotype - were retrieved from the Mouse Genome Database (MGD) (26) at the Mouse Genome Informatics website, the Jackson laboratory, Bar Harbor, Maine. (URL: http://www.informatics.jax.org) [Oct, 2017].

1.9 Structural Variants Structural Variants (SVs) (>50 kilobases in length) were called using a combination of two SV calling algorithms: CNVnator (v0.3.3) (27) and Lumpy (v 0.2.13) (28). These algorithms complement each other regarding the kinds of signals in WGS data that they can detect, as CNVnator is based on read-depth, whereas Lumpy is based on paired-end mapping (29). For CNVnator the bin size was set to 100 base pairs for all samples except for two, where it was 150 base pairs (we determined a roughly optimal bin size for each subject’s genome, such that that the average read depth for that genome would be at least 4 standard deviations from zero). Lumpy was run using default parameters in “lumpyexpress”. SVs were then annotated using SV2 (30). Variants that were present in any of the 15 healthy controls were removed from consideration. Only variants that passed the default SV2 filtering criteria for quality were included (30).

2.0 Supplementary Results

2.1 Additional observations – recessive mutations in unsolved cases (see Table S5) We note possible compound heterozygous mutations in PKD1 [MIM:601313], as a paralogue of PKD1L1 [MIM:609721], in subject SI14. However, recessive mutations would be expected to cause Polycystic Kidney Disease (31, 32), and since there was no such diagnostic record for this subject, one or both of these specific mutations probably has limited functional impact and is therefore unlikely to be a monogenic cause for SIT either. The gene RP1L1 [MIM:608581], which is a member of the gene set GO:005930 ‘Axoneme’ (which was a significant set in GSEA of recessively mutated genes in Kartagener subjects, see Table S3), was putatively recessively mutated in subject SI03. However, mutations in this gene are associated with occult macular dystrophy (33), and again no such diagnosis had been made, so that this also appears an unlikely causal gene for non-PCD SIT in this subject. KIF13B [MIM:607350] was putatively recessively mutated in subjects SI14 and SI12, and is a member of the gene sets GO:007018 (Microtubule-based movement) and GO:0003777 (Microtubule motor activity), which again had shown significant results in GSEA of the Kartagener subjects (Table S3). Although no literature has linked KIF13B to PCD or laterality phenotypes, KIF3A [MIM:604683], another kinesin encoding gene, is known to be involved in LR determination (34). Together with dyneins, kinesins allow ciliary proteins to enter the organelle via the transition zone by transporting them as cargo (35, 36), and accordingly play an important role in ciliary construction and maintenance (35, 36). The mutations in KIF13B might therefore potentially cause SIT without PCD in subject SI14, and perhaps also affect the phenotypic outcome in subject SI12 who has likely causal mutations in DNAH5, but we cannot confidently assign a role to KIF13B on the basis of this evidence.

2.2. Additional observations – dominant model in unsolved cases (see Table S6) We furthermore note a heterozygous mutation in WDR62 [MIM:613583] in the unsolved case SI09. Although mice with mutations in this gene demonstrated dextrocardia and right pulmonary isomerim (MGI:5437081) (26), humans with recessive WDR62 mutations do not show altered laterality. Instead, they have shown infantile spasm, microcephaly and intellectual disability (37). It is therefore unlikely that WDR62 is a dominant cause of altered laterality in humans. The same subject (i.e. SI09) also showed a mutation in PLXND1 [MIM:604282]. This gene is involved in cardiovascular development (38), and recessive mutations in this gene may lead to congenital heart defects in humans (39), but dominant mutations have not been reported in humans. Since there is no clear evidence for a role of PLNXD1 in laterality – including not in mice– it is improbable that this gene is involved in non-PCD SIT. The gene CEP290 [MIM:610142], mutated in subject SI14, is linked to left sided isomerism in mice (MGI:5438068)(26). In humans, recessive mutations have been linked to a variety of ciliopathies, ranging from nephronophthisis, retinal degeneration and Joubert syndrome, to Bardet-Biedl syndrome and Meckel-Grüber syndrome (40, 41). However, similar to the aforementioned genes, mutations in CEP290 have not been associated with laterality phenotypes in humans. A heterozygous mutation in LRRC6 [MIM:614930] was observed in subject SI05. Since recessive – but not dominant - mutations in this gene have been associated with PCD (42), it cannot be concluded that this mutation is causal for non-PCD SIT in this subject.

2.3 Structural variants SIT cases had SVs affecting an average of 9.7 genes per subject (min = 2, max = 16), and controls had SVs affecting an average of 9.6 genes per subject (min = 5, max = 16). None of 6 the cases showed SVs in genes that were annotated SIT, PCD, Kartagener, situs ambiguus (SA), or Heterotaxy (HTX) in Clinvar, nor in genes from the broader list of candidate genes (Table S4).

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Supplementary Tables and Figures

Table S1. Subject characteristics

Sample Type Sex Age Educ EHI Hand PCD CHD SI02 non-PCD SIT Male 50 8 0.9 R 0 0 SI03 non-PCD SIT Female 26 12 -0.8 L 0 0 SI04 non-PCD SIT Male 23 13 -1 L 0 1 SI05 non-PCD SIT Male 27 12 0.9 R 0 0 SI07 non-PCD SIT Female 35 12 0.9 R 0 1 SI09 non-PCD SIT Female 36 15 0.7 L* 0 0 SI12 non-PCD SIT Female 40 12 0.9 R 0 0 SI14 non-PCD SIT Male 18 12 -0.8 L 0 1 SI16 non-PCD SIT Male 21 14 -1 L 0 0 SI06 Kartagener Male 46 12 1 R 1 0 SI08 Kartagener Female 23 15 0.9 R 1 0 SI11 Kartagener Female 32 11 0.9 R 1 0 SI13 Kartagener Male 48 14 0.6 L* 1 0 SI15 Kartagener Female 31 18 0.7 R 1 0 SI17 Kartagener Male 39 14 0.5 R 1 0 CO02 Healthy control Male 51 10 1 R CO03bis Healthy control Female 26 15 -1 L CO04 Healthy control Male 22 13 -0.5 L CO15 Healthy control Male 27 12 1 R CO06bis Healthy control Male 43 12 1 R CO07 Healthy control Female 33 12 1 R CO08 Healthy control Female 22 14 1 R CO09bis Healthy control Female 38 15 0.6 R CO11 Healthy control Female 34 12 1 R CO12 Healthy control Female 38 12 1 R CO13bis Healthy control Male 46 12 1 R CO14 Healthy control Male 19 12 -1 L CO15 Healthy control Female 35 16 1 R CO16bis Healthy control Male 20 14 -0.7 L CO17 Healthy control Male 41 12 1 R Educ: years of formal eduction; EHI: Edinburgh Handedness Inventory score; Hand: natural handedness; CHD: Congenital Heart Disease. *Self-identified natural lefthander made to convert to right-handedness.

Table S2. Inbreeding coefficients per subject.

Subject Type O(HOM) E(HOM) NOMISS Fhat1 Fhat2 Fhat3 SI02 non-PCD SIT 21098 21680 40351 -0.034 -0.034 -0.034 SI03 non-PCD SIT 20997 21680 40352 -0.035 -0.038 -0.036 SI04 non-PCD SIT 21099 21680 40350 -0.027 -0.035 -0.031 SI05 non-PCD SIT 21161 21680 40337 -0.026 -0.030 -0.028 SI07 non-PCD SIT 20935 21690 40354 -0.047 -0.039 -0.043 SI09 non-PCD SIT 21090 21690 40363 -0.037 -0.033 -0.035 SI12 non-PCD SIT 21274 21690 40359 -0.032 -0.023 -0.028 SI14 non-PCD SIT 20952 21690 40359 -0.041 -0.040 -0.040 SI16 non-PCD SIT 21282 21680 40352 -0.018 -0.023 -0.021 SI06 Kartagener 22503 21680 40348 0.048 0.041 0.044 SI08 Kartagener 21039 21690 40357 -0.033 -0.035 -0.034 SI11 Kartagener 21059 21680 40346 -0.036 -0.034 -0.035 SI13 Kartagener 21011 21690 40360 -0.036 -0.036 -0.036 SI15 Kartagener 21000 21690 40355 -0.032 -0.038 -0.035 SI17 Kartagener 20888 21690 40363 -0.042 -0.047 -0.045 CO02 Healthy control 21221 21690 40358 -0.031 -0.025 -0.028 CO03bis Healthy control 21028 21680 40341 -0.036 -0.036 -0.036 CO04 Healthy control 21192 21680 40351 -0.026 -0.030 -0.028 CO15 Healthy control 21177 21680 40348 -0.037 -0.025 -0.031 CO06bis Healthy control 21151 21680 40346 -0.035 -0.029 -0.032 CO07 Healthy control 20989 21680 40349 -0.038 -0.038 -0.038 CO08 Healthy control 21128 21680 40354 -0.034 -0.030 -0.032 CO09bis Healthy control 20923 21680 40350 -0.039 -0.045 -0.042 CO11 Healthy control 21095 21680 40344 -0.038 -0.029 -0.033 CO12 Healthy control 21095 21690 40356 -0.031 -0.034 -0.032 CO13bis Healthy control 21067 21690 40358 -0.039 -0.033 -0.036 CO14 Healthy control 20983 21690 40362 -0.041 -0.039 -0.040 CO15 Healthy control 20965 21690 40360 -0.036 -0.042 -0.039 CO16bis Healthy control 21007 21690 40356 -0.038 -0.038 -0.038 CO17 Healthy control 21170 21680 40345 -0.023 -0.031 -0.027 Subject SI06 (bold font) is the only subject with positive inbreeding coefficients, indicating an excess of homozygosity. O(HOM): observed number of homozygotes; E(HOM): expected number of homozygotes; NOMISS: Number of non-missing genotype calls; Fhat1: variance-standardized relationship minus 1; Fhat2: Excess homozygosity-based inbreeding estimate; Fhat3: estimate based on correlation between uniting gametes.

