Suppl Table 1. Detailed Data of MCOLN1 Molecular Variants Used in Table 1

Suppl Table 1. Detailed data of MCOLN1 molecular variants used in Table 1.

Gene and Variant type cDNA Some phenotype protein and Additional information Pathogenicity prediction2 Database status3 and protein level correlation localization MAF [%]1 Deletion of 6434 bps spans exon 1-6 and the Functional analysis: gross deletion first 12 bps of exon 7 and affecting the c.-1015_789del exon 1-7 Pathogenic (null mutation to HGMD accession: CG005059 (DM: Associated with a severe (minor AJ) translation initiation codon (Met1). The p.0 null yield no mRNA and no Mucolipidosis IV) MLIV [no data] consequence of this change is no protein protein product). product. The 93bp segment from mitochondrial NADH dehydrogenase 5 (MTND5 ) is inserted inframe prior to the last nucleotide of exon 2 of MCOLN1. This variation creates a frame shift starting at codon Gln79. The new reading MutationTaster: disease frame ends in a STOP codon at position 8. causing inframe c.236_237ins exon 2 Alteration abolish proper splicing of Functional analysis: HGMD accession: CN044483 (DM: Associated with moderate insertion p.0? null MCOLN1. The splice site at the end of the Pathogenic (null mutation to Mucolipidosis IV) phenotype of MLIV [no data] exon 2 is not used due to an inhibitory effect yield no mRNA and no of the inserted segment, resulting in two protein product). aberrant splice products containing stop codons in the downstream intron and these products are eliminated via nonsense-mediated decay - NMD). HGMD accession: CM003596 (DM: Mucolipidosis IV)

Nonsense ClinVar: RCV000005443.5 SIFT: Deleterious exon 3 [ALL:0.0024 The reading frame is interrupted by a (Pathogenic- Mucolipidosis type IV), c.304C>T MutationTaster: disease Associated with a severe TM1-TM2 AFR:0.0062 premature STOP codon at position 102. The RCV000498354.4 (Pathogenic - not p.(Arg102*) causing MLIV loop SAS:0.0033 mRNA produced might be targeted for NMD. provided). Predicted truncated protein NFE:0.0035] dbSNP: rs121908373 (validated or no product. dbSNP entry - Clinical significance: CLIN_pathogenic). HGMD accession: CM026007 (DM: The substitution of the highly conserved Mucolipidosis IV) amino acid. Moderate physicochemical ClinVar: RCV000194523.1 exon 3 difference between Leu and Pro. Variant SIFT: Deleterious (Pathogenic - Mucolipidosis type IV) c.317T>C missense Associated with milder TM1-TM2 decreases formation and extrusion of MutationTaster: disease Uniprot : this variant is reported as p.(Leu106Pro) [no data] phenotype loop tubulovesicular structures when causing possibly pathogenic overexpressed; disrupts tetrameric assembly; dbSNP: rs797044825 (not validated abolishes lysosomal localization. dbSNP entry - Clinical significance: CLIN_pathogenic) exon 3 inframe c.395_397del Microdeletion of 3 nucleotides CTG occurring MutationTaster: disease HGMD accession: CP149142 (DM: Associated with a severe TM1-TM2 deletion p.? in cis (on the same allele) with c.468_474dup causing Mucolipidosis IV) MLIV loop [no data] HGMD accession: CS002473 (DM: Mucolipidosis IV) splice site MutationTaster: disease ClinVar: RCV000825567.1 (major AJ) Substitution affecting the splice site acceptor causing (Pathogenic- Mucolipidosis), [ALL:0.019 site in intron 3 leading to frame shift. The Predicted change at acceptor c.406-2A>G intron 3 RCV000058927.3 (Pathogenic - not Associated with a severe AMR:0.0085 reading frame is interrupted by a premature site 2 bps downstream: - p.? null provided), RCV000005438.7 MLIV ASJ:0.41 STOP codon. The mRNA produced might be 100.0% (MaxEnt: -100.0%, (Pathogenic - Mucolipidosis type IV) NFE:0.0054 targeted for nonsense mediated decay (NMD). NNSPLICE: -100.0%, SSF: