The neurodevelopmental transcriptome of theDrosophila melanogastermicrocephaly geneabnormal spindlereveals a role for temporal transcription factors and the immune system in regulating brain size

Abstract

ABSTRACTThe coordination of cellular behaviors during neurodevelopment is critical for determining the form, function, and size of the central nervous system. Mutations in the vertebrateAbnormal Spindle-Like, Microcephaly Associated (ASPM)gene and itsDrosophila melanogasterorthologabnormal spindle (asp)lead to microcephaly, a reduction in overall brain size whose etiology remains poorly defined. Here we provide the neurodevelopmental transcriptional landscape for aDrosophilamodel for autosomal recessive primary microcephaly (MCPH) and extend our findings into the functional realm in an attempt to identify the key cellular mechanisms responsible for Asp-dependent brain growth and development. We identify multiple transcriptomic signatures, including new patterns of co-expressed genes in the developing CNS. Defects in optic lobe neurogenesis were detected in larval brains through downregulation of temporal transcription factors (tTFs) and Notch signaling targets, which correlated with a significant reduction in brain size and total cell numbers during the neurogenic window of development. We also found inflammation as a hallmark ofaspMCPH brains, detectable throughout every stage of CNS development, which also contributes to the brain size phenotype. Finally, we show that apoptosis is not a primary driver of theaspMCPH phenotype, further highlighting an intrinsic Asp-dependent neurogenesis promotion mechanism that is independent of cell death. Collectively, our results suggest that the etiology ofaspMCPH is complex and that a comprehensive view of the cellular basis of the disorder requires an understanding of how multiple pathway inputs collectively determine the microcephaly phenotype.AUTHOR SUMMARYAutosomal recessive primary microcephaly (MCPH) is a neurodevelopmental disorder characterized by a reduction in brain size, intellect, and life span. Over 30 genes have been found mutated in human MCPH patients, withAbnormal Spindle-Like, Microcephaly Associated (ASPM)being the most common. Although the clinical aspects of the disorder are well-characterized, the underlying cellular and molecular mechanisms are not. The fruit fly,Drosophila melanogaster,has an ortholog of theASPMgene namedabnormal spindle (asp),and mutations also give rise to fruit flies with small brains. It had previously been suggested that mitotic spindle defects were responsible for theaspMCPH phenotype, preventing neural stem cells from dividing properly to generate the necessary number of neurons and glia in the brain. However, genetic studies in flies showed that this wasn’t the case, suggesting that our knowledge of MCPH remains incomplete and must be revised. In this manuscript, we identified new pathways important forasp-dependent brain growth through transcriptional profiling. We found a number of key pathways disrupted inaspmutants, which have not been described previously. Using genetic tools in the fly, we tested a subset of these pathways to identify their contributions to the brain growth defects seen inaspmutants. Our results add to the growing number of pathways necessary for brain growth control, and provide a suitable foundation for follow-up genetic studies to assess MCPH.