The cerebellum, situated at the rear of the skull, has long been recognised for its role in coordinating muscle movements. Recent research, however, indicates it also contributes to higher‑order cognitive processes. The Purkinje cells are the sole neurons within the cerebellar cortex that integrate incoming signals and transmit them to other brain regions.
These Purkinje cells are large, highly branched neurons that can assume a range of functions. Although many studies have examined their roles, the precise neural and genetic mechanisms underlying their diversity are still not fully understood.
Scientists from the University of Connecticut School of Medicine conducted a study to determine whether FOXP genes—a family known to regulate the activation and repression of other genes—affect the composition of Purkinje cells and the wiring of cerebellar circuits. Results published in Nature Neuroscience reveal at least eleven distinct Purkinje cell subtypes, implicating FOXP1 and FOXP2 in their diversification.
The FOXP family comprises four genes (FOXP1‑FOXP4). Alterations in FOXP1 and FOXP2 have previously been associated with neurodevelopmental and language disorders.
“FOXP2 is essential for vocalisation across species, supporting song learning in birds, echolocation in bats, pup calling in mice, and proper speech in humans,” senior author James Y.H. Li told Medical Xpress.
“Mutations in FOXP1 produce a syndrome marked by delayed development, language impairment, intellectual disability, and autism‑like traits. Despite their clinical relevance, the functions of FOXP genes during brain development remain poorly defined, so we set out to fill this gap.”
Previous work by Li and colleagues demonstrated that Foxp1 and Foxp2 are expressed at different levels in cerebellar Purkinje cells. The new study aimed to assess how conditional deletion of these genes from the cerebellum influences mouse development and behaviour.
“Using single‑cell RNA sequencing (scRNA‑seq) we profiled gene expression within individual embryonic mouse cerebellar cells at key developmental stages,” Li explained. “This revealed multiple molecularly distinct Purkinje cell subtypes. Because scRNA‑seq lacks spatial information, we developed scANKRS (single‑cell Anchoring Network of Key Regulators to Space), which remaps the scRNA‑seq‑identified groups back into their native tissue context.”
To map FOXP expression, the team combined scANKRS with 3‑D light‑sheet fluorescence imaging. The integration allowed direct overlay of Foxp1, Foxp2, and Foxp4 patterns onto the emerging Purkinje cell subtypes.
The researchers discovered that deleting Foxp1 or Foxp2 from the mouse cerebellum impaired vocalisation, pointing to a potential cerebellar role in speech and vocalisation.
“We also found that Purkinje cells are not all uniform but comprise several subtypes with distinct molecular signatures, each occupying specific regions of the cerebellar cortex,” Li said. “Because they are the primary output neurons of the cortex, this diversity underpins functional specialisation across cortical areas.”
The experiments also illuminated the mechanisms that may govern cerebellar hemisphere formation. These bilateral structures are particularly large in primates and are associated with advanced motor and cognitive abilities.
The deletion of Foxp1 and Foxp2 prevented the development of the two hemispheres in mice, providing the first direct genetic evidence linking hemisphere formation to specific molecular pathways.
“In addition, Foxp1‑positive Purkinje cells—the subtype most affected by the knockouts—are plentiful in the human foetal cerebellum but rare in birds,” Li noted. “Their presence may have driven the evolutionary expansion of mammalian cerebellar hemispheres, enabling higher cognitive functions.”
Overall, the study supplies strong evidence that FOXP genes contribute to the diversification of Purkinje cell subtypes and to the establishment of connections with other brain regions. If similar patterns are confirmed in primates and humans, these findings could shed light on the links between certain neurodevelopmental disorders, such as autism spectrum disorder, and FOXP deficiencies.
“Our next steps will be to delineate the molecular mechanisms of FOXP transcription factors—their binding partners, target genes, and regulatory networks,” Li added. “The recognition of Purkinje cell diversity now allows us to test how perturbing specific subtypes, especially Foxp1‑positive cells, influences cerebellar hemisphere expansion.”
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