Figure S1. Multidimensional Scaling (MDS) to capture overall genomic diversity among the 30 study samples (black dots), in relation to the 1000 Genomes populations of known geographic ancestries. Subject SI06 is clearly distinct from the European-descent population (yellow dots).

Table S3. Significantly enriched gene-sets under a recessive mutation model

Group p-value1 ter query overlap term ID term name intersection samples Pval OR CI low CI m size size Fisher2 up size Kartagener 6.27E-05 258 41 8 GO:0007018 microtubule-based DNAH5, CCDC114, SI06, SI08, SI11, 2.94E-15 Inf 143 Inf (n=6) movement DNAH11, WDR60, SI12, SI13, SI15, LRRC6, DNAAF1, SI17 FMN2, DNAH12 Kartagener 4.32E-05 17 41 4 GO:0036158 outer dynein arm DNAH5, CCDC114, SI06, SI11, SI12, 7.39E-13 Inf 1007 Inf (n=6) assembly LRRC6, DNAAF1 SI13, SI15, SI17 Kartagener 7.00E-05 50 41 5 GO:0030286 dynein complex DNAH5, CCDC114, SI08, SI11, SI12, 1.97E-13 Inf 434 Inf (n=6) DNAH11, WDR60, SI13, SI15, SI17 DNAH12 Kartagener 3.07E-03 106 41 5 GO:0005930 axoneme DNAH5, CCDC114, SI03, SI08, SI11, 9.43E-12 Inf 196 Inf (n=6) DNAH11, DNAAF1, SI12, SI13, SI15, RP1L1 SI17 Kartagener 2.10E-02 25 41 3 GO:0045503 dynein light chain DNAH5, DNAH11, SI08, SI11, SI12, 2.98E-09 Inf 408 Inf (n=6) binding WDR60 SI15, SI17 Kartagener 3.31E-02 85 41 4 GO:0003777 microtubule motor DNAH5, DNAH11, SI08, SI11, SI12, 6.29E-10 Inf 179 Inf (n=6) activity WDR60, DNAH12 SI15, SI17 Kartagener 2.18E-02 15 18 3 HP:0005938 Abnormal CCDC114, LRRC6, SI06, SI11, SI13 5.89E-10 Inf 702 Inf (n=6) respiratory motile DNAAF1 cilium morphology Kartagener 1.01E-02 74 18 5 HP:0001696 Situs inversus DNAH5, CCDC114, SI06, SI08, SI11, 1.50E-12 Inf 283 Inf (n=6) totalis DNAH11, LRRC6, SI12, SI13, SI15, DNAAF1 SI17 Kartagener 1.21E-04 31 18 5 HP:0012261 Abnormal DNAH5, CCDC114, SI06, SI08, SI11, 1.58E-14 Inf 715 Inf (n=6) respiratory motile DNAH11, LRRC6, SI12, SI13, SI15, cilium physiology DNAAF1 SI17 Non-PCD SIT No significant gene sets (n=9) Non-PCD SIT No significant gene sets LH (n=5) Non-PCD SIT No significant gene sets unsolved (n=6) Healthy No significant gene sets Controls (n=15) 1 P-values are corrected for multiple testing using the gSCS method. 2 Uncorrected P-values were calculated using Fisher’s exact test. LH: left-handed.

Table S4. Broader list of candidate genes, and the sources that led to their inclusion.

Gene Location Ensembl Gene ID Literature AK7 14q32.2 ENSG00000140057 Reiter & Leroux (2017) AMN 14q32 ENSG00000166126 Mouse Genome Database (2018) ANKS3 16p13.3 ENSG00000168096 Reiter & Leroux (2017) ANKS6 9q22.33 ENSG00000165138 Reiter & Leroux (2017) AP1B1 22q12 ENSG00000100280 Mouse Genome Database (2018) ATMIN 16q23.2 ENSG00000166454 Mouse Genome Database (2018) BBS1 11q13 ENSG00000174483 Mouse Genome Database (2018) BBS2 16q21 ENSG00000125124 Deng, Xia, & Deng (2015) BBS4 15q22.3-q23 ENSG00000140463 Mouse Genome Database (2018) BICC1 10q21.2 ENSG00000122870 Mouse Genome Database (2018) CC2D2A 4p15.3 ENSG00000048342 Mouse Genome Database (2018) CEP290 12q21.3 ENSG00000198707 Mouse Genome Database (2018) CFAP54 Reiter & Leroux (2017) CITED2 6q23.3 ENSG00000164442 Deng, Xia, & Deng (2015) DAND5 19p13.2-p13.13 ENSG00000179284 Mouse Genome Database (2018) DCTN5 16p12.2 ENSG00000166847 Mouse Genome Database (2018) DNAH6 2p11.2 ENSG00000115423 Reiter & Leroux (2017) DNAH9 17p12 ENSG00000007174 Reiter & Leroux (2017) DPCD 10q24.32 ENSG00000166171 Reiter & Leroux (2017) DRC3 Reiter & Leroux (2017) EPB41L5 2q14 ENSG00000115109 Deng, Xia, & Deng (2015) FGF10 5p13-p12 ENSG00000070193 Mouse Genome Database (2018) FOXA2 20p11 ENSG00000125798 Peeters & Devriendt (2006) FOXH1 8q24.3 ENSG00000160973 Deng, Xia, & Deng (2015) FOXI2 10q26.2 ENSG00000186766 Hagen et al. (2016) FOXJ1 17q22-q25 ENSG00000129654 Mouse Genome Database (2018) FOXL2 3q23 ENSG00000183770 Mouse Genome Database (2018) GALNT11 7q36.1 ENSG00000178234 Reiter & Leroux (2017) GDF1 19p12 ENSG00000223802 Deng, Xia, & Deng (2015) GJA1 5q22.31 ENSG00000152661 Hagen et al. (2016) HES7 17p13.2 ENSG00000179111 Deng, Xia, & Deng (2015) IFT122 3q21 ENSG00000163913 Mouse Genome Database (2018) IFT140 16p13.3 ENSG00000187535 Mouse Genome Database (2018) IFT27 22q12.3 ENSG00000100360 Mouse Genome Database (2018) IFT74 9p21.2 ENSG00000096872 Mouse Genome Database (2018) IFT88 13q12.1 ENSG00000032742 Reiter & Leroux (2017) IHH 2q33-q35 ENSG00000163501 Mouse Genome Database (2018) INVS 9q31 ENSG00000119509 Reiter & Leroux (2017) KIF3A 5q31 ENSG00000131437 Reiter & Leroux (2017) LEFTY1 1q42.1 ENSG00000243709 Mouse Genome Database (2018) LETYA Peeters & Devriendt (2006) LZTFL1 3p21.3 ENSG00000163818 Deng, Xia, & Deng (2015) MBD4 3q21-q22 ENSG00000129071 Mouse Genome Database (2018) MCIDAS 5q11.2 ENSG00000234602 Reiter & Leroux (2017)

Table S4 [Continued]. Broader list of candidate genes, and the sources that led to their inclusion.

Gene Location Ensembl Gene ID Literature MED13L 12q24 ENSG00000123066 Deng, Xia, & Deng (2015) MEGF8 19q12 ENSG00000105429 Deng, Xia, & Deng (2015) MGAT1 5q35 ENSG00000131446 Mouse Genome Database (2018) MGRN1 16p13.3 ENSG00000102858 Mouse Genome Database (2018) MKS1 17q23 ENSG00000011143 Mouse Genome Database (2018) MNS1 15q21.3 ENSG00000138587 Reiter & Leroux (2017) NEK2 1q32.2-q41 ENSG00000117650 Deng, Xia, & Deng (2015) NEK8 17q11.1 ENSG00000160602 Deng, Xia, & Deng (2015) NKX2.5 Peeters & Devriendt (2006) NKX2-5 5q34 ENSG00000183072 Deng, Xia, & Deng (2015) NKX6-2 10q26 ENSG00000148826 Hagen et al. (2016) NME7 1q24 ENSG00000143156 Mouse Genome Database (2018) NOTCH1 9q34.3 ENSG00000148400 Deng, Xia, & Deng (2015) NOTCH2 1p13-p11 ENSG00000134250 Deng, Xia, & Deng (2015) NOTO Reiter & Leroux (2017) NPHP3 3q22 ENSG00000113971 Deng, Xia, & Deng (2015) NPHP4 1p36 ENSG00000131697 Deng, Xia, & Deng (2015) NUP188 9q34.11 ENSG00000095319 Deng, Xia, & Deng (2015) OFD1 Xp22.3-p22.2 ENSG00000046651 Reiter & Leroux (2017) PAX8 2q12-q14 ENSG00000125618 Mouse Genome Database (2018) PCSK5 9q21.3 ENSG00000099139 Mouse Genome Database (2018) PCSK6 15q26 ENSG00000140479 Mouse Genome Database (2018) PITX2 4q25-q26 ENSG00000164093 Deng, Xia, & Deng (2015) PKD2 4q21-q23 ENSG00000118762 Deng, Xia, & Deng (2015) PLXND1 3q22 ENSG00000004399 Mouse Genome Database (2018) POLB 8p11.2 ENSG00000070501 Mouse Genome Database (2018) PSKH1 16q22.1 ENSG00000159792 Mouse Genome Database (2018) RFX3 9p24.2 ENSG00000080298 Reiter & Leroux (2017) ROCK2 2p24 ENSG00000134318 Deng, Xia, & Deng (2015) RPGR Xp11.4 ENSG00000156313 Reiter & Leroux (2017) RPGRIP1L 16q12.2 ENSG00000103494 Mouse Genome Database (2018) SESN1 6q21 ENSG00000080546 Deng, Xia, & Deng (2015) SHH 7q36 ENSG00000164690 Mouse Genome Database (2018) SHROOM3 4q21.1 ENSG00000138771 Deng, Xia, & Deng (2015) SLIT2 4p15.2 ENSG00000145147 Mouse Genome Database (2018) SMAD2 18q21 ENSG00000175387 Deng, Xia, & Deng (2015) T 6q27 ENSG00000164458 Mouse Genome Database (2018) TBC1D32 6q22.31 ENSG00000146350 Mouse Genome Database (2018) TGFBR2 3p22 ENSG00000163513 Deng, Xia, & Deng (2015) TGIF1 18p11.3 ENSG00000177426 Mouse Genome Database (2018) TMEM67 8q21.13-q22.1 ENSG00000164953 Mouse Genome Database (2018) TRAPPC10 21q22.3 ENSG00000160218 Mouse Genome Database (2018) UVRAG 11q13 ENSG00000198382 Deng, Xia, & Deng (2015) WDR62 19q13.12 ENSG00000075702 Mouse Genome Database (2018)

