Normalizing translation through 4E-BP prevents mTOR-driven cortical mislamination and ameliorates aberrant neuron integration

Abstract

Hyperactive mammalian target of rapamycin complex 1 (mTORC1) is a shared molecular hallmark in several neurodevelopmental disorders characterized by abnormal brain cytoarchitecture. The mechanisms downstream of mTORC1 that are responsible for these defects remain unclear. We show that focally increasing mTORC1 activity during late corticogenesis leads to ectopic placement of upper-layer cortical neurons that does not require altered signaling in radial glia and is accompanied by changes in layer-specific molecular identity. Importantly, we found that decreasing cap-dependent translation by expressing a constitutively active mutant of the translational repressor eukaryotic initiation factor 4E-binding protein 1 (4E-BP1) prevents neuronal misplacement and soma enlargement, while partially rescuing dendritic hypertrophy induced by hyperactive mTORC1. Furthermore, overactivation of translation alone through knockdown of 4E-BP2 was sufficient to induce neuronal misplacement. These data show that many aspects of abnormal brain cytoarchitecture can be prevented by manipulating a single intracellular process downstream of mTORC1, cap-dependent translation.veractive mammalian target of rapamycin complex 1 (mTORC1) signaling is a signature of many disorders with cortical malformations (1), ranging from tuberous sclerosis complex with focal dysplasias to hemimegalencephaly with more diffuse, hemispheric aberrations. The high incidence of negative outcomes in individuals with such malformations (2), which are often associated with intractable childhood seizures, underscores the need to better understand the molecular etiology of these developmental lesions. Animal models have demonstrated the causative effect of increased mTORC1 signaling on mislamination (3-7). The pharmacological mTORC1 blocker rapamycin has also been shown to reverse some of the developmental abnormalities and associated seizure activity in several of these mouse models (5, 6, 8-10), further emphasizing the importance of mTORC1 in the disease pathogenesis.Despite the demonstrated relevance of mTORC1 signaling, there is less known about the molecular mechanisms by which mTORC1 alters cortical development. Addressing this question is complicated by the wide range of cellular processes regulated by mTORC1 through independent downstream targets. Among these regulated processes are autophagy, lysosomal function, lipid synthesis, and, one of the best-studied functions, cap-dependent translation (11). Because current drugs that suppress mTORC1 activity can have serious side effects (12, 13) and do not fully block some of mTORC1's functions (14), a more specific understanding of how mTORC1 contributes to cortical mislamination could yield better targets for treatment.This study therefore aimed to more closely characterize the cytoarchitectural aberrations generated by hyperactive mTORC1 and to examine the contribution of translational regulation to these cortical malformations. Using in utero electroporation, we generated and characterized focal mislamination an...

Hypervascularization in mTOR‐dependent focal and global cortical malformations displays differential rapamycin sensitivity

“…28,34 In this model, a focal MCD in the somatosensory cortex did not lead to seizures while a focal MCD in the medial prefrontal cortex led to seizures that were visible starting at P21. In addition, using the transgenic Tsc1 mice with global cortical malformation leads to seizure activity, preventing us from assessing whether the presence of dysmorphic neurons alone without seizure is sufficient to induce hypervascularization.…”

“…We thus used our previously described model of a focal MCD, in which we manipulated mTOR activity in a subset of cortical neural progenitors during corticogenesis. 28,34 In this model, a focal MCD in the somatosensory cortex did not lead to seizures while a focal MCD in the medial prefrontal cortex led to seizures that were visible starting at P21. 28 To generate this model, we introduced a plasmid expressing a constitutively active form of Rheb (Rheb CA , 1.5 μg/μL at E15), 28,34,39,40 the canonical activator of mTOR, into neural progenitors lining the lateral ventricles of the developing cortex through in utero electroporation (IUE) (Figure 2A).…”

“…30,34 Vessels were analyzed in layer 2/3 in Rheb CA -expressing mice and in all layers in transgenic Tsc1 mice using automatic software AngioTool. Only cells within the gray matter of the cortex were quantified.…”

“…For the dendritic analyses, we performed Sholl analysis using Fuji simple neurite tracer as described before. 30,34 Vessels were analyzed in layer 2/3 in Rheb CA -expressing mice and in all layers in transgenic Tsc1 mice using automatic software AngioTool. 35 Analyses were performed in a blinded fashion.…”

Removing 4E-BP Enables Synapses to Refine without Postsynaptic Activity

“…And, interfering with pre- or postsynaptic activity in developing circuits or manipulating postsynaptic calcium influx prevents inputs from refining (Fitzsimonds and Poo, 1998; Balice-Gordon and Lichtman, 1994; Hata and Stryker, 1994; Hashimoto et al, 2011), indicating clearly that the processes that lead to synaptic refinement require activity. Given recent evidence that activity-dependent mechanisms promote cap-dependent mRNA translation in developing dendrites and axons and play essential roles in forming neural circuits (Bramham and Wells, 2007; Lin et al, 2016; Jung et al, 2012; Wang et al, 2010), we asked whether cap-dependent translation has any role in re-organizing neural connections during post-natal development.…”