Figure Legends

Figure e-1. MRI of patient #11. A. sagittal T1-weighted and C. parasagittal T1-weighted images show hypoplastic inferior vermis associated with a small pons and thin corpus callosum.

B. Coronal inversion recovery image shows markedly hypoplastic cerebellum occupying the superior portion of the posterior cranial fossa, with dilatation of the cisterna magna inferiorly. D. axial T2-weighted image shows reduced size of the ponto-bulbar junction which is overall slightly hyperintense.

Figure e-2. MRI of the youngest sister, patient #12. A. sagittal and C. parasagittal T1- weighted images showing hypoplastic inferior vermis associated with a small pons and thin corpus callosum similarly to her eldest brother. B. Coronal inversion recovey image shows markedly hypoplastic cerebellum occupying the superior portion of the posterior cranial fossa particularly on the left side, with dilatation of the cisterna magna inferiorly. D. axial T2-weighted image shows reduced size of the pons which shows hyperintensity of the medial rafe.

Figure e-3. Mutations revealed in the TSEN54 gene. (A) Left: Electropherogram of the sequence flanking the c.919G>T detected in patient #1 as compared to a normal control (Ctrl).

Mutation is indicated by an arrow. Right: Rapid determination by PCR-restriction fragment length polymorphism analysis of the c.919G>T/p.A307S mutation in patient #1. Using oligonucleotide primers TSEN54-8F (5’-3’ GGA CGT GTG CAC CTT AGC A) and TSEN54-

8R (5’-3’- CCA GTT CAT CCA TGC CCT TT), we PCR-amplified a 751-base pairs (bp) fragment. The presence of the p.A307S mutation introduces a single site of cleavage for the endonuclease NspI (RCATG’Y) (New England Biolabs, Milan, Italy), resulting in fragments sized 402- and 349-bp (arrow heads). The wild-type sequence remains uncleaved whereas the heterozygote show fragments sized 751-, 402-, and 349-bp. Individuals correspond to the superimposed family tree. Ctrl, Control DNA; MW, 100-bp DNA molecular marker size. (B)

Electropherogram of the sequence flanking the novel c.1170_1183del mutation (arrow) detected in patient #10. The sequence of a normal control (Ctrl) is also shown.

Figure e-4 Haplotype reconstruction in seven Italian cases harboring the common c.919G>T/p.A307S mutation in the TSEN54 gene.

The genotype of polymorphic microsatellite markers scattered along chromosome 17q25 and

TSEN54 intragenic single nucleotide polymorphisms (boxed) is shown. The narrowed disease interval flanked by markers D17S1603 and D17S801 is also boxed. The location of the markers was obtained from the UniSTS database (http://www.ncbi.nlm.nih.gov/sites/entrez?db=unists).

Numbers refer to allele sizes in base pairs.

Figure e-5. Pedigree structure of the family of patients #11 & #12, and genotypes of the individuals tested on chromosome 7q11–21. Seven markers tested in the region of chromosome 7q11–21 are indicated. The marker order is cen-D7S502-D7S630-tel. Haplotypes were manually reconstructed to assure the minimal recombination. The PCH3-associated region is also indicated3.