dbSNP: rs104886461 (validated OTH:0.014] -100.0%) dbSNP entry - Clinical significance: CLIN_pathogenic) Microduplication of 7 nucleotides TTGGACC occurring in cis (on the same allele) with frameshift c.468_474dup exon 4 c.395_397del. This variation probable creates MutationTaster: disease HGMD accession: CP149142 (DM: Associated with a severe duplication p.? null a frame shift starting at codon Asn159. The causing Mucolipidosis IV) MLIV [no data] new reading frame ends in a STOP codon at position 27. HGMD accession: CD003634 (DM: Mucolipidosis IV) Microdeletion of 2bps CC. This variation frameshift ClinVar: RCV000192306.1 c.473_474del exon 4 creates a frame shift starting at codon Thr158. MutationTaster: disease deletion (Pathogenic - Mucolipidosis type IV) No data p.(Thr158Lysfs*25) null The new reading frame ends in a STOP codon causing [0.00] dbSNP: rs797044821 (not validated at position 25. dbSNP entry - Clinical significance: CLIN_pathogenic) HGMD accession: CM003597 (DM: Mucolipidosis IV) The reading frame is interrupted by a ClinVar: RCV000195067.2 (Likely c.514C>T exon 4 Nonsense premature STOP codon at position 172. The MutationTaster: disease pathogenic - Mucolipidosis type IV) No data p.(Arg172*) null [0.00] mRNA produced might be targeted for causing dbSNP: rs797044824 (not validated nonsense mediated decay (NMD). dbSNP entry - Clinical significance: CLIN_likely_pathogenic) HGMD accession: CM011400 (DM: The substitution of the highly conserved Mucolipidosis IV) amino acid. ClinVar: RCV000192300.2 (Likely exon 6 missense Small physicochemical difference between SIFT: Damaging/ Tolerated c.694A>C pathogenic - Mucolipidosis type IV) The typical, rather severe TM1-TM2 [ALL:0.0016 Thr and Pro. Protein fails to localize to late MutationTaster (v2013): p.(Thr232Pro) Uniprot: this variant is reported as presentation MLIV loop NFE:0.0035] endosomes; abolishes Fe(2+) permeability; disease causing possibly pathogenic disrupts tetrameric assembly; abolishes dbSNP: rs767122713 (Clinical lysosomal localization. significance: CLIN_pathogenic) Pathogenic variants found in the loop between the first and second transmembrane domain The substitution of the highly conserved (87-298aa) possibly reduce exon 7 SIFT: Deleterious c.785T>C missense amino acid. the stability of mucolipin- TM1-TM2 MutationTaster: disease no data p.(Phe262Ser) [no data] Large physicochemical difference between 1. Individuals with these loop causing Phe and Ser. pathogenic variants had a mild phenotype, an independent ataxic gait, and the ability to use their hands to feed themselves. This pathogenic variant HGMD accession: CM003598 (DM: was associated with mild Mucolipidosis IV) psychomotor involvement; ClinVar: RCV000727664.1 a slower progression of the The substitution of the highly conserved (Pathogenic - not provided), retinal disease and a missense amino acid. SIFT: Deleterious RCV000005442.7 (Likely pathogenic c.1084G>T exon 9 relatively mild neurologic [ALL:0.0008 Large physicochemical difference between MutationTaster: disease - Mucolipidosis type IV) p.(Asp362Tyr) TM3 phenotype, although NFE:0.0018] Asp and Tyr. Variant affects channel activity; causing Uniprot: this variant is reported as membrane preparations abolishes Fe(2+) permeability. possibly pathogenic containing mucolipin-1 dbSNP: rs121908372 (validated with this pathogenic dbSNP entry - Clinical significance: variant had no channel CLIN_likely_pathogenic) activity. Microdeletion of 3 bps TTC leads to the loss of residue Phe408, which is highly conserved HGMD accession: CD003635 (DM: inframe amino acid in protein domain Polycystin Mucolipidosis IV) c.1222_1224del exon 10 deletion Associated with the cation channel, PKD1/PKD2. Variant does not MutationTaster: disease dbSNP: rs797044817 (not validated p.