Supplementary Information

Supplementary Table S1.Variant prioritisation strategy

Prioritisation Parameter / II:1 and II:2
Variants and indels identified / 47,908
+ Autosomal / 40,302
+ Absent or present with a frequency <1% in dbSNP, NHLBI ESP and 1000G / 3,817
+ Coding (missense, nonsense, splice site and indel) / 1,004
+ Homozygous / 103
+ Absent or present with a frequency <1% in control exomes / 16

Assuming an autosomal recessive model, we prioritised autosomal coding (missense, nonsense, splice site and indels) variants that were (i) autosomal, (ii) absent or present with a frequency <1% in dbSNP130,NHLBI Exome Variant Server database and 1000 Genomes, (iii) coding, (iv) homozygous, (v)absent or present with a frequency <1% in our 60 control exomes. As the DNA samples from the two affected children were pooled for exome sequencing, variants designated homozygous means that both children are homozygous for the alternate allele.

Supplementary Table S2. Primer sequences for RMND1 mutation validation

Gene / Variant / Forward primer 5’-3’ / Reverse primer 5’-3’ / Product size (bp)
RMND1 / c.1250G>A / CCTTTGGGGAGAGGACTGAG / AGCATAGCCCCTGTATTTGTG / 295

Primer sequences were designed using the Exon Primer program available in the UCSC Genome Browser.

Supplementary Table S3. Rare homozygous variants shared by the affected siblings

Gene / Transcript / Change at cDNA level / Change at protein level / dbSNP ID / MAF / Associated human disorder
AMPD2 / NM_203404.1 / c.1521C>G / p.(Ser507Arg) / rs116415979 / 0.4% / Pontocerebellar hypoplasia type 9
OR2T29 / NM_001004694.2 / c.97G>T / p.(Val33Leu) / novel / - / -
FRG2 / NM_001286820.1 / c.329C>G / p.(Thr110Arg) / rs143585119 / 0.1% / -
RMND1 / NM_017909.3 / c.1250G>A / p.(Arg417Gln) / - / - / Combined oxidative phosphorylation deficiency 11
SYNE1 / NM_182961.3 / c.8825G>T / p.(Ser2942Ile) / novel / - / Emery-Dreifuss muscular dystrophy (AD)
Spinocerebellar ataxia 8 (AR)
MROH5 / NM_207414.2 / c.3439G>C / p.(Gly1147Arg) / novel / - / -
PDILT / NM_174924.1 / c.139G>T / p.(Val47Leu) / rs193158575 / 0.09% / -
MUC16 / NM_024690.2 / c.19499C>T / p.(Thr6500Ile) / novel / - / -
MUC16 / NM_024690.2 / c.19486T>G / p.(Tyr6496Asp) / novel / - / -
MUC16 / NM_024690.2 / c.13808G>A / p.(Arg4603Gln) / rs201713021 / n/a / -
ZNF846 / NM_001077624.1 / c.1069G>C / p.(Val357Leu) / rs139394148 / 0.2% / -
ZNF846 / NM_001077624.1 / c.793C>A / p.(Gln265Lys) / rs182871540 / 0.2% / -
DOCK6 / NM_020812.3 / c.322C>T / p.(Gln108*) / novel / - / Adams-Oliver syndrome 2 (AR)
RFX1 / NM_002918.4 / c.2869C>A / p.(Leu957Met) / rs111774835 / n/a / -
PSG6 / NM_002782 / c.136delT / p.(Ser46Profs*27) / novel / - / -
HELZ2 / NM_001037335.2 / c.4766G>A / p.(Gly1589Asp) / rs145214493 / 0.2% / -

Whole exome sequencing and variant prioritisation identified 16 novel or rare homozygous candidate variants that were shared by two of the affected siblings. AD; autosomal dominant, AR; autosomal recessive, MAF; minor allele frequency, n/a; not available

Supplementary Table S4. Genes associated with calcification, abnormal pterins or dry thickened skin

Brain calcification and/or abnormal pterins
Gene / Associated disorder
CA2 / Carbonic Anhydrase II
CPT2 / Carnitine palmitoyltransferase II deficiency
CRY1 / Pterin cofactor
CRY2 / Pterin cofactor
ECM1 / Urbach-Wiethe disease
ENPP1 / Arterial calcification, generalized, of infancy, 1
GFAP / Alexander disease
GNAQ / Sturge-Weber syndrome
LIPA / Wolman disease
MYO5A / Griscelli syndrome
OCLN / Microcephaly, intracranial calcification, intellectual disability
PCBD1 / Hyperphenylalaninemia, BH4-deficient, D
PDGFRB / Basal ganglia calcification
QDPR / Dihydropteridine reductase deficiency
RNASEH2A / Aicardi-Goutieres syndrome 4
RNASEH2B / Aicardi-Goutieres syndrome 2
RNASEH2C / Aicardi-Goutieres syndrome 3
SAMHD1 / Aicardi-Goutieres syndrome 5
TREX1 / Aicardi-Goutieres syndrome 1
SPR / Sepiapterin reductase deficiency
Dry, thickened or pigmented skin
CTSC / Palmoplantar hyperkeratosis
DSP / Keratosis palmoplantarisstriata II
ELOVL4 / Ichthyosis, spastic quadriplegia, and mental retardation
PAH / Phenylketonuria (pigmentation and rashes)
HRAS / Costello syndrome
KRT1 / Hyperkeratosis
KRT6C / Palmoplantar keratoderma, nonepidermolytic, focal or diffuse
KRT16 / Palmoplantar keratoderma, nonepidermolytic, focal
LIPN / Congenital ichthyosis type 8
RSPO1 / Palmoplantar hyperkeratosis
TINF2 / Dyskeratosiscongenita

The exome data was analysed to determine if patient II:1 had an additional mutation that could account for the extra clinical features of intracranial calcification, abnormal pterins and dry, thickened and pigmented skin. Genes associated with disorders that include any of these three features were obtained from Phenomizer ( Orphanet ( and OMIM (