Phosphorylation Modulates the Subcellular Localization of SOX11

SOX11 is a key Transcription Factor (TF) in the regulation of embryonic and adult neurogenesis, whose mutation has recently been linked to an intellectual disability syndrome in humans. SOX11’s transient activity during neurogenesis is critical to ensure the precise execution of the neurogenic program. Here, we report that SOX11 displays differential subcellular localizations during the course of neurogenesis. Western-Blot analysis of embryonic mouse brain lysates indicated that SOX11 is post-translationally modified by phosphorylation. Using Mass Spectrometry, we found 10 serine residues in the SOX11 protein that are putatively phosphorylated. Systematic analysis of phospho-mutant SOX11 resulted in the identification of the S30 residue, whose phosphorylation promotes nuclear over cytoplasmic localization of SOX11. Collectively, these findings uncover phosphorylation as a novel layer of regulation of the intellectual disability gene Sox11

Phosphorylation of the neurogenic transcription factor SOX11 modulates its subcellular localization and fine-tunes its function during neuronal differentiation

SOX11 gehört zur Familie der SoxC (SRY-verwandten HMG-Box C) Transkriptionsfaktoren und fungiert als Schlüsselregulator der embryonalen und adulten Neurogenese. Sox11 hat nach der Entdeckung, dass Mutationen in Sox11 das Coffin-Siris-Syndrom, eine angeborene Störung, die mit Entwicklungsverzögerung und Mikrozephalie verbunden ist, verursachen, zusätzliche Aufmerksamkeit gefunden. Die Funktion von SOX11 in der Neurogenese reicht vom Überleben der Vorläufer und der Schicksalsbestimmung bis zur dendritischen Morphogenese und Reifung. Die Mechanismen, die SOX11 Expression und Funktion regeln, und damit die SOX11-abhängigen Transkriptionsprogramme, sind jedoch noch nicht verstanden. Um die Mechanismen zu verstehen, durch die das SOX11-Protein reguliert wird, wurde eine Massenspektrometrie durchgeführt, die ergab, dass SOX11 mindestens 10 (phosphorylierte) p-Serinreste aufweist. Um die biologische Rolle der SOX11-Phosphorylierungen aufzuzeigen, wurden phosphomimetische und nicht phosphorylierbare Mutanten erzeugt und funktionell getestet. Diese Studien identifizierten die N-terminalen Serine, die die HMG-Box flankieren, als kritisch für die Funktion von SOX11, indem sie ihre Kernverteilung und ihre Transkriptionsaktivität steuern. Darüber hinaus zeigt diese Studie, dass SOX11 an S133 durch PKA einer Phosphorylierung unterworfen wird, die die dendritische Verfeinerung möglicherweise moduliert, indem möglicherweise die Zielgene differenziell moduliert werden. Insgesamt erweitern diese Daten unser Wissen darüber, wie ein wichtiger Transkriptionsfaktor für die Neurogenese, SOX11, reguliert wird, wodurch das Verständnis der Regulation der SOX11-Aktivität um eine neue Komplexitätsebene erweitert wird. Darüber hinaus war SOX11 in dieser Studie mit einem zentralen neuronalen Entwicklungspfad verbunden, dem cAMP / PKA-Signalweg. Dies ist ein Ansatzpunkt für das Verständnis, wie die SOX11-abhängigen Transkriptionsprogramme von wichtigen neuronalen Entwicklungssignalen reguliert werden.SOX11 belongs to the SoxC (SRY-related HMG-box C) transcription factor family and functions as a key regulator of embryonic and adult neurogenesis. Sox11 has gained attention following the discovery that mutations in Sox11 cause Coffin-Siris Syndrome, a congenital disorder connected with developmental delay and microcephaly. SOX11’s function in neurogenesis ranges from precursor survival and fate commitment to dendritic morphogenesis and maturation. The mechanisms, though, that regulate SOX11 itself and thus the SOX11-dependent transcriptional programs are not yet understood. Aiming to understand the mechanisms by which SOX11 protein is regulated, mass spectrometry was performed and revealed that SOX11 has at least 10 (phosphorylated) p-Serine residues. To unveil the biological role of SOX11’s phosphorylations, phosphomimetic and non-phosphorylatable mutants were generated and functionally tested. These studies identified the N-terminal serines flanking the HMG-box as being critical for the function of SOX11 by controlling its nuclear distribution and its transcriptional activity. Moreover, this study indicates that SOX11 is subjected to phosphorylation on S133 by PKA which may modulate dendritic refinement perhaps by allowing differential modulation of target genes. Overall, these data expand our knowledge of how a key transcription factor for neurogenesis, SOX11, is regulated thereby introducing a new layer of complexity to our understanding of the regulation of SOX11’s activity. Additionally, in this study SOX11 was linked to a central neurodevelopmental pathway, the cAMP/PKA-signaling, providing a starting point into understanding how the SOX11-dependent transcriptional programs are regulated from key neurodevelopmental signals

Analysis of the expression pattern of the schizophrenia-risk and intellectual disability gene TCF4 in the developing and adult brain suggests a role in development and plasticity of cortical and hippocampal neurons