(Phe408del) TM4 [ALL:0.0004 mildest MLIV phenotype affect channel activity; affects channel causing dbSNP entry - Clinical significance: NFE:0.0009] inhibition by low pH; still localizes to late CLIN_pathogenic) endosomes. The substitution of the highly conserved amino acid. exon 11 SIFT: Deleterious c.1256G>C missense Moderate physicochemical difference between TM4-TM5 MutationTaster: disease novel variant (this study) No data p.(Arg419Pro) [< 0.01] Arg and Pro. loop causing This variant is in protein domain: Polycystin cation channel, PKD1/PKD2. HGMD accession: CM026008 (DM: Mucolipidosis IV) The substitution of the highly conserved ClinVar: RCV000192526.1 amino acid. SIFT: Deleterious (Pathogenic - Mucolipidosis type IV) c.1340T>C exon 11 missense Associated with a more Moderate physicochemical difference between MutationTaster: disease Uniprot : this variant is reported as p.(Leu447Pro) TM5 [no data] severe MLIV Leu and Pro. causing possibly pathogenic This variant is in protein domain: Polycystin dbSNP: rs797044827 (not validated cation channel, PKD1/PKD2 dbSNP entry - Clinical significance: CLIN_pathogenic) SIFT: Deleterious MutationTaster: disease The substitution of the highly conserved causing exon 12 c.1367C>T missense amino acid. Large physicochemical difference Predicted change at acceptor Associated with a severe channel pore p.(Ser456Leu) [no data] between Ser and Leu. site 8 bps upstream: -20.5% MLIV loop This variant is in protein domain: Polycystin (MaxEnt: 0.0%, cation channel, PKD1/PKD2 NNSPLICE: -41.0%, SSF: 0.0%) Substitution affecting the HGMD accession: CS003630 (DM: splice site acceptor site in Atypical moderate form of Mucolipidosis IV) The substitution of the highly conserved intron 11 leading to frame mucolipidosis IV - exon 12 ClinVar: RCV000192302.1 c.1406A>G splice site amino acid. shift. The reading frame is affected individuals walk channel pore (Pathogenic - Mucolipidosis type IV). p.? [< 0.01] This variant is in protein domain: Polycystin interrupted by a premature independently and have loop, null rs797044818 (validated dbSNP entry - cation channel, PKD1/PKD2. STOP codon. The mRNA better communicative Clinical significance: produced might be targeted skills CLIN_pathogenic) for NMD. Microduplication of 11bps MutationTaster: disease ClinVar: RCV000193040.2 exon 12 frameshift GGCCGCAGCAG. This variation creates a c.1453_1463dup causing (Pathogenic - Mucolipidosis type IV) Associated with a severe channel pore duplication frame shift starting at codon Ser488. The new p.(Ser488Argfs*96) The mRNA produced might dbSNP: rs1207178149 (validated MLIV loop [< 0.01] reading frame ends in a STOP codon at be targeted for NMD. dbSNP entry) position 96.

AJ - Ashkenazi Jewish; DM - disease mutation, MLIV - mucolipidosis IV, n/a - not applicable, NMD - nonsense mediated decay , TM - transmembrane domain. 1 MAF - minor allele frequency in gnomAD public database (gnomad.broadinstitute.org) cluster ed with the major populations: [ALL: general population, AFR: African/African American, AMR - Admixed American, ASJ - Ashkenazi Jewish, OTH: other ancestry, NFE: Non-Finnish European, SAS: South Asian] 2 Molecular variants were assessed by pathogenicity prediction tools: MutationTaster (www.mutationtaster.org) and SIFT (sift.bii.a-star.edu.sg) for missense changes localized in coding sequence and MaxEnt, NNSPLICE or SSF for nucleotide changes identified in splice-site, respectively. 3 According database: The Human Gene Mutation Database (HGMD, www.hgmd.cf.ac.uk) , ClinVar aggregates information about genomic variation (ClinVar, www.ncbi.nlm.nih.gov), human single nucleotide variations (dbSNP, www.ncbi.nlm.nih.gov).