Table S5. Potential recessive and X-linked mutations in the unsolved cases (note that genes recessively mutated in any of the 15 healthy control subjects are not included in this list).

impact Polyphen/ model hom samples het samples gene location rs ID start end ref alt MAF impact severity Clinvar Sift hom SI04 not applicable AC092170.1 NA None 117938751 117938754 CTT C -1 splice acceptor HIGH None NA/ NA variant hom SI04 not applicable AC092170.1 NA None 117938751 117938754 CTT C -1 splice acceptor HIGH None NA/ NA variant chet not applicable SI03 AC093323.1 NA rs180678565 6693854 6693855 T C 0.0014 missense variant MED None unknown/ deleterious low confidence chet not applicable SI03 AC093323.1 NA rs201074425, 6693879 6693880 A ATG 0 frameshift variant HIGH None NA/ NA rs746556174 chet not applicable SI07, SI09, AL390778.1 NA rs760375607 138150834 138150841 ACGGC A 0 inframe deletion MED None NA/ NA SI12, CO03bis, GG CO07 chet not applicable SI02, SI07, AL390778.1 NA None 138151168 138151169 T TGACGGCGG -1 frameshift variant HIGH None NA/ NA SI14, SI16, TGATGCCGA CO11 CCGTATGGG GTGACGGTG GTGATGCCG ACCGTATGG GGC hom SI03 not applicable AL591806.1 NA rs774404701, 161035657 161035658 C CTTAT 0 frameshift variant HIGH None NA/ NA rs560515090 hom SI03 not applicable AL591806.1 NA rs774404701, 161035657 161035658 C CTTAT 0 frameshift variant HIGH None NA/ NA rs560515090 chet not applicable SI09, SI14 AP003041.2 NA rs397848292 129936974 129936976 CT C -1 frameshift variant HIGH None NA/ NA chet not applicable SI09, SI14 AP003041.2 NA rs552607281 129936974 129936984 CTTTTT C 0 splice donor HIGH None NA/ NA TTTT variant chet not applicable SI14 ASB10 7q36.1 rs61735130 150878219 150878220 G A 0.0037 missense variant MED glaucoma possibly 27 damaging/ deleterious chet not applicable SI14 ASB10 7q36.1 rs104886481 150878143 150878144 G A 0.0015 missense variant MED glaucoma probably 2754 damaging/ deleterious chet not applicable SI05 C2CD5 NA rs192026857 22650521 22650522 A T 0.002 missense variant MED None NA/ NA chet not applicable SI05 C2CD5 NA rs768626962 22622730 22622731 T C 0.0003 splice acceptor HIGH None NA/ NA 8586 variant hom SI16 not applicable CCNJ NA rs200852141 97819345 97819347 GT G -1 splice donor HIGH None NA/ NA variant hom SI05 not applicable COL4A6 Xq22.3 rs756582576 107433663 107433664 C T 0.0005 missense variant MED alport possibly 7128 syndrome | damaging/ deafness NA

Table S5 [Continued]. Potential recessive and X-linked mutations in the unsolved cases.

impact Polyphen/ model hom samples het samples gene location rs ID start end ref alt MAF impact severity Clinvar Sift hom SI05 not applicable COL4A6 Xq22.3 rs750814219 107422569 107422570 C T 0.0006 missense variant MED alport probably 3994 syndrome | damaging/ deafness NA chet not applicable SI08, SI14, FAM120B 6q26-q27 None 170627694 170627695 A ACCCTGAAC -1 inframe insertion MED None NA/ NA CO08, CCAGGCAAG CO09bis, CO14 AAGTTCCCA TGTGTACAG G hom SI14 not applicable FAM9A Xp22.33- rs199608514 8763339 8763340 C T 0.0002 missense variant MED None unknown/ p22.32 3997 tolerated low confidence hom SI14 not applicable FOXR2 Xp11.21 None 55650996 55650997 C T -1 missense variant MED None possibly damaging/ deleterious hom SI14 not applicable GOLGA6L6 NA rs754572511 20740290 20740296 CGTGA C 0.0011 frameshift variant HIGH None NA/ NA A 5562 hom SI14 not applicable GOLGA6L6 NA rs754572511 20740290 20740296 CGTGA C 0.0011 frameshift variant HIGH None NA/ NA A 5562 hom SI11 not applicable HLA-DRB5 6p21.3 rs138885982 32487157 32487158 G GCA -1 frameshift variant HIGH None NA/ NA hom SI11 not applicable HLA-DRB5 6p21.3 None 32487214 32487215 C CT -1 frameshift variant HIGH None NA/ NA hom SI11 not applicable HLA-DRB5 6p21.3 rs41549318 32487210 32487212 AC A -1 frameshift variant HIGH None NA/ NA hom SI05 not applicable IGHV3-42 NA rs73372475 106919381 106919382 T C -1 stop lost HIGH None NA/ NA hom SI05 not applicable IGHV3-42 NA rs73372475 106919381 106919382 T C -1 stop lost HIGH None NA/ NA chet not applicable SI12, SI14 KIF13B 8p12 None 28974427 28974428 C A -1 missense variant MED None possibly damaging/ deleterious chet not applicable SI12, SI14 KIF13B 8p12 rs753108980 28974427 28974428 C T 0.0001 missense variant MED None probably 8827 damaging/ deleterious hom SI08 not applicable KIR3DL1 19q13.4 rs652641 55331286 55331287 G T 0 missense variant MED None possibly damaging/ deleterious hom SI08 not applicable KIR3DL1 19q13.4 rs643861 55330035 55330036 G A 0.0003 missense variant MED None benign/ 8485 deleterious hom SI05, SI12, not applicable KRTAP5-1 11p15.5 None 1605903 1605904 T TCCCCCACA 0.0045 inframe insertion MED None NA/ NA SI16 AGAACCGCA 2282 GCCCCCCTT GCAGCCTCC ACAGGAGCC ACAGCCCCC CTTGGAG

Table S5 [Continued]. Potential recessive and X-linked mutations in the unsolved cases.

locat impact Polyphen/ model hom samples het samples gene ion rs ID start end ref alt MAF impact severity Clinvar Sift chet not applicable SI09 LOC100132874 NA None 45120089 45120150 GGCCTCTCCAGC G -1 inframe deletion MED None NA/ NA GTGACCCTGTCT GCATGGGGCCTT TCCAGCGTGACC CTGTCTGCCTGT T chet not applicable SI08, SI09, LOC100132874 NA None 45120094 45120125 CTCCAGCGTGAC C -1 inframe deletion MED None NA/ NA CO02, CO08, CCTGTCTGCATG CO12, CO13bis GGGCCTT hom SI14 not applicable LOC100506248 NA rs12831542 12264204 12264205 A G -1 missense variant MED None NA/ NA hom SI14 not applicable LOC100506248 NA rs12831542 12264204 12264205 A G -1 missense variant MED None NA/ NA hom SI09, SI13 not applicable LOC100653133 NA rs202162063 32787186 32787187 T C -1 missense variant MED None NA/ NA hom SI08 not applicable LOC100996276 NA rs113311249 101327887 101327888 A C -1 missense variant MED None NA/ NA hom SI04 not applicable LOC100996350 NA rs62248006 75719341 75719342 C T -1 missense variant MED None NA/ NA hom SI04 not applicable LOC100996350 NA rs62248006 75719341 75719342 C T -1 missense variant MED None NA/ NA hom SI15 not applicable LOC101926998 NA None 77645488 77645519 CCTTCCTTTCTTT C -1 splice donor HIGH None NA/ NA CTTTTTCTTTCTT variant TCTTT hom SI12 not applicable LOC101926998 NA None 77645488 77645515 CCTTCCTTTCTTT C -1 splice donor HIGH None NA/ NA CTTTTTCTTTCTT variant T hom SI14 not applicable LOC101926998 NA None 77645489 77645504 CTTCCTTTCTTTC C -1 splice donor HIGH None NA/ NA TT variant hom SI14 not applicable LOC101926998 NA None 77645489 77645504 CTTCCTTTCTTTC C -1 splice donor HIGH None NA/ NA TT variant hom SI14 not applicable LOC101926998 NA None 77645517 77645524 TTCTTTC T -1 inframe deletion MED None NA/ NA hom SI02 not applicable LOC101928723 NA rs745758 6164679 6164680 C G -1 missense variant MED None NA/ NA chet not applicable SI07, SI14, LOC101928884 NA rs748700261 1328429 1328459 ACTCCTCTACGG A -1 frameshift variant HIGH None NA/ NA SI17, CO06bis, ACACATTTCATC CO11, CO12, CAGCTC CO15, CO16bis, CO17 chet not applicable SI07, SI09 LOC101928884 NA rs868671387 1328564 1328565 T C -1 missense variant MED None NA/ NA chet not applicable SI09 LOC101928884 NA rs866503911 1327424 1327425 A G -1 missense variant MED None NA/ NA