Abstract Background Haploinsufficiency of the class I bHLH transcription factor TCF4 causes Pitt-Hopkins syndrome (PTHS), a severe neurodevelopmental disorder, while common variants in the TCF4 gene have been identified as susceptibility factors for schizophrenia. It remains largely unknown, which brain regions are dependent on TCF4 for their development and function. Methods We systematically analyzed the expression pattern of TCF4 in the developing and adult mouse brain. We used immunofluorescent staining to identify candidate regions whose development and function depend on TCF4. In addition, we determined TCF4 expression in the developing rhesus monkey brain and in the developing and adult human brain through analysis of transcriptomic datasets and compared the expression pattern between species. Finally, we morphometrically and histologically analyzed selected brain structures in Tcf4-haploinsufficient mice and compared our morphometric findings to neuroanatomical findings in PTHS patients. Results TCF4 is broadly expressed in cortical and subcortical structures in the developing and adult mouse brain. The TCF4 expression pattern was highly similar between humans, rhesus monkeys, and mice. Moreover, Tcf4 haploinsufficiency in mice replicated structural brain anomalies observed in PTHS patients. Conclusion Our data suggests that TCF4 is involved in the development and function of multiple brain regions and indicates that its regulation is evolutionary conserved. Moreover, our data validate Tcf4-haploinsufficient mice as a model to study the neurodevelopmental basis of PTHS

Image_2_Phosphorylation Modulates the Subcellular Localization of SOX11.TIF

<p>SOX11 is a key Transcription Factor (TF) in the regulation of embryonic and adult neurogenesis, whose mutation has recently been linked to an intellectual disability syndrome in humans. SOX11’s transient activity during neurogenesis is critical to ensure the precise execution of the neurogenic program. Here, we report that SOX11 displays differential subcellular localizations during the course of neurogenesis. Western-Blot analysis of embryonic mouse brain lysates indicated that SOX11 is post-translationally modified by phosphorylation. Using Mass Spectrometry, we found 10 serine residues in the SOX11 protein that are putatively phosphorylated. Systematic analysis of phospho-mutant SOX11 resulted in the identification of the S30 residue, whose phosphorylation promotes nuclear over cytoplasmic localization of SOX11. Collectively, these findings uncover phosphorylation as a novel layer of regulation of the intellectual disability gene Sox11.</p

Table_1_Phosphorylation Modulates the Subcellular Localization of SOX11.XLSX

<p>SOX11 is a key Transcription Factor (TF) in the regulation of embryonic and adult neurogenesis, whose mutation has recently been linked to an intellectual disability syndrome in humans. SOX11’s transient activity during neurogenesis is critical to ensure the precise execution of the neurogenic program. Here, we report that SOX11 displays differential subcellular localizations during the course of neurogenesis. Western-Blot analysis of embryonic mouse brain lysates indicated that SOX11 is post-translationally modified by phosphorylation. Using Mass Spectrometry, we found 10 serine residues in the SOX11 protein that are putatively phosphorylated. Systematic analysis of phospho-mutant SOX11 resulted in the identification of the S30 residue, whose phosphorylation promotes nuclear over cytoplasmic localization of SOX11. Collectively, these findings uncover phosphorylation as a novel layer of regulation of the intellectual disability gene Sox11.</p

Table_2_Phosphorylation Modulates the Subcellular Localization of SOX11.XLSX

<p>SOX11 is a key Transcription Factor (TF) in the regulation of embryonic and adult neurogenesis, whose mutation has recently been linked to an intellectual disability syndrome in humans. SOX11’s transient activity during neurogenesis is critical to ensure the precise execution of the neurogenic program. Here, we report that SOX11 displays differential subcellular localizations during the course of neurogenesis. Western-Blot analysis of embryonic mouse brain lysates indicated that SOX11 is post-translationally modified by phosphorylation. Using Mass Spectrometry, we found 10 serine residues in the SOX11 protein that are putatively phosphorylated. Systematic analysis of phospho-mutant SOX11 resulted in the identification of the S30 residue, whose phosphorylation promotes nuclear over cytoplasmic localization of SOX11. Collectively, these findings uncover phosphorylation as a novel layer of regulation of the intellectual disability gene Sox11.</p

Image_4_Phosphorylation Modulates the Subcellular Localization of SOX11.TIF

<p>SOX11 is a key Transcription Factor (TF) in the regulation of embryonic and adult neurogenesis, whose mutation has recently been linked to an intellectual disability syndrome in humans. SOX11’s transient activity during neurogenesis is critical to ensure the precise execution of the neurogenic program. Here, we report that SOX11 displays differential subcellular localizations during the course of neurogenesis. Western-Blot analysis of embryonic mouse brain lysates indicated that SOX11 is post-translationally modified by phosphorylation. Using Mass Spectrometry, we found 10 serine residues in the SOX11 protein that are putatively phosphorylated. Systematic analysis of phospho-mutant SOX11 resulted in the identification of the S30 residue, whose phosphorylation promotes nuclear over cytoplasmic localization of SOX11. Collectively, these findings uncover phosphorylation as a novel layer of regulation of the intellectual disability gene Sox11.</p