X-Linked Congenital Bilateral Absence of Vas Deferens

Transcription

1 The American Journal of Human Genetics, Volume 99 Supplemental Data Truncating Mutations in the Adhesion G Protein- Coupled Receptor G2 Gene ADGRG2 Cause an X-Linked Congenital Bilateral Absence of Vas Deferens Olivier Patat, Adrien Pagin, Aurore Siegfried, Valérie Mitchell, Nicolas Chassaing, Stanislas Faguer, Laetitia Monteil, Véronique Gaston, Louis Bujan, Monique Courtade-Saïdi, François Marcelli, Guy Lalau, Jean-Marc Rigot, Roger Mieusset, and Eric Bieth

2 Supplemental Tables: Table S1: Inclusion and exclusion criteria. CFTR Genotype: Inclusion criteria CBAVD Phenotype Available CFTR mutation screening results Individual addressed to the Hospital Centers of Toulouse or Lille, France, for a CBAVD Sufficient amount of DNA to perform complementary analyses defined by the total absence of sperm cells on microscopic analysis of centrifuged complete semen Sequencing of the 27 exons of CFTR Morphological description of the scrotal and intra-abdominal portions of the vasa deferentia (a) When technically possible, search for intragenic rearrangement by a semi quantitative method (b) CFTR Genotype: At least one pathogenic mutation in CFTR in a heterozygous or homozygous state c t[5] allele in a homozygous state regardless of the c tg(n) alleles present c t[5] allele in a heterozygous state in cis with the c tg[12] or c tg[13] allele Exclusion criteria: Proved bilateral absence of vasa deferentia either complete or incomplete CBAVD Phenotype Absence of kidney ultra-sound examination Any kidney abnormality apart from unilateral agenesis Suspicion of an acquired cause of absence of the vasa deferentia CBAVD: congenital bilateral absence of vas deferens; (a): See table S2; (b): Multiplex ligation dependent probe amplification MLPA or quantitative multiplex PCR on short fragments QMPSF

3 Table S2: Phenotypic evaluation criteria of the CBAVD subjects. Familial medical history of infertility Personal medical history Infectious medical history (STD, mumps, epididymitis ) Cryptorchidism Surgery (cryptorchidism, inguinal hernia, scrotal incision ) Occupation Toxic/heat exposure Clinical examination Testis volume & position Epididymis volume & position Hydrocele, varicocele, cyst Vas deferens Scrotal description Testis volume Caput Peroperative Epididymis Corpus description Vas deferens Cauda Permeability test Testis volume Caput Scrotal ultra-sound Epididymis Corpus Vas deferens Cauda Intra Abdominal description (a) Vas deferens Deferential bulb Ejaculatory duct Seminal vesicle Renal phenotype Renal ultra-sound Semen analyses Volume ph Sperm count Seminal L-Carnitine Seminal glycerophosphocholine Seminal alpha-glucosidase Fructose Seminal acyl-carnitine Hormonal tests LH FSH Testosterone Inhibin (a) Trans Rectal Ultra Sound results

4 Table S3: Phenotypic description of the 12 individuals with Congenital Bilateral Absence of Vas Deferens without kidney abnormality T1 Scrotal (a) Epididymis (c) Cap Cor Cau Vas deferens Intra Abdominal (b) Vas Deferential deferens bulb T2 T3 T4 T5 R: segmental absence L: segmental absence L1 L2 L3 L4 L5 L6 L7

5 T1 Intra Abdominal (b) Ejaculatory Seminal duct Vesicle (d) R: hypoplasia Testis (e) Semen (f) L: hypoplasia, acid ph T2 T3 T4 T5 L1 L2 R: hypoplasia L3 L4 L5 L6 R: hypoplasia L: hypoplasia L7 R: hypoplasia L: hypoplasia L: hypotrophy R: hypotrophy L: hypotrophy R: hypotrophy,, acid ph acid ph,, acid ph, R = right side; L = left side; = not done; (a): surgically observed and/or by scrotal ultrasonography and clinical examination; (b): observed by transrectal ultrasonography; (c): Cap = caput, Cor = corpus, Cau = cauda of the epididymis; (d): hypoplasia = present seminal vesicle with reduced size; (e): hypotrophy = testis volume < 13 ml; (f): = low semen volume (< 1.5 ml); acid ph = semen ph < 7.2. Individuals are anonymously referred to as T1, T2,, L7.

6 Table S4: Number of genes carrying one variant in a heterozygous state in at least two individuals. Number of individals Before standard filtering After standard filtering (a) With LoF variants Restricted/strong expression in male genital tract (b) (a) Standard filtering of variants predicted as benign by PolyPhen2, variants with allelic frequency over 1% in EVS, HapMap, dbsnp, 1000genomes and the in-house database of IntegraGen, variants non covered by at least one read, depth <10X. LoF: loss-of-function (nonsense, frameshift or canonic splice site variants) (b) The expression pattern was analyzed using the database Human Protein Atlas and data of the literature. Table S5: Number of genes carrying one variant in a homozygous state in at least two individuals. Number of individuals Before standard filtering After standard filtering (a) With LoF variants Restricted/strong expression in male genital tract (b) (a) Standard filtering of variants predicted as benign by PolyPhen2, variants with allelic frequency over 1% in EVS, HapMap, dbsnp, 1000genomes and the in-house database of IntegraGen, variants non covered by at least one read, depth <10X. LoF: loss-of-function (nonsense, frameshift or canonic splice site variants) (b) The expression pattern was analyzed using the database Human Protein Atlas and data of the literature..

7 Table S6: Number of genes carrying two variants in a heterozygous state in at least two individuals. Number of individuals 2 3 Before standard filtering After standard filtering (a) 10 1 With LoF variants 0 0 Restricted/strong expression in male genital tract (b) 2 0 (a) Standard filtering of variants predicted as benign by PolyPhen2, variants with allelic frequency over 1% in EVS, HapMap, dbsnp, 1000genomes and the in-house database of IntegraGen, variants non covered by at least one read, depth <10X. LoF: loss-of-function (nonsense, frameshift or canonic splice site variants) (b) The expression pattern was analyzed using the database Human Protein Atlas and data of the literature. Table S7: Number of X-linked genes carrying one variants in a hemizygous state in at least two individuals. Number of individuals Before standard filtering After standard filtering (a) With LoF variants Restricted/strong expression in male genital tract (b) 1 (c) 1 (c) 0 0 (a) Standard filtering of variants predicted as benign by PolyPhen2, variants with allelic frequency over 1% in EVS, HapMap, dbsnp, 1000genomes and the in-house database of IntegraGen, variants non covered by at least one read, depth <10X. LoF: loss-of-function (nonsense, frameshift or canonic splice site variants) (b) The expression pattern was analyzed using the database Human Protein Atlas and data of the literature. (c) ADGRG2

8 Table S8: List of genes carrying loss-of-function mutations in at least 3 individuals of the WES cohort, by mode of inheritance. Gene Number of individuals carrying mutations 1 heterozygous mutation CHADL 5 TYRO3 4 BAZ2A 4 C4orf21 3 CLTCL1 3 FRG1 3 1 hemizygous mutation (X-linked) NUDT11 12 ADGRG2 3 1 homozygous mutations GALC 12 MAML3 12 C8orf59 12 DDHD1 3 PCSK6 3