Table S5 [Continued]. Potential recessive and X-linked mutations in the unsolved cases.

impact Polyphen/ model hom samples het samples gene location rs ID start end ref alt MAF impact severity Clinvar Sift chet not applicable SI07, SI14, LOC101928884 NA rs748700261 1328429 1328459 ACTCCTCT A -1 frameshift variant HIGH None NA/ NA SI17, CO06bis, ACGGACAC CO11, CO12, ATTTCATC CO15, CAGCTC CO16bis, CO17 hom SI04 not applicable LOC101928917 NA None 148630211 148630212 G T -1 missense variant MED None NA/ NA chet not applicable SI09 LOC101929113 NA None 4848346 4848347 C CTTC -1 frameshift variant HIGH None NA/ NA CGAC GCCA CTGG CCCC G chet not applicable SI09 LOC101929113 NA rs528814344 4848369 4848370 C A -1 missense variant MED None NA/ NA hom SI14 not applicable LOC338797 NA None 131851942 131851998 TGCGGGGC T -1 splice acceptor HIGH None NA/ NA ACTCCACA variant GAAGCTGG GTCTGGAG ATGAGGGT CCCCGTGA GGCCATGC hom SI14 not applicable LOC338797 NA None 131851942 131851998 TGCGGGGC T -1 splice acceptor HIGH None NA/ NA ACTCCACA variant GAAGCTGG GTCTGGAG ATGAGGGT CCCCGTGA GGCCATGC hom SI14 not applicable MAP3K15 Xp22.12 rs771700881 19506958 19506959 G C 2.59E- missense variant MED pyruvate possibly 05 dehydrogenas damaging/ e e1-alpha deleterious deficiency chet not applicable SI05 MUC16 19p13.2 rs374715126 9084244 9084245 G A 0.0001 missense variant MED abnormality unknown/ NA 1729 of neuronal migration chet not applicable SI05 MUC16 19p13.2 rs145105175 9076005 9076006 A G 0.0024 missense variant MED abnormality unknown/ NA 6063 of neuronal migration hom SI06, SI14, not applicable MZT2A 2q21.1 rs200921574 132250066 132250067 G GGGA -1 inframe insertion MED None NA/ NA SI15 GCGG CA

Table S5 [Continued]. Potential recessive and X-linked mutations in the unsolved cases.

impact Polyphen/ model hom samples het samples gene location rs ID start end ref alt MAF impact severity Clinvar Sift hom SI06, SI14, not applicable MZT2A 2q21.1 rs200921574 132250066 132250067 G GGGAGCGGC -1 inframe insertion MED None NA/ NA SI15 A hom SI07, SI11 not applicable NA NA rs76161293 31362995 31362996 T C -1 splice acceptor HIGH None NA/ NA variant hom SI16 not applicable NA NA None 157732348 157732383 TGTTCA T -1 splice donor HIGH None NA/ NA CCTAG variant CCTCAT CATAG TGTCCC AGTGT GC hom SI07, SI11 not applicable NA NA rs76161293 31362995 31362996 T C -1 splice acceptor HIGH None NA/ NA variant chet not applicable SI07 NCOR2 12q24 rs184942554 124838654 124838655 C T 0.0049 missense variant MED None probably 9266 damaging/ NA chet not applicable SI07,SI14 NCOR2 12q24 rs61754987 124817778 124817779 C T 0.003 missense variant MED None possibly damaging/ NA hom SI13 not applicable NOP9 NA rs113258190, 24769848 24769849 A AGAG -1 inframe insertion MED None NA/ NA rs753459758 chet not applicable SI05 PASK 2q37.3 rs201982321 242066164 242066165 A G 0.0045 missense variant MED None probably 1621 damaging/ deleterious chet not applicable SI05 PASK 2q37.3 rs146901373 242079957 242079958 G A 0.0002 missense variant MED None probably 3256 damaging/ deleterious hom SI14 not applicable PCDH19 Xq22 rs749818933 99551551 99551552 T C 0 missense variant MED early infantile probably epileptic damaging/ encephalopat tolerated low hy 9 confidence hom SI06 not applicable PCLO 7q11.23- None 82784832 82784833 T TGAGCTGGA -1 inframe insertion MED pontocerebell NA/ NA q21.1 GGCTTAGCA ar hypoplasia GGACCAAGA type 3 G chet not applicable SI14 PKD1 16p13.3- rs199700485 2154530 2154531 G T 0.0015 missense variant MED 3-4 toe probably p13.12 9084 syndactyly damaging/ |abnormality NA of the kidney etc. chet not applicable SI14 PKD1 16p13.3- rs142733588 2153266 2153267 C T 0.0002 missense variant MED 3-4 toe benign/ NA p13.12 4516 syndactyly |abnormality of the kidney etc.

Table S5 [Continued]. Potential recessive and X-linked mutations in the unsolved cases.

impact Polyphen/ model hom samples het samples gene location rs ID start end ref alt MAF impact severity Clinvar Sift hom SI04 not applicable PLA2G4E NA None 42302329 42302330 G GCCCCCCCC -1 inframe insertion MED abnormality NA/ NA CCCC of neuronal migration hom SI16 not applicable PPP2R2B 5q31-q33 rs142461655 146258289 146258290 A AGCTGCTGC -1 inframe insertion MED None NA/ NA TGCTGCT hom SI07 not applicable RP11- NA rs148035776 81425350 81425351 C T -1 splice acceptor HIGH None NA/ NA 119F19.2 variant hom SI07 not applicable RP11- NA rs148035776 81425350 81425351 C T -1 splice acceptor HIGH None NA/ NA 119F19.2 variant hom SI12 not applicable RP11- NA None 85076258 85076259 C CTTTT -1 splice acceptor HIGH None NA/ NA 120I21.3 variant hom SI07 not applicable RP13- NA None 133264704 133264744 AGGCG A -1 splice donor HIGH None NA/ NA 672B3.2 CGGGT variant GAAGA CAGGG CGAGG GGAGC GGGTC TGCGG hom SI03,SI15 not applicable RP1L1 8p23 None 10467588 10467589 T TCCTCTAACT -1 inframe insertion MED occult NA/ NA GCACCCTCT macular CTTCTTGCAG dystrophy|reti CCCTTCTATT nitis ACTTTAGTCC pigmentosa etc. hom SI03,SI15 not applicable RP1L1 8p23 None 10467588 10467589 T TCCTCTAACT -1 inframe insertion MED occult NA/ NA GCACCCTCT macular CTTCTTGCAG dystrophy|reti CCCTTCTATT nitis ACTTTAGTCC pigmentosa etc. hom SI17 not applicable RP3-358H7.1 NA rs758255165 132276757 132276765 CTTTTT C -1 splice acceptor HIGH None NA/ NA TT variant hom SI11 not applicable RP4-683M8.2 NA None 48520861 48520875 GAGCA G -1 splice donor HIGH None NA/ NA GGGTG variant AGTA hom SI03 not applicable SGK223 NA None 8176319 8176320 T A -1 missense variant MED None possibly damaging/ NA hom SI05,SI14 not applicable SLC38A5 Xp11.23 None 48317040 48317041 G GCCT -1 inframe insertion MED abnormality NA/ NA of neuronal migration hom SI05,SI14 not applicable SLC38A5 Xp11.23 None 48317040 48317041 G GCCT -1 inframe insertion MED abnormality NA/ NA of neuronal migration hom SI12 not applicable TAF9 5q11.2- rs200123392 68665052 68665054 AC A -1 splice donor HIGH None NA/ NA q13.1 variant

Table S5 [Continued]. Potential recessive and X-linked mutations in the unsolved cases.

impact Polyphen/ model hom samples het samples gene location rs ID start end ref alt MAF impact severity Clinvar Sift chet not applicable SI09 TMPO 12q22 rs370309699 98909593 98909594 C T 0.00127421 splice donor HIGH cardiomyopathy|car NA/ NA variant diovascular phenotype etc. chet not applicable SI09 TMPO 12q22 None 98940187 98940188 A G 6.28E-05 missense MED cardiomyopathy|car possibly variant diovascular damaging/ phenotype etc. deleterious hom SI15 not applicable TSSC2 11p15.5 rs576755840 3418630 3418675 TGGGAGGT T -1 splice donor HIGH None NA/ NA GAGGAGCG variant TCTCCACC CGGCAGCC ACCCCGTC CGGGA chet not applicable SI14 WDR16 NA rs148905544 9489207 9489208 T A 0.00066303 missense MED None probably variant damaging/ deleterious chet not applicable SI14 WDR16 NA rs374035006 9536224 9536225 C T 0.00011628 stop gained HIGH None NA/ NA hom SI06 not applicable YBX1P6 NA None 112295223 112295226 GTT G -1 splice HIGH None NA/ NA acceptor variant hom SI03 not applicable ZDHHC11 NA rs200955248 814894 814895 G A 0.0014 missense MED None possibly variant damaging/ tolerated low confidence hom SI04, SI13 not applicable ZNF66 NA rs374547894 20981866 20981867 C CT 0 frameshift HIGH None NA/ NA variant hom SI04, SI13 not applicable ZNF66 NA rs374547894 20981866 20981867 C CT 0 frameshift HIGH None NA/ NA variant Chet = compound heterozygous; hom = homozoygous; MAF: Minor Allele Frequency; alt = alternate allele; ref = reference allele.

