Genetic variation in MKL2 and decreased downstream PCTAIRE1 expression in extreme, fatal primary human microcephaly.

The genetic mechanisms driving normal brain development remain largely unknown. We performed genomic and immunohistochemical characterization of a novel, fatal human phenotype including extreme microcephaly with cerebral growth arrest at 14-18 weeks gestation in three full sisters born to healthy, non-consanguineous parents. Analysis of index cases and parents included familial exome sequencing, karyotyping, and genome-wide single nucleotide polymorphism (SNP) array. From proband, control and unrelated microcephalic fetal cortical tissue, we compared gene expression of RNA and targeted immunohistochemistry. Each daughter was homozygous for a rare, non-synonymous, deleterious variant in the MKL2 gene and heterozygous for a private 185 kb deletion on the paternal allele, upstream and in cis with his MKL2 variant allele, eliminating 24 CArG transcription factor binding sites and MIR4718. MKL1 was underexpressed in probands. Dysfunction of MKL2 and its transcriptional coactivation partner, serum response factor (SRF), was supported by a decrease in gene and protein expression of PCTAIRE1, a downstream target of MKL2:SRF heterodimer transcriptional activation, previously shown to result in severe microcephaly in murine models. While disruption of the MKL2:SRF axis has been associated with severe microcephaly and disordered brain development in multiple model systems, the role of this transcription factor complex has not been previously demonstrated in human brain development.

Two novel translocation breakpoints upstream of SOX9 define borders of the proximal and distal breakpoint cluster region in campomelic dysplasia.

The semilethal skeletal malformation syndrome campomelic dysplasia (CD) with or without XY sex reversal is caused by mutations within the SOX9 gene on 17q24.3 or by chromosomal aberrations (translocations, inversions or deletions) with breakpoints outside the SOX9 coding region. The previously published CD translocation breakpoints upstream of SOX9 fall into two clusters: a proximal cluster with breakpoints between 50-300 kb and a distal cluster with breakpoints between 899-932 kb. Here, we present clinical, cytogenetic and molecular data from two novel CD translocation cases. Case 1 with karyotype 46,XY,t(1;17)(q42.1;q24.3) has characteristic symptoms of CD, including mild tibial bowing, cryptorchidism and hypospadias. By standard fluorescence in situ hybridization (FISH) and by high-resolution fiber FISH, the 17q breakpoint was mapped 375 kb from SOX9, defining the centromeric border of the proximal breakpoint cluster region. Case 2 with karyotype 46,X,t(Y;17)(q11.2;q24.3) has the acampomelic form of CD and complete XY sex reversal. By FISH and somatic cell hybrid analysis, the 17q breakpoint was mapped 789 kb from SOX9, defining the telomeric border of the distal breakpoint cluster region. We discuss the structure of the 1 Mb cis-control region upstream of SOX9 and the correlation between the position of the 14 mapped translocation breakpoints with respect to disease severity and XY sex reversal.