The Liberfarb syndrome, a multisystem disorder affecting eye, ear, bone, and brain development, is caused by a founder pathogenic variant in the PISD gene

We observed four individuals in two unrelated but consanguineous families from Portugal and Brazil affected by early-onset retinal degeneration, sensorineural hearing loss, microcephaly, intellectual disability, and skeletaldysplasia with scoliosis and short stature. The phenotype precisely matched that of an individual of Azorean descent published in 1986 by Liberfarb andcoworkers. Methods: Patients underwent specialized clinical examinations (including ophthalmological, audiological, orthopedic, radiological, and developmental assessment). Exome and targeted sequencing was performed on selected individuals. Minigene constructs were assessed by quantitative polymerase chainreaction (qPCR) and Sanger sequencing. Results: Affected individuals shared a 3.36-Mb region of autozygosity onchromosome 22q12.2, including a 10-bp deletion (NM_014338.3:c.904-12_904-3delCTATCACCAC), immediately upstream of the last exon of the PISD (phosphatidylserinedecarboxylase) gene. Sequencing of PISD from paraffin-embedded tissue from the 1986 case revealed the identical homozygous variant. In HEK293T cells, this variant led to aberrant splicing of PISD transcripts. Conclusion: We have identified the genetic etiology of the Liberfarb syndrome, affecting brain, eye, ear, bone, and connective tissue. Our work documents the migration of a rare Portuguese founder variant to two continents and highlights the link between phospholipid metabolism and bone formation, sensory defects,and cerebral development, while raising the possibility of therapeutic phospholipid replacement

Summary information for each of the germline and somatic mutations found in <i>MAP3K6</i>.

a<p>The H506Y mutation is a somatic second-hit observed in FFPE tumor tissue from patient 1884.</p><p>Summary information for each of the germline and somatic mutations found in <i>MAP3K6</i>.</p

Pedigree of the Maritime Canadian family.

<p>Clinically affected individuals are indicated with shaded symbols. Individuals for whom DNA samples were collected are indicated by a number. Individuals shaded within 2 quarter sections were affected but were negative for the MAP3K6 mutation. Individuals shaded on one half had another, non-gastric, cancer. Generations I–VI are indicated.</p

Regions with LOD>0.5 from parametric linkage analyses using Merlin when individual 2447 is treated as unknown (sub-pedigree).

<p>Genomic intervals and associated LOD scores are shown under dominant models with 50% or 99% penetrance. Regions are defined by their 1-LOD support interval. Base pair positions are from hg19. An asterisk indicates the SNP was the first or last analyzed marker on the chromosome.</p><p>Regions with LOD>0.5 from parametric linkage analyses using Merlin when individual 2447 is treated as unknown (sub-pedigree).</p

Regions with LOD>1 from pedigree-wide parametric linkage analyses using Merlin.

<p>Genomic intervals and associated LOD scores are shown under dominant models with 50% or 99% penetrance. Regions are defined by their 1-LOD support interval. Base pair positions are from hg19. An asterisk indicates the SNP was the first or last analyzed marker on the chromosome.</p><p>Regions with LOD>1 from pedigree-wide parametric linkage analyses using Merlin.</p

A) Pedigree of the Portuguese Familial Gastric Cancer family.

<p>Affected individuals are shaded in black with the sequenced proband indicated with a triangle. Deceased individuals are marked with a strike-through. Generations I–III are indicated. <b>B</b>) Tumor cells showing signet ring cell morphology (H&E, 200×). <b>C</b>) Tumor cells retaining E-cadherin protein expression (IHC analysis performed with the rabbit anti-E-cadherin Antibody (24E10 Cell Signaling, MA, USA), according to manufacturer's instructions, 200×).</p

Summary of SNVs observed in the Maritime sub-pedigree (Proband, Mother, Maternal Aunts) and in 115 probands from an additional screened cohort, including a frameshift-truncating mutation Portuguese pedigree (F849Sfs*142).

<p>All SNVs are shown in relation to the predicted functional domains of <i>MAP3K6</i>.</p

Methylation analysis of the Portuguese family.

<p>Left panel: Schematic representation of the MAP3K6 gene adapted from Ensembl genome browser (release 75). The two CpG islands analyzed are represented. CpG island 1 is mainly non-methylated for several normal tissues and cells lines represented in the scheme, while CpG island 2 displays low methylation frequency (light green) in normal tissues such as B-cells, Colon, Liver and Whole Blood, and high methylation (blue) in colon (HCT116), liver (HepG2) and blood (Jurkat) cancer cell lines. A DNase HSS predicted to harbor a promoter-associated regulatory element overlapping with CpG island 2. Right panel: For the CpG island 1, no hypermethylation was detected (white circles). For the CpG island 2, we observed complete methylation in the proband's tumor DNA (black circles) and no methylation in the PBLs' DNA. The DNA of normal gastric mucosa from controls displayed partial methylation (grey circles). All gastric cancer cell lines mimicked the full methylation observed for the tumor DNA (black circles).</p

Histology of two gastric cancer patients from the Maritime Canadian family.

<p><b>A</b>) Patient 1826. Section shows a tumor composed of solid and glandular components (H&E, 40×). <b>B</b>) The tumor cells show signet ring cell morphology in solid areas (H&E, 200×). <b>C</b>) Patient 1884. Section shows sheet of tumor cells infiltrated lamina propria and submucosa (H&E, 40×). <b>D</b>) The tumor entirely consists of signet ring cells (H&E 200×).</p

Predicted pathogenicity of germline and somatic variants in <i>MAP3K6</i> observed in a Maritime Canadian family and probands from a screen of 115 FGC cases negative for <i>CDH1</i> mutations.

<p>* FATHMM predictions for Missense Variants and Cancer-Associated Variants tools are both given.</p>a<p>The H506Y mutation is a somatic second-hit observed in FFPE tumor tissue from patient 1884.</p><p>Predicted pathogenicity of germline and somatic variants in <i>MAP3K6</i> observed in a Maritime Canadian family and probands from a screen of 115 FGC cases negative for <i>CDH1</i> mutations.</p