Table S6. Potential mutations in the unsolved cases, dominant model. (Note that genes dominantly mutated in any of the 15 healthy control subjects are not included in this list.)

Samples Samples impact hom het gene Location rs ID start end ref alt MAF impact severity Clinvar Polyphen/ Sift NA SI05 ABCF1 6p21.33 None 30550199 30550200 A AGAG -1 inframe MED long qt syndrome NA/ NA insertion NA SI03 AC012313.1 NA None 58907492 58907493 C T 2.21E-05 missense MED None possibly damaging/ variant deleterious low confidence NA SI14 AC079354.1 NA None 203058061 203058062 G A 0 stop gained HIGH None NA/ NA

NA SI03 AC093323.1 NA rs201074425, 6693879 6693880 A ATG 0 frameshift HIGH None NA/ NA rs746556174 variant NA SI07 ACADS 12q22-qter None 121174787 121174788 G A -1 splice HIGH deficiency of NA/ NA acceptor butyryl-coa variant dehydrogenase NA SI14 ADAT2 6q24.2 rs745389133 143753694 143753695 A G 1.50E-05 missense MED zellweger syndrome benign/ deleterious variant NA SI03 AFAP1 4p16.1 None 7774610 7774611 G A 9.13E-06 missense MED None possibly damaging/ variant deleterious NA SI07 AKAP10 17p11.1 None 19861880 19861881 C G -1 splice HIGH cardiac conduction NA/ NA acceptor defect variant NA SI07 AKAP9 7q21-q22 None 91700267 91700268 C T -1 missense MED cardiac probably damaging/ NA variant arrest|cardiac arrhythmia etc. NA SI05 AKAP9 7q21-q22 rs551918552 91672672 91672673 C T -1 missense MED cardiac NA/ NA variant arrest|cardiac arrhythmia etc. NA SI07 AL357673.1 NA None 54637044 54637046 AT A -1 frameshift HIGH None NA/ NA variant NA SI05 ALG1L2 NA None 129831617 129831626 CCAG C -1 splice HIGH None NA/ NA GGGC acceptor T variant NA SI03 ALG5 13q13.3 None 37573429 37573430 G A 0 missense MED pontocerebellar unknown/ tolerated low variant hypoplasia confidence NA SI09, AP003041.2 NA rs552607281 129936974 129936984 CTTTT C 0 splice HIGH None NA/ NA SI14 TTTTT donor variant NA SI03 AP3M2 8p11.2 None 42022662 42022663 C T -1 missense MED None possibly damaging/ variant deleterious NA SI05 ARHGAP19- NA None 99003855 99003857 TC T -1 frameshift HIGH None NA/ NA SLIT1 variant NA SI05 ASS1 9q34.1 None 133339553 133339554 G T -1 missense MED citrullinemia probably damaging/ variant deleterious NA SI09 ATP2B3 Xq28 None 152845483 152845484 A T -1 stop gained HIGH aldosterone NA/ NA producing adrenal cortex adenoma|inborn genetic diseases|spinocerebe llar ataxia

Table S6 [Continued]. Potential mutations in the unsolved cases, dominant model.

Samples Samples impact hom het gene Location rs ID start end ref alt MAF impact severity Clinvar Polyphen/ Sift NA SI03 BCAS3 NA None 59112109 59112110 G A 8.99E-06 missense MED None probably damaging/ variant deleterious NA SI07 C10orf53 NA None 50887747 50887748 C G -1 missense MED None probably damaging/ variant deleterious NA SI09 C15orf43 NA None 45270705 45270706 C CA -1 frameshift HIGH None NA/ NA variant NA SI09 C15orf43 NA None 45270708 45270714 GAAG G -1 frameshift HIGH None NA/ NA AA variant NA SI07 C2orf54 NA None 241827721 241827722 A G -1 missense MED None possibly damaging/ variant deleterious NA SI05 C2orf73 NA None 54587586 54587587 C G -1 missense MED None probably damaging/ variant tolerated NA SI03 C6orf48 6p21.3 rs748729448 31805121 31805122 T C -1 missense MED None benign/ NA variant NA SI14 C7orf50 NA None 1068074 1068075 C CG -1 frameshift HIGH None NA/ NA variant NA SI14 C9orf153 NA None 88870900 88870901 C A -1 splice HIGH None NA/ NA donor variant NA SI14 CAPN5 11q14 None 76799025 76799026 G A -1 stop gained HIGH vitreoretinopathy NA/ NA

NA SI14 CCDC169 NA None 36857816 36857817 A C -1 stop gained HIGH None NA/ NA

NA SI07 CCDC28A 6q24.1 None 139101026 139101027 A T -1 missense MED None probably damaging/ variant deleterious NA SI04 CDH4 20q13.3 rs754818766 60509128 60509129 C A 1.57E-05 missense MED None possibly damaging/ variant deleterious NA SI05 CDHR2 NA None 176005531 176005532 G A -1 missense MED None probably damaging/ variant tolerated NA SI14 CEACAM1 19q13.2 rs751330079 43026320 43026321 T C 4.48E-05 missense MED None probably damaging/ variant deleterious NA SI14 CEP290 12q21.3 None 88462420 88462421 C A -1 missense MED abnormality of the benign/ NA variant kidney|agenesis of cerebellar vermis|bardet-biedl syndrome|joubert syndrome |meckel- gruber syndrome|nephrono phthisis|senior-loken syndrome 6 etc. NA SI08, CHST12 7p22 None 2443287 2443302 GCGC G -1 splice HIGH None NA/ NA SI09 GAGG donor TGAG variant GGT

Table S6 [Continued]. Potential mutations in the unsolved cases, dominant model.

Samples Samples impact hom het gene Location rs ID start end ref alt MAF impact severity Clinvar Polyphen/ Sift NA SI05 CHST15 10q26 None 125804305 125804306 G A -1 missense MED None probably damaging/ variant deleterious NA SI03 CIC 19q13.2 None 42798817 42798818 C T -1 missense MED inborn genetic probably damaging/ NA variant diseases NA SI14 CIC 19q13.2 rs747754098 42796861 42796862 C T 4.48E-05 missense MED inborn genetic benign/ NA variant diseases NA SI07 CLCN3 4q33 None 170618621 170618622 G A -1 missense MED None probably damaging/ variant tolerated NA SI09 CLIC1 NA None 31701995 31701996 C G -1 missense MED None possibly damaging/ variant deleterious NA SI04 CNNM3 2q11.2 None 97494302 97494303 G T -1 splice HIGH None NA/ NA acceptor variant NA SI14 COL6A6 3q21 None 130300859 130300860 G T -1 missense MED None possibly damaging/ variant deleterious NA SI05 CSK 15q23-q25 None 75093892 75093893 G A -1 missense MED None probably damaging/ variant deleterious NA SI14 CTR9 11p15.3 rs753928657 10789928 10789929 G A 3.25E-05 missense MED None probably damaging/ variant deleterious NA SI03 CYFIP1 15q11 None 22958289 22958290 C T -1 missense MED None probably damaging/ variant deleterious NA SI04 DACT3 19q13.3 None 47152388 47152389 C T -1 missense MED None probably damaging/ variant tolerated NA SI05 DDX54 12q22-q23 rs757607025 113623243 113623244 T TGTC 0 frameshift HIGH None NA/ NA GGCC variant GCCA NA SI05, DGKQ 4p16.3 None 962078 962079 G GTGC -1 frameshift HIGH None NA/ NA SI13 CTCT variant CCTG CCCC GCCC NA SI05 CHST15 10q26 None 125804305 125804306 G A -1 missense MED None probably damaging/ variant deleterious NA SI03 CIC 19q13.2 None 42798817 42798818 C T -1 missense MED inborn genetic probably damaging/ NA variant diseases NA SI14 CIC 19q13.2 rs747754098 42796861 42796862 C T 4.48E-05 missense MED inborn genetic benign/ NA variant diseases NA SI07 CLCN3 4q33 None 170618621 170618622 G A -1 missense MED None probably damaging/ variant tolerated NA SI09 CLIC1 NA None 31701995 31701996 C G -1 missense MED None possibly damaging/ variant deleterious NA SI04 CNNM3 2q11.2 None 97494302 97494303 G T -1 splice HIGH None NA/ NA acceptor variant NA SI14 COL6A6 3q21 None 130300859 130300860 G T -1 missense MED None possibly damaging/ variant deleterious NA SI05 CSK 15q23-q25 None 75093892 75093893 G A -1 missense MED None probably damaging/ variant deleterious

Table S6 [Continued]. Potential mutations in the unsolved cases, dominant model.

