Satb2 Cre/+ mouse as a tool to investigate cell fate determination in the developing neocortex

Ambrozkiewicz, Mateusz C.; Bessa, Paraskevi; Salazar-Lázaro, Andrea; Salina, V. A.; Tarabykin, Victor

doi:10.1016/j.jneumeth.2017.07.023

Cited by 6 publications

(17 citation statements)

References 25 publications

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“…Primary antibodies used for immunocytochemistry were used at dilutions indicated: anti-Satb2 (1:500, rabbit, home-made; (Ambrozkiewicz et al, 2017)), anti-Bcl11b (1:500, rat, Abcam, 25B6, anti-Ctip2), anti-GFP…”

Section: Antibodies For Immunohistochemistrymentioning

confidence: 99%

A critical period of translational control during brain development at codon resolution

Harnett

Ambrozkiewicz

Zinnall

et al. 2021

Preprint

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Translation modulates the timing and amplification of gene expression after transcription. Development of the brain's neocortex requires precisely timed and spatially targeted gene expression, but the relationship between mRNA vs. protein synthesis throughout the genome is unknown. We perform a comprehensive analysis of the reactants, synthesis, and products of mRNA translation spanning mouse neocortex neurogenesis. Ribosome number in the cortical plate decreases sharply at mid-neurogenesis during a transition in neuronal subtype specification, shifting the fundamental kinetics of protein synthesis, with mRNA and protein levels frequently divergent. Satb2, which drives an essential neuronal subtype-specific program, is a highly dynamically translated mRNA with surprisingly broad transcription across diverse neuronal lineages. Satb2 protein achieves its neuronal subtype expression through timed regulation by the RNA-binding protein Pumilio2. Thus, the refinement of transcriptional programs by protein synthesis is a widespread feature of neuronal specification. Developmental neocortex translatome data are provided in an open-source resource: https://shiny.mdc-berlin.de/cortexomics/.

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Section: Antibodies For Immunohistochemistrymentioning

confidence: 99%

A critical period of translational control during brain development at codon resolution

Harnett

Ambrozkiewicz

Zinnall

et al. 2021

Preprint

Self Cite

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“…In order to identify signaling pathways orchestrating neuronal subtype specification, we have previously developed a method for screening small molecule inhibitors for their role in the control of cell fate acquisition (Ambrozkiewicz et al, 2017), which utilizes the Satb2 Cre/+ mouse line (Britanova et al, 2006). By expressing a Cre-inducible fluorescent reporter loxP-Stop-loxP-tdTomato in Satb2 Cre/+ primary cortical cells (Fig.…”

Section: Ire1a Is a Positive Regulator Of Satb2 And Axon Specification In Developing Cortical Neuronsmentioning

confidence: 99%

“…For our screening, we used beta-actin driven expression constructs pCAG-EGFP and pCAGflox-stop-flox-tdTomato, which are described in our previous work in detail (Ambrozkiewicz et al, 2017) Cloning strategies for constructs generated in this study Cloning of all expression vectors in this study was performed using the NEBuilder system according to manufacturer's protocol (New England BioLabs). DNA fragments were amplified using GXL Prime Star DNA polymerase (Takara) using cDNA libraries as templates.…”

Section: Expression Vectorsmentioning

confidence: 99%

“…Transfection of primary cortical cells was performed using nucleofection (Lonza Bioscience) according to our previously published protocol (Ambrozkiewicz et al, 2017).…”

Section: Nucleofection Of Primary Neuronsmentioning

confidence: 99%

“…Two hours post-plating, cultures were treated with compounds at two concentrations, in technical duplicates. Cells were then cultivated until DIV2, when the proportion of Satb2 tdTom neurons normalized to EGFP positive cells was determined using FACS as described (Ambrozkiewicz et al, 2017).…”

Section: Small Molecule Inhibitor Screeningmentioning

confidence: 99%

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Ire1α-Regulated mRNA Translation Rate Controls the Identity and Polarity of Upper Layer Cortical Neurons

Ambrozkiewicz

Борисова

Newman

et al. 2021

Preprint

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Establishment of cortical layers and axon-dendrite polarity in neurons is fundamental for brain connectivity. Here, we present that timed mRNA translation control by Inositol-Requiring Enzyme 1α, Ire1α, is necessary for acquisition of upper layer neuronal identity and a single axon. We demonstrate that Ire1α acts as a canonical regulator of global protein synthesis in developing cortical neurons, controlling the level of actively translating ribosomes, expression of translation factors, and ribosomal proteins. Translation rates distinguish early and late neuronal progenitors and early- and late-born postmitotic neurons, indicative of developmental stage- and differentiation-specific requirements for protein synthesis rates in the formation of upper and deeper cortical layers. We demonstrate that specification and polarization of upper layer neurons is uniquely sensitive to translation rate, in contrast to deep layer neurons. Our data shed light onto the post-transcriptional source of cellular diversity in the developing cortex and unveils stress-independent homeostatic functions of Ire1α.

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Protein translation rate determines neocortical neuron fate

Borisova,

Newman,

Couce Iglesias

et al. 2024

Nat Commun

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The mammalian neocortex comprises an enormous diversity regarding cell types, morphology, and connectivity. In this work, we discover a post-transcriptional mechanism of gene expression regulation, protein translation, as a determinant of cortical neuron identity. We find specific upregulation of protein synthesis in the progenitors of later-born neurons and show that translation rates and concomitantly protein half-lives are inherent features of cortical neuron subtypes. In a small molecule screening, we identify Ire1α as a regulator of Satb2 expression and neuronal polarity. In the developing brain, Ire1α regulates global translation rates, coordinates ribosome traffic, and the expression of eIF4A1. Furthermore, we demonstrate that the Satb2 mRNA translation requires eIF4A1 helicase activity towards its 5’-untranslated region. Altogether, we show that cortical neuron diversity is generated by mechanisms operating beyond gene transcription, with Ire1α-safeguarded proteostasis serving as an essential regulator of brain development.

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Satb2 Cre/+ mouse as a tool to investigate cell fate determination in the developing neocortex

Cited by 6 publications

References 25 publications

A critical period of translational control during brain development at codon resolution

A critical period of translational control during brain development at codon resolution

Ire1α-Regulated mRNA Translation Rate Controls the Identity and Polarity of Upper Layer Cortical Neurons

Protein translation rate determines neocortical neuron fate

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