Differential Song Deficits after Lentivirus-Mediated Knockdown of FoxP1, FoxP2, or FoxP4 in Area X of Juvenile Zebra Finches

Abstract

Mutations in the transcription factors FOXP1 and FOXP2 are associated with speech impairments. FOXP1 is additionally linked to cognitive deficits, as is FOXP4. These FoxP proteins are highly conserved in vertebrates and expressed in comparable brain regions, including the striatum. In male zebra finches, experimental manipulation of FoxP2 in Area X, a striatal song nucleus essential for vocal production learning, affects song development, adult song production, dendritic spine density, and dopamine-regulated synaptic transmission of striatal neurons. We previously showed that, in the majority of Area X neurons FoxP1, FoxP2, and FoxP4 are coexpressed, can dimerize and multimerize with each other and differentially regulate the expression of target genes. These findings raise the possibility that FoxP1, FoxP2, and FoxP4 (FoxP1/2/4) affect neural function differently and in turn vocal learning. To address this directly, we downregulated FoxP1 or FoxP4 in Area X of juvenile zebra finches and compared the resulting song phenotypes with the previously described inaccurate and incomplete song learning after FoxP2 knockdown. We found that experimental downregulation of FoxP1 and FoxP4 led to impaired song learning with partly similar features as those reported for FoxP2 knockdowns. However, there were also specific differences between the groups, leading us to suggest that specific features of the song are differentially impacted by developmental manipulations of FoxP1/2/4 expression in Area X.SIGNIFICANCE STATEMENT We compared the effects of experimentally reduced expression of the transcription factors FoxP1, FoxP2, and FoxP4 in a striatal song nucleus, Area X, on vocal production learning in juvenile male zebra finches. We show, for the first time, that these temporally and spatially precise manipulations of the three FoxPs affect spectral and temporal song features differentially. This is important because it raises the possibility that the different FoxPs control different aspects of vocal learning through combinatorial gene expression or by acting in different microcircuits within Area X. These results are consistent with the deleterious effects of human FOXP1 and FOXP2 mutations on speech and language and add FOXP4 as a possible candidate gene for vocal disorders.

Keywords: CAS; FoxP1; FoxP2; FoxP4; developmental verbal dyspraxia; song learning.