Samples Samples impact hom het gene Location rs ID start end ref alt MAF impact severity Clinvar Polyphen/ Sift NA SI14 CTR9 11p15.3 rs753928657 10789928 10789929 G A 3.25E-05 missense MED None probably damaging/ variant deleterious NA SI03 CYFIP1 15q11 None 22958289 22958290 C T -1 missense MED None probably damaging/ variant deleterious NA SI04 DACT3 19q13.3 None 47152388 47152389 C T -1 missense MED None probably damaging/ variant tolerated NA SI05 DDX54 12q22-q23 rs757607025 113623243 113623244 T TGTC 0 frameshift HIGH None NA/ NA GGCC variant GCCA NA SI05, DGKQ 4p16.3 None 962078 962079 G GTGC -1 frameshift HIGH None NA/ NA SI13 CTCT variant CCTG CCCC GCCC NA SI03 DNAH7 2q33.1 None 196737122 196737123 C A 8.95E-06 stop gained HIGH None NA/ NA

NA SI07 DNAJC27 2p23.3 None 25174265 25174266 G C -1 stop gained HIGH None NA/ NA

NA SI03 DONSON NA None 34950632 34950636 TGTA T -1 inframe MED None NA/ NA deletion NA SI09 DSEL 18q21-q22 None 65181598 65181599 G T -1 missense MED None probably damaging/ variant deleterious NA SI09 EEF2 19pter-q12 None 3976607 3976608 G A 9.31E-06 missense MED None probably damaging/ variant deleterious NA SI09 ENKD1 NA None 67700084 67700085 G A -1 missense MED None probably damaging/ variant deleterious NA SI09 ETFB 19q13.3 rs762647069 51857659 51857675 ACAG A 1.51E-05 inframe MED glutaric acidemia NA/ NA CCTG deletion iib|inborn genetic GAAA diseases GGGG NA SI05 EXOSC10 1p36.22 None 11147607 11147608 A T -1 missense MED None probably damaging/ variant deleterious NA SI14 CTR9 11p15.3 rs753928657 10789928 10789929 G A 3.25E-05 missense MED None probably damaging/ variant deleterious NA SI03 CYFIP1 15q11 None 22958289 22958290 C T -1 missense MED None probably damaging/ variant deleterious NA SI04 DACT3 19q13.3 None 47152388 47152389 C T -1 missense MED None probably damaging/ variant tolerated NA SI05 DDX54 12q22-q23 rs757607025 113623243 113623244 T TGTC 0 frameshift HIGH None NA/ NA GGCC variant GCCA NA SI05, DGKQ 4p16.3 None 962078 962079 G GTGC -1 frameshift HIGH None NA/ NA SI13 CTCT variant CCTG CCCC GCCC

Table S6 [Continued]. Potential mutations in the unsolved cases, dominant model.

Samples Samples impact hom het gene Location rs ID start end ref alt MAF impact severity Clinvar Polyphen/ Sift NA SI03 DNAH7 2q33.1 None 196737122 196737123 C A 8.95E-06 stop gained HIGH None NA/ NA

NA SI07 DNAJC27 2p23.3 None 25174265 25174266 G C -1 stop gained HIGH None NA/ NA

NA SI04 FAM217B NA None 58519806 58519807 C A -1 missense MED None probably damaging/ variant deleterious NA SI03 FAM65B NA None 24843189 24843190 G C -1 missense MED deafness possibly damaging/ variant deleterious NA SI14 FAM73B NA None 131822909 131822910 C T -1 missense MED None probably damaging/ variant deleterious NA SI14 FAM83D NA rs368793717 37576567 37576568 G T 4.48E-05 missense MED None probably damaging/ variant deleterious NA SI04 FBN2 5q23-q31 None 127685696 127685697 T C 8.96E-06 missense MED arterial benign/ deleterious variant dissection|cerebral ischemia etc. NA SI07 FLCN 17p11.2 None 17125975 17125976 C T -1 splice HIGH birt-hogg-dub NA/ NA acceptor syndrome|carcinoma variant of colon etc. NA SI04 FLYWCH1 NA rs753873848 2980436 2980437 A G -1 missense MED None benign/ deleterious variant NA SI05 FOXK2 17q25 None 80526038 80526039 G A -1 missense MED None possibly damaging/ variant deleterious SI14 NA FOXR2 Xp11.21 None 55650996 55650997 C T -1 missense MED None possibly damaging/ variant deleterious NA SI14 FOXS1 20q11.1-q11.2 None 30433096 30433097 G T -1 missense MED None probably damaging/ variant deleterious NA SI09 FRK 6q21-q22.3 rs754085532 116263752 116263753 G C 3.00E-05 missense MED None benign/ NA variant NA SI03 GAK 4p16 None 905873 905877 CAGG C -1 stop lost HIGH None NA/ NA

NA SI09 GALNT10 5q33.2 None 153597595 153597596 C T -1 missense MED None NA/ NA variant

Table S6 [Continued]. Potential mutations in the unsolved cases, dominant model.

Samples Samples impact hom het gene Location rs ID start end ref alt MAF impact severity Clinvar Polyphen/ Sift NA SI14 GEN1 2p24.2 rs777769429 17942778 17942779 G C 1.50E-05 missense MED hereditary cancer- probably damaging/ variant predisposing deleterious syndrome NA SI14 GFRA4 NA None 3641189 3641190 G C -1 missense MED None probably damaging/ variant deleterious NA SI03 GRIK5 19q13.2 None 42506667 42506668 A G -1 missense MED None benign/ deleterious variant NA SI04 GUCY1A2 11q21-q22 None 106681034 106681035 G T -1 missense MED None probably damaging/ variant deleterious NA SI07 HAL 12q22-q23 rs779836173 96371760 96371761 C T 4.50E-05 missense MED histidinemia|increas probably damaging/ variant ed histidine deleterious NA SI09 HAND2-AS1 4q34.1 None 174480352 174480353 G A -1 splice HIGH None NA/ NA donor variant NA SI07 HAUS6 9p22.1 None 19080518 19080519 G T 8.99E-06 missense MED None probably damaging/ variant deleterious NA SI07 HIST1H4D 6p21.3 None 26189194 26189195 C A -1 missense MED None probably damaging/ variant deleterious NA SI03 HNRNPF 10q11.21- None 43882953 43882954 T C -1 missense MED None probably damaging/ q11.22 variant deleterious NA SI09 ICA1L NA None 203682255 203682256 T C -1 missense MED None possibly damaging/ variant deleterious NA SI14 IFI44L 1p31.1 None 79095586 79095587 G A 9.00E-06 missense MED None probably damaging/ variant tolerated NA SI03 IGHV7-81 NA None 107282970 107282971 G A -1 stop gained HIGH None NA/ NA

NA SI03 IMPA1 8q21.13-q21.3 None 82591397 82591398 T A -1 missense MED None probably damaging/ variant deleterious NA SI03 INTS8 8q22.1 rs755830705 95879374 95879375 T G 4.51E-05 missense MED long qt possibly damaging/ variant syndrome|malignant deleterious tumor of prostate NA SI09 IQCE NA None 2652349 2652350 A AGGT -1 splice HIGH None NA/ NA GAGC acceptor TGTG variant GGCG GGGC CTAG TCAT GGGA GG NA SI14 KCNAB2 1p36.3 rs531444097 6146022 6146023 T C 0 missense MED nephronophthisis|ren unknown/ NA variant al dysplasia and retinal aplasia NA SI04 KCNQ1 11p15.5 rs749073770 2594174 2594175 G A 4.58E-05 missense MED long qt syndrome possibly damaging/ variant 1|arrhythmia|atrial deleterious fibrillation etc. NA SI05 KLB 4p14 None 39450305 39450306 A C -1 stop lost HIGH None NA/ NA

Table S6 [Continued]. Potential mutations in the unsolved cases, dominant model.

Samples Samples impact Polyphen/ hom het gene Location rs ID start end ref alt MAF impact severity Clinvar Sift

NA SI14 KLKB1 4q35 rs200255707 187179270 187179271 C G 3.58E-05 missense MED prekallikrein probably variant deficiency damaging/ deleterious NA SI03 KMT2C 7q36 None 151836310 151836311 C T -1 missense MED None probably variant damaging/ NA NA SI14 KRT16 17q12-q21 None 39766693 39766694 C A 8.98E-06 missense MED inborn genetic possibly variant diseases etc. damaging/ deleterious NA SI03 KRTAP4-3 NA None 39324083 39324084 C G -1 missense MED None unknown/ variant deleterious NA SI03, LCORL 4p15.3 None 18023311 18023312 G GGCGG -1 inframe MED None NA/ NA SI15 CGGCG insertion GCGGC GGCAG CA NA SI03 LMCD1 3p26-p24 rs761332410 8578988 8578989 C T 2.69E-05 missense MED None probably variant damaging/ deleterious NA SI09 LOC100132874 NA None 45120089 45120150 GGCCTCT G -1 inframe MED None NA/ NA CCAGCGT deletion GACCCTG TCTGCAT GGGGCCT TTCCAGC GTGACCC TGTCTGC CTGTT NA SI04 LOC100652807 NA None 28617507 28617508 G T -1 missense MED None NA/ NA variant SI09, SI17 LOC100653133 NA rs202162063 32787186 32787187 T C -1 missense MED None NA/ NA SI13 variant NA SI09 LOC101059952 NA rs747411142 37116683 37116684 G A -1 missense MED None NA/ NA variant NA SI05 LOC101927594 NA rs200623579 102304672 102304673 T C -1 missense MED None NA/ NA variant NA SI03, LOC101927848 NA rs148517271 3170820 3170821 T TCCCTG -1 inframe MED None NA/ NA SI11, GCGCC insertion SI16 TCCTC NA SI03 LOC101927903 NA None 45442373 45442396 GTGTGTG G -1 frameshift HIGH None NA/ NA CGCGCGC variant GCGCGTG TC

Table S6 [Continued]. Potential mutations in the unsolved cases, dominant model.

