Cytoskeletal proteins in the cell nucleus influence brain development

The cytoskeleton gives cells their shape and helps them move. Researchers at Helmholtz Munich and the Ludwig Maximilian University now show that, in neural stem cells, proteins of the cytoskeleton are also found in the cell nucleus, where they can influence developmental programs. Using the protein MAP1B as an example, they explain how this previously underestimated connection may contribute to abnormal brain development. The results are published in the journal Cell.

During brain development, neural stem cells gradually give rise to specialized nerve cells. What matters is not only which cell types are formed, but also when they arise and how they then find their place in the developing tissue – for example in the cerebral cortex. Researchers have often studied these processes on two separate levels: in the cytoskeleton, the cell's internal scaffold that enables shape and movement, and in the nucleus, where genetic programs are regulated.

A team led by Prof. Magdalena Götz, Director of the Institute of Stem Cell Research at Helmholtz Munich and Professor at the Ludwig Maximilian University, now shows that this separation is too simple. In the nuclei of neural stem cells, the researchers found numerous proteins of the cytoskeleton. Surprisingly, these proteins were present in the nucleus in large numbers and appear to be involved in developmental programs. One of these proteins is MAP1B – a protein whose mutations are linked to developmental disorders of the brain.

The starting point of the study was a comprehensive analysis of proteins in neural stem cells. To do this, the researchers examined cell nuclei and cytoplasm separately – both in cells from the embryonic mouse brain and in human neural stem cells generated in the laboratory from reprogrammed body cells. "What surprised us was not that we found individual cytoskeletal proteins in the nucleus, but how many there were," says Dr. Florencia Merino, first author of the study and a PhD student in the Götz lab at the time the research was carried out. For further investigation, the team focused on the protein MAP1B. The reason: mutations in MAP1B had previously been described in patients with periventricular heterotopia – a developmental disorder in which some nerve cells in the brain are not located in the correct position.

To understand the role of MAP1B in neural stem cells, the team studied the function of the protein separately in the cytoplasm and in the nucleus. This revealed a clear contrast: in the cytoplasm, MAP1B promotes the differentiation of neural stem cells into nerve cells. In the nucleus, by contrast, MAP1B helps maintain the neural stem cell state for longer. "The function of MAP1B clearly depends on the cell compartment in which it is active," says Götz. "In the cytoplasm and in the nucleus, MAP1B binds to different protein complexes – and thereby influences different developmental programs."

The results change the view of periventricular heterotopia, in which some nerve cells are not located where they should be during brain development. Instead of migrating into the neuronal layers, they remain below them in the wrong place. Until now, the most obvious explanation was that the migration of these nerve cells was impaired. The new experiments, however, suggest that the abnormal development begins earlier – namely in the neural stem cells from which the nerve cells arise: if mutations or experimental interventions disrupt the function of MAP1B already at this stage, the cells maintain their stem cell program for longer. This leads to misspecification of some nerve cells that they generate: some migrate more slowly and do not reach the correct position.

Periventricular heterotopia is therefore not only a disorder of cell migration. Our results show that an early dysregulation of cell identity causes the disease."

Prof. Magdalena Götz, Director, Institute of Stem Cell Research at Helmholtz Munich and Professor, Ludwig Maximilian University

To examine the link with the disorder in more detail, the team also studied human cell models. The researchers first generated neural stem cells and then three-dimensional brain organoids – laboratory-grown models of early brain structures. These organoids carried mutations in the cytoskeletal protein MAP1B that are known from people with periventricular heterotopia. In the models, mutant MAP1B accumulated more strongly in the nucleus. At the same time, the researchers found nerve cells in the organoids in places where they should not occur during normal development. The models therefore reproduced key features of the disorder. "This supports our assumption that increased accumulation of MAP1B in the nucleus contributes to abnormal development," says Götz.

But what exactly does MAP1B change in the nucleus? The researchers found a clue in the so-called BAF protein complex. It influences which regions of the DNA are accessible – and therefore which genes can be read. MAP1B binds to this complex in the nucleus. In neural stem cells with disease-associated MAP1B mutations, the binding of a central component of BAF to the DNA changed: the complex was more strongly detected at regions close to genes involved in the neural stem cell state, cell movement and the cytoskeleton. As a result, genes involved in such developmental programs could be read at the wrong time or at altered levels. "For us, it was crucial that MAP1B is not merely present in the nucleus," says Merino. "There, it is connected to a molecular machinery that regulates developmental programs."

"Our results open up a broader view of the role of the cytoskeleton in cell development, beyond MAP1B," says Magdalena Götz. "They suggest that proteins of the cytoskeleton not only influence the shape and movement of cells, but may also be involved in regulating developmental programs in the nucleus."

Götz's team now aims to investigate whether other cytoskeleton-associated proteins have similar functions in the nucleus – and whether comparable mechanisms also play a role in other stem cells and developmental processes. In the long term, this new understanding could also help classify developmental disorders more precisely: not only according to where cells are ultimately located, but according to when and how their development is misregulated.