*Result*: Cerebral organoids expressing mutant actin genes reveal cellular mechanism underlying microcephaly.

*Actins are cytoskeletal proteins that are essential for multiple cellular processes. Mutations in the ACTB and ACTG1 genes, encoding the ubiquitous beta- and gamma-cytoskeletal actin isoforms, respectively, cause a broad spectrum of neurodevelopmental disorders, with microcephaly as the most frequent one. To investigate the pathogenesis underlying this cortical malformation, we studied patient-derived cerebral organoids from induced pluripotent stem cells of individuals with the Baraitser–Winter-CerebroFrontoFacial syndrome (BWCFF-S) carrying an ACTB/ACTG1 missense mutation. These organoids were reduced in size, showing a thinner ventricular zone (VZ) due to reduced VZ progenitor abundance. Strikingly, VZ progenitors in BWCFF-S cerebral organoids displayed a shift in the orientation of their cleavage plane from a predominantly vertical to a majoritarian horizontal orientation. The latter cleavage plane orientation is incompatible with increasing VZ progenitor abundance and instead promotes basal progenitor generation. Various cytoskeletal and morphological irregularities of BWCFF-S VZ progenitors, notably in the apical region, seemingly contribute to this change in cleavage plane orientation. Our results provide insight into the cell biological basis of the microcephaly associated with BWCFF-S caused by actin mutations. Synopsis: BWCFF-S cerebral organoids mimic microcephaly with fewer VZ progenitors due to increased delamination and elevated basal progenitor output. This is likely caused by altered mitotic spindle orientation, possibly via cytoskeletal and structural changes. Patient-derived and CRISPR/Cas9-generated Baraitser-Winter-CerebroFrontoFacial Syndrome (BWCFF-S) cerebral organoids recapitulate BWCFF-S-associated microcephaly. BWCFF-S cerebral organoids exhibit a reduced number of VZ progenitors due to increased delamination, which leads to aberrant basal progenitor output. This increased delamination is likely caused by a change in cleavage plane orientation in mitotic anaphase VZ progenitors from a predominantly vertical to a nearly randomized or majoritarian horizontal orientation. This altered cleavage plane orientation is likely caused by cytoskeletal and morphological irregularities in VZ progenitors of BWCFF-S cerebral organoids. Baraitser-Winter-CerebroFrontoFacial Syndrome (BWCFF-S) cerebral organoids mimic microcephaly with fewer ventricular zone progenitors. [ABSTRACT FROM AUTHOR]

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