Samples Samples MA impact Polyphen/ hom het gene Location rs ID start end ref alt F impact severity Clinvar Sift NA SI14 LOC101928218 NA None 11135656 11135657 G A -1 missense variant MED None NA/ NA

NA SI14 LOC101928218 NA None 11135653 11135655 CT C -1 frameshift variant HIGH None NA/ NA

NA SI04 LOC101928672 NA None 139957996 139957997 G A -1 missense variant MED None NA/ NA

NA SI07, SI14 LOC101928841 NA rs56193086 114058440 114058441 C T -1 missense variant MED None NA/ NA

NA SI07, SI14 LOC101928841 NA None 114058426 114058432 CCCGTT C -1 frameshift variant HIGH None NA/ NA

NA SI07, SI14 LOC101928841 NA None 114058443 114058444 T A -1 missense variant MED None NA/ NA

NA SI07, SI09 LOC101928884 NA rs868671387 1328564 1328565 T C -1 missense variant MED None NA/ NA

NA SI09 LOC101928884 NA rs866503911 1327424 1327425 A G -1 missense variant MED None NA/ NA

NA SI09 LOC101928884 NA rs866831612 1327425 1327426 C G -1 missense variant MED None NA/ NA

SI04 NA LOC101928917 NA None 148630211 148630212 G T -1 missense variant MED None NA/ NA

NA SI04 LOC101929103 NA None 1286805 1286806 G GGAGGGGGC -1 frameshift variant HIGH None NA/ NA CTGGACGGG GCTGGAGCG GTGGGGAGA GT NA SI07 LOC101929113 NA None 4848588 4848609 GACGC G -1 frameshift variant HIGH None NA/ NA CACTG CCCCC GTTCC A NA SI09 LOC101929113 NA None 4848346 4848347 C CTTCCGACG -1 frameshift variant HIGH None NA/ NA CCACTGGCC CCG NA SI09 LOC101929113 NA rs528814344 4848369 4848370 C A -1 missense variant MED None NA/ NA

NA SI14 LOC101929300 NA None 16130630 16130631 C A -1 missense variant MED None NA/ NA

NA SI03 LOC101929740 NA None 38408214 38408215 C T -1 missense variant MED None NA/ NA

NA SI03, SI13, LOC101929747 NA rs17515799 96271601 96271602 G GC -1 frameshift variant HIGH None NA/ NA SI16 NA SI14 LOC729900 NA None 23104933 23104934 C G -1 missense variant MED None NA/ NA

NA SI09 LONRF2 NA None 100915379 100915380 C T -1 missense variant MED None probably damaging/ deleterious NA SI05 LRRC6 8q24.22 None 133687517 133687518 A T -1 splice donor HIGH PCD | NA/ NA variant Kartagener

Table S6 [Continued]. Potential mutations in the unsolved cases, dominant model.

Samples Samples impact hom het gene Location rs ID start end ref alt MAF impact severity Clinvar Polyphen/ Sift NA SI04 LRRC8E 19p13.2 rs371880747 7965192 7965193 C T 2.70E-05 missense MED long qt syndrome benign/ variant deleterious NA SI05 LSR 19q13.12 rs748542666 35757272 35757273 G A 1.99E-05 missense MED None probably variant damaging/ deleterious SI14 NA MAP3K15 Xp22.12 rs771700881 19506958 19506959 G C 2.59E-05 missense MED pyruvate dehydrogenase possibly variant e1-alpha deficiency damaging/ deleterious NA SI04 MGC4294 NA rs558457591 57210232 57210233 C T -1 missense MED None unknown/ NA variant NA SI09 MMP17 12q24.33 None 132325178 132325179 T C -1 missense MED None probably variant damaging/ tolerated NA SI07 MRC2 17q23.2 rs753363212 60768391 60768392 C G 1.85E-05 missense MED None probably variant damaging/ deleterious NA SI09 MVP 16p13.1-p11.2 rs576877456 29832694 29832695 G C -1 splice HIGH None NA/ NA donor variant NA SI07 MYCN 2p24.1 None 16085637 16085638 G A -1 missense MED feingold syndrome 1 probably variant damaging/ deleterious NA SI03 MYH11 16p13.13- rs760908992 15813471 15813472 G C 2.69E-05 missense MED abnormality of the left benign/ NA p13.12 variant ventricle|altered myosin contractile function etc. NA SI03 MYH7 14q12 None 23897781 23897782 T C -1 missense MED arrhythmia|atrial septal probably variant defect etc. damaging/ tolerated NA SI05 MYO5B 18q21 None 47527667 47527668 G T -1 missense MED congenital microvillous possibly variant atrophy|diarrhea with damaging/ microvillus atrophy deleterious NA SI04 MYOF 10q24 None 95109726 95109727 C T 0 splice HIGH None NA/ NA acceptor variant NA SI07 NEB 2q22 rs112229327 152531893 152531894 C A 2.58E-05 missense MED congenital muscular probably variant dystrophy|inborn genetic damaging/ NA diseases|limb pain|muscle weakness etc. NA SI07 NEB 2q22 rs574363877 152531872 152531873 T G 2.25E-05 missense MED congenital muscular probably variant dystrophy|inborn genetic damaging/ NA diseases|limb pain|muscle weakness etc. NA SI07 NHSL2 NA None 71352069 71352070 A T 2.65E-05 missense MED spinocerebellar ataxia benign/ variant deleterious NA SI14 OR5AN1 11q12.1 None 59132864 59132865 T C 2.99E-05 stop lost HIGH None NA/ NA

Table S6 [Continued]. Potential mutations in the unsolved cases, dominant model.

Samples Samples impact hom het gene Location rs ID start end ref alt MAF impact severity Clinvar Polyphen/ Sift NA SI07 P2RY6 11q13.5 None 72983452 72983453 G C -1 missense MED None NA/ NA variant NA SI09 PAPPA2 NA None 176675534 176675535 G A 8.97E-06 missense MED None possibly variant damaging/ deleterious NA SI03 PARK2 NA rs137853059 162864345 162864346 A T 3.00E-05 missense MED parkinson disease possibly variant damaging/ deleterious SI14 NA PCDH19 Xq22 rs749818933 99551551 99551552 T C 0 missense MED early infantile epileptic probably variant encephalopathy 9 damaging/ tolerated low confidence NA SI03 PCDHB9 5q31 None 140569234 140569239 GAA G -1 frameshift HIGH None NA/ NA AT variant NA SI09 PCK2 14q11.2-q12 None 24566156 24566157 C T -1 missense MED None benign/ variant deleterious NA SI03 PDE12 3q21.2 rs780463808 57640449 57640451 AC A -1 frameshift HIGH None NA/ NA variant NA SI04 PGLYRP1 19q13 rs770993705 46526193 46526194 C T 1.84E-05 missense MED None possibly variant damaging/ deleterious NA SI03 PGS1 17q25.3 None 76415730 76415731 C T 1.79E-05 stop gained HIGH None NA/ NA

NA SI03 PKD1 16p13.3- None 2163259 2163260 T C -1 missense MED 3-4 toe benign/ NA p13.12 variant syndactyly|abnormality of the kidney|cerebral aneurysm etc. NA SI05 PLEKHG2 19q13.1 None 39914997 39914998 C T -1 missense MED abnormality of neuronal unknown/ NA variant migration NA SI05 PLEKHM2 1p36.21 None 16047825 16047826 T A -1 missense MED None probably variant damaging/ deleterious low confidence NA SI09 PLXND1 3q22 None 129286636 129286640 AGA A 9.01E-06 inframe MED None NA/ NA C deletion NA SI14 POM121C 7q11.23 None 75051114 75051115 G C -1 missense MED None benign/ NA variant NA SI05 POMGNT1 1p34-p33 None 46662407 46662408 T A -1 missense MED congenital muscular possibly variant dystrophy damaging/ NA NA SI04 POTED 21q11.2 None 14982960 14982961 G A -1 missense MED None probably variant damaging/ tolerated NA SI07 PPCS 1p13.1-p12 None 42938850 42938851 T G -1 missense MED None NA/ NA variant

Table S6 [Continued]. Potential mutations in the unsolved cases, dominant model.

Samples Samples impact hom het gene Location rs ID start end ref alt MAF impact severity Clinvar Polyphen/ Sift NA SI03 PRDM15 NA None 43221742 43221743 G A -1 missense MED None possibly variant damaging/ deleterious low confidence NA SI04 PRDM7 NA None 90124698 90124699 A C -1 stop lost HIGH None NA/ NA

NA SI03 PRR24 NA None 47778504 47778505 C G 4.59E-05 missense MED None unknown/ variant deleterious low confidence NA SI05 PSG8 19q13.2 None 43259248 43259249 C G -1 missense MED None probably variant damaging/ deleterious NA SI14 PTH2R 2q33 rs781693841 209302299 209302300 T C 4.50E-05 missense MED fleck corneal probably variant dystrophy damaging/ deleterious NA SI14 PTPDC1 NA None 96846905 96846906 C A 9.11E-06 missense MED None probably variant damaging/ deleterious low confidence NA SI03 PTPRH 19q13.4 None 55708150 55708151 C G -1 missense MED None probably variant damaging/ deleterious NA SI05 R3HCC1 NA None 23153511 23153512 A C -1 missense MED None possibly variant damaging/ deleterious NA SI03 RASSF8- NA None 26109157 26109158 A C -1 splice donor HIGH None NA/ NA AS1 variant NA SI09 RELN 7q22 None 103234178 103234179 C T -1 missense MED abnormality of probably variant neuronal damaging/ migration|epilepsy deleterious |lissencephaly NA SI07 RMI2 16p13.13 rs768453762 11389070 11389076 AGGGGT A -1 splice acceptor HIGH None NA/ NA variant NA SI05 RP11- NA None 83837272 83837273 C T -1 splice donor HIGH None NA/ NA 483P21.2 variant NA SI03 RP11- NA rs769866764 144898867 144898897 TTAGAATA T -1 splice donor HIGH None NA/ NA 673E1.1 CAAAACAT variant GTAAGTAA CTATTA SI07 NA RP13- NA None 133264704 133264744 AGGCGCGG A -1 splice donor HIGH None NA/ NA 672B3.2 GTGAAGAC variant AGGGCGAG GGGAGCGG GTCTGCGG NA SI04 RP5- NA None 22381749 22381750 C T -1 splice acceptor HIGH None NA/ NA 1004I9.1 variant

Table S6 [Continued]. Potential mutations in the unsolved cases, dominant model.

