Mutations in microcephalin cause aberrant regulation of chromosome condensation

Abstract

Microcephalin (MCPH1) is a gene mutated in primary microcephaly, an autosomal recessive neurodevelopmental disorder in which there is a marked reduction in brain size. PCC syndrome is a recently described disorder of microcephaly, short stature, and misregulated chromosome condensation. Here, we report the finding that MCPH1 primary microcephaly and PCC syndrome are allelic disorders, both having mutations in the MCPH1 gene. The two conditions share a common cellular phenotype of premature chromosome condensation in the early G2 phase of the cell cycle, which, therefore, appears to be a useful diagnostic marker for individuals with MCPH1 gene mutations. We demonstrate that an siRNA-mediated depletion of MCPH1 is sufficient to reproduce this phenotype and also show that MCPH1-deficient cells exhibit delayed decondensation postmitosis. These findings implicate microcephalin as a novel regulator of chromosome condensation and link the apparently disparate fields of neurogenesis and chromosome biology. Further characterization of MCPH1 is thus likely to lead to fundamental insights into both the regulation of chromosome condensation and neurodevelopment.

Figure 1

Depiction of the 64-cM homozygous region, identified by a genomewide search, in consanguineous siblings with PCC syndrome. The region of homozygous markers is boxed. Maximum two-point LOD scores of 1.37 (θ=0) were obtained for markers D8S264, D8S277, D8S550, and D8S560.

Figure 2

Findings indicating that MCPH1 primary microcephaly and PCC syndrome are allelic disorders. Patients with MCPH1 primary microcephaly (black diamonds) and patients with PCC syndrome (unblackened squares) have a similar degree of microcephaly, and both groups also exhibit short stature of varying severity. a, Head circumference (OFC), height, and weight, plotted as SDs relative to the appropriate age- and sex-related population mean. b and c, Neuroimaging in MCPH1 primary microcephaly, demonstrating a small but structurally normal brain. In PCC syndrome, however, there is more marked gyral simplification resembling pachygyria (d), and periventricular neuronal heterotopias are also present (e). Subjects include an adult with MCPH1 primary microcephaly (b, T1W sagittal; c, axial T2W) and an 18-mo–old child with PCC syndrome (d, axial T2W; e, axial T1W and T2W images of lateral ventricles). f–h, MRI scans of normal subjects age matched to those of images b–d, respectively. The scale bar is 1 cm.

Figure 3

Premature chromosome condensation, present in both MCPH1 primary microcephaly and PCC syndrome. Shown are prophase-like cells in peripheral blood lymphocytes of patients with MCPH1 primary microcephaly (a) and patients with PCC syndrome (b).

Figure 4

Homozygous ins427A mutation (arrow) of the MCPH1 gene, which is present in patients with PCC syndrome.

Figure 5

The siRNA-mediated depletion of MCPH1 is sufficient to cause premature chromosome condensation. a, Prophase-like phenotype observed in HeLa cells after siRNA-mediated depletion of MCPH1. A marked increase in prophase cells is seen after MCPH1 siRNA transfection in HeLa and LN9SV fibroblasts (b), whereas there is no significant change in mitotic index (% metaphases) (c).

Figure 6

Delayed decondensation observed in MCPH1-deficient cells. a, Binucleate MCPH1 mutant cell with condensed chromosomes indicating a delay in decondensation. b, Binucleate control lymphoblastoid cell with decondensed chromatin. c, Proportion of cells with condensed chromosomes postmitosis in patient and control cell lines. Many cytochalasin-treated binucleate G1-phase PCC (29%) and MCPH1 (36%) mutant cells show condensed chromatin, compared with control lymphoblastoid cells (4%).