Dataset Information

AGS3 and G?i3 Are Concomitantly Upregulated as Part of the Spindle Orientation Complex during Differentiation of Human Neural Progenitor Cells.

ABSTRACT: Adult neurogenesis is modulated by many Gi-coupled receptors but the precise mechanism remains elusive. A key step for maintaining the population of neural stem cells in the adult is asymmetric cell division (ACD), a process which entails the formation of two evolutionarily conserved protein complexes that establish the cell polarity and spindle orientation. Since ACD is extremely difficult to monitor in stratified tissues such as the vertebrate brain, we employed human neural progenitor cell lines to examine the regulation of the polarity and spindle orientation complexes during neuronal differentiation. Several components of the spindle orientation complex, but not those of the polarity complex, were upregulated upon differentiation of ENStem-A and ReNcell VM neural progenitor cells. Increased expression of nuclear mitotic apparatus (NuMA), G?i subunit, and activators of G protein signaling (AGS3 and LGN) coincided with the appearance of a neuronal marker (?-III tubulin) and the concomitant loss of neural progenitor cell markers (nestin and Sox-2). Co-immunoprecipitation assays demonstrated that both G?i3 and NuMA were associated with AGS3 in differentiated ENStem-A cells. Interestingly, AGS3 appeared to preferentially interact with G?i3 in ENStem-A cells, and this specificity for G?i3 was recapitulated in co-immunoprecipitation experiments using HEK293 cells transiently overexpressing GST-tagged AGS3 and different G?i subunits. Moreover, the binding of G?i3 to AGS3 was suppressed by GTP?S and pertussis toxin. Disruption of AGS3/G?i3 interaction by pertussis toxin indicates that AGS3 may recognize the same site on the G? subunit as G protein-coupled receptors. Regulatory mechanisms controlling the formation of spindle orientation complex may provide novel means to manipulate ACD which in turn may have an impact on neurogenesis.

SUBMITTER: Yip JLK

PROVIDER: S-EPMC7664263 | biostudies-literature | 2020 Nov

REPOSITORIES: biostudies-literature

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Publications

AGS3 and Gα<sub>i3</sub> Are Concomitantly Upregulated as Part of the Spindle Orientation Complex during Differentiation of Human Neural Progenitor Cells.

Yip Jackson L K JLK Lee Maggie M K MMK Leung Crystal C Y CCY Tse Man K MK Cheung Annie S T AST Wong Yung H YH

Molecules (Basel, Switzerland) 20201106 21

Adult neurogenesis is modulated by many G<sub>i</sub>-coupled receptors but the precise mechanism remains elusive. A key step for maintaining the population of neural stem cells in the adult is asymmetric cell division (ACD), a process which entails the formation of two evolutionarily conserved protein complexes that establish the cell polarity and spindle orientation. Since ACD is extremely difficult to monitor in stratified tissues such as the vertebrate brain, we employed human neural progeni ...[more]

PMID: 33172018

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Project description:BackgroundMutations in the Abnormal Spindle Microcephaly related gene (ASPM) are the commonest cause of autosomal recessive primary microcephaly (MCPH) a disorder characterised by a small brain and associated mental retardation. ASPM encodes a mitotic spindle pole associated protein. It is suggested that the MCPH phenotype arises from proliferation defects in neural progenitor cells (NPC).ResultsWe show that ASPM is a microtubule minus end-associated protein that is recruited in a microtubule-dependent manner to the pericentriolar matrix (PCM) at the spindle poles during mitosis. ASPM siRNA reduces ASPM protein at the spindle poles in cultured U2OS cells and severely perturbs a number of aspects of mitosis, including the orientation of the mitotic spindle, the main determinant of developmental asymmetrical cell division. The majority of ASPM depleted mitotic cells fail to complete cytokinesis. In MCPH patient fibroblasts we show that a pathogenic ASPM splice site mutation results in the expression of a novel variant protein lacking a tripeptide motif, a minimal alteration that correlates with a dramatic decrease in ASPM spindle pole localisation. Moreover, expression of dominant-negative ASPM C-terminal fragments cause severe spindle assembly defects and cytokinesis failure in cultured cells.ConclusionsThese observations indicate that ASPM participates in spindle organisation, spindle positioning and cytokinesis in all dividing cells and that the extreme C-terminus of the protein is required for ASPM localisation and function. Our data supports the hypothesis that the MCPH phenotype caused by ASPM mutation is a consequence of mitotic aberrations during neurogenesis. We propose the effects of ASPM mutation are tolerated in somatic cells but have profound consequences for the symmetrical division of NPCs, due to the unusual morphology of these cells. This antagonises the early expansion of the progenitor pool that underpins cortical neurogenesis, causing the MCPH phenotype.

Dataset Information

AGS3 and G?i3 Are Concomitantly Upregulated as Part of the Spindle Orientation Complex during Differentiation of Human Neural Progenitor Cells.

Publications

AGS3 and Gα<sub>i3</sub> Are Concomitantly Upregulated as Part of the Spindle Orientation Complex during Differentiation of Human Neural Progenitor Cells.

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