Samples Samples impact hom het gene Location rs ID start end ref alt MAF impact severity Clinvar Polyphen/ Sift NA SI05 RP5- NA None 111860550 111860551 T C -1 splice acceptor HIGH None NA/ NA 1125M8.2 variant NA SI07 SALL1 16q12.1 None 51173502 51173503 A G -1 missense MED townes-brocks- possibly variant branchiootorenal- damaging/ like deleterious syndrome|townes syndrome NA SI03, SI14 SEC16A 9q34.3 None 139368939 139368940 T G -1 splice acceptor HIGH familial aplasia of NA/ NA variant the vermis NA SI03 SEMA6B 19p13.3 None 4550799 4550834 GGGGTTCT G -1 splice donor HIGH None NA/ NA CACCGGGG variant TCGAGGCA CCTGATCC TCC NA SI09 SHF 15q21.1 None 45492926 45492927 C A -1 missense MED None benign/ NA variant NA SI04 SI 3q25-q26 rs540333172 164780174 164780175 A C 1.80E-05 missense MED malignant tumor probably variant of damaging/ prostate|sucrase- deleterious isomaltase deficiency NA SI04 SLC16A4 NA rs766095761 110921586 110921587 C T 4.50E-05 stop gained HIGH None NA/ NA

NA SI14 SLC35E4 NA None 31032504 31032505 G T -1 missense MED None benign/ variant deleterious NA SI03 SLC4A3 2q36 None 220494118 220494119 C CG -1 frameshift HIGH None NA/ NA variant NA SI05 SLC7A14 3q26.2 None 170244546 170244547 G C -1 missense MED retinitis probably variant pigmentosa 68 damaging/ deleterious NA SI05 SNPH 20p13 None 1277711 1277712 G A -1 missense MED None NA/ NA variant NA SI03 SNX7 1p21.3 rs757467199 99161080 99161081 C T 2.69E-05 missense MED None possibly variant damaging/ deleterious NA SI07 SNX9 6q26.3 None 158330819 158330820 A G -1 missense MED None possibly variant damaging/ deleterious NA SI03 SP100 2q37.1 None 231367817 231367818 A T -1 missense MED None possibly variant damaging/ deleterious NA SI14 SPATA3 NA None 231865151 231865152 C T 3.49E-05 missense MED None possibly variant damaging/ deleterious NA SI07 SPCS2 NA None 74660593 74660594 C A -1 missense MED None unknown/ variant tolerated low confidence

Table S6 [Continued]. Potential mutations in the unsolved cases, dominant model.

Samples Samples impact hom het gene Location rs ID start end ref alt MAF impact severity Clinvar Polyphen/ Sift NA SI03 SPEF2 5p13.2 None 35776366 35776367 A T -1 missense MED None benign/ variant deleterious NA SI14 STARD9 15q15 None 42956050 42956051 C A -1 missense MED None probably variant damaging/ NA NA SI09 STS Xp22.32 None 7177755 7177756 T C -1 missense MED x-linked benign/ variant ichthyosis with deleterious steryl-sulfatase deficiency NA SI07 SVEP1 9q32 None 113173542 113173543 G A -1 missense MED None probably variant damaging/ tolerated NA SI03 TAMM41 3p25.2 None 11858658 11858659 C CAG -1 frameshift HIGH None NA/ NA variant NA SI04 TANC1 2q24.2 rs749103357 160082213 160082214 C T 3.58E-05 missense MED radiotherapy possibly variant response - damaging/ toxicity/adr deleterious NA SI07 TAS1R1 1p36 None 6639279 6639280 C T -1 missense MED None possibly variant damaging/ deleterious NA SI07 TAS2R50 12p13.2 None 11138568 11138571 AAT A -1 frameshift HIGH None NA/ NA variant NA SI04 TDRD6 6p12.3 None 46668775 46668776 C T -1 missense MED long qt syndrome possibly variant damaging/ deleterious NA SI05 TGM4 3p22- None 44943292 44943293 G A -1 missense MED None possibly p21.33 variant damaging/ deleterious NA SI04 THADA 2p21 None 43459988 43459989 C G -1 missense MED abnormality of probably variant neuronal damaging/ migration deleterious NA SI05 THUMPD2 2p22-p21 None 39997172 39997176 CAAG C 4.23E-05 inframe deletion MED None NA/ NA

NA SI04 TMEM174 5q13.2 None 72469134 72469135 C T -1 missense MED None probably variant damaging/ tolerated NA SI03 TMEM74 8q23.2 rs768684104 109796943 109796947 TGGC T 1.50E-05 inframe deletion MED None NA/ NA

NA SI04 TMPRSS11E 4q13.2 None 69342002 69342009 CTCTA C -1 inframe deletion MED None NA/ NA GG NA SI03 TMTC1 12p11.22 None 29920912 29920913 C T -1 missense MED None probably variant damaging/ deleterious NA SI09 TNNI3 19q13.4 rs397516344 55666173 55666174 G A 8.96E-06 missense MED cardiomyopathy|c possibly variant ardiovascular damaging/ phenotype etc. deleterious NA SI03 TNS3 7p12.3 None 47521266 47521267 G A -1 missense MED None NA/ NA variant

Table S6 [Continued]. Potential mutations in the unsolved cases, dominant model.

Samples Samples impact hom het gene Location rs ID start end ref alt MAF impact severity Clinvar Polyphen/ Sift NA SI03 TOM1L1 17q22 None 53016308 53016313 TAACA T -1 frameshift HIGH None NA/ NA variant NA SI04 TRIM61 NA rs752246419 165890280 165890281 G A -1 stop gained HIGH None NA/ NA

NA SI04 TRPV2 Chr.17 None 16329583 16329584 G A -1 splice donor HIGH None NA/ NA variant NA SI07 TRRAP 7q21.2-q22.1 None 98567767 98567768 A G -1 missense MED malignant tumor benign/ NA variant of prostate NA SI07 TSLP 5q22.1 rs11539838 110409260 110409261 G C 4.50E-05 missense MED None probably variant damaging/ deleterious NA SI07 TTLL8 NA None 50454849 50454850 C T -1 splice donor HIGH megalencephalic NA/ NA variant leukoencephalopat hy with subcortical cysts NA SI04 TTYH3 NA None 2685393 2685394 T G -1 missense MED None unknown/ variant deleterious low confidence NA SI09 UACA 15q24 None 70959465 70959466 C A -1 missense MED None benign/ variant deleterious NA SI07 UBR4 1p36.13 None 19412653 19412654 G T -1 missense MED long qt syndrome benign/ NA variant NA SI05 USP8 15q21.2 None 50776554 50776559 TAAAG T -1 splice donor HIGH pituitary NA/ NA variant dependent hypercortisolism NA SI04 USPL1 NA None 31205448 31205453 TGTTG T -1 frameshift HIGH None NA/ NA variant NA SI09 WDR49 NA None 167196713 167196714 C G 9.05E-06 missense MED malignant tumor possibly variant of prostate damaging/ deleterious NA SI07 WDR6 3p21.31 rs758104902 49046360 49046363 CTG C 0 frameshift HIGH None NA/ NA variant NA SI07 WDR6 3p21.31 rs747354637 49046364 49046367 AGT A 0 frameshift HIGH None NA/ NA variant NA SI09 WDR62 19q13.12 None 36594252 36594254 CT C -1 frameshift HIGH abnormality of NA/ NA variant neuronal migration|microce phaly etc. NA SI14 WDR65 NA None 43700195 43700196 C T -1 stop gained HIGH van der woude NA/ NA syndrome 2 NA SI14 ZAN 7q22 None 100349448 100349449 T C 2.98E-05 missense MED None benign/ NA variant NA SI02, SI04, ZDHHC11 NA rs199757139 796007 796029 GGCTC G -1 splice donor HIGH None NA/ NA SI07, SI14 CCATTT variant CCCAG TACTGT

Table S6 [Continued]. Potential mutations in the unsolved cases, dominant model.

Samples Samples impact hom het gene Location rs ID start end ref alt MAF impact severity Clinvar Polyphen/ Sift NA SI07 ZFHX3 16q22.3-q23.1 None 72830442 72830443 G GGG -1 inframe MED inborn genetic NA/ NA TGG insertion diseases|prostate A cancer NA SI14 ZFHX3 16q22.3-q23.1 None 72828343 72828344 G A -1 missense MED inborn genetic unknown/ NA variant diseases|prostate cancer NA SI09 ZNF106 NA rs775858778 42713296 42713297 C A 1.79E-05 missense MED limb-girdle muscular benign/ variant dystrophy|limb-girdle deleterious muscular dystrophy NA SI04 ZNF33A 10q11.2 None 38344861 38344862 A G -1 missense MED None probably variant damaging/ deleterious NA SI05 ZNF43 19p13.1-p12 rs536331309 21991297 21991298 T C -1 missense MED None possibly variant damaging/ deleterious NA SI09 ZNF638 2p13.2 rs771571099 71575988 71575989 A G 1.83E-05 missense MED None NA/ NA variant NA SI07 ZNF679 NA rs765986106 63726311 63726312 C G 3.06E-05 missense MED None benign/ variant deleterious NA SI04 ZNF786 NA None 148768920 148768921 G A -1 missense MED None benign/ variant deleterious NA SI05 ZNF83 19q13.3-q13.4 rs199877693 53117417 53117418 A AT -1 frameshift HIGH None NA/ NA variant MAF: Minor Allele Frequency; alt = alternate allele; ref = reference allele