Multifaceted actions of Zeb2 in postnatal neurogenesis from the ventricular-subventricular zone to the olfactory bulb.

The transcription factor Zeb2 controls fate specification and subsequent differentiation and maturation of multiple cell types in various embryonic tissues. It binds many protein partners, including activated Smad proteins and the NuRD co-repressor complex. How Zeb2 subdomains support cell differentiation in various contexts has remained elusive. Here, we studied the role of Zeb2 and its domains in neurogenesis and neural differentiation in the young postnatal ventricular-subventricular zone (V-SVZ), in which neural stem cells generate olfactory bulb-destined interneurons. Conditional Zeb2 knockouts and separate acute loss- and gain-of-function approaches indicated that Zeb2 is essential for controlling apoptosis and neuronal differentiation of V-SVZ progenitors before and after birth, and we identified Sox6 as a potential downstream target gene of Zeb2. Zeb2 genetic inactivation impaired the differentiation potential of the V-SVZ niche in a cell-autonomous fashion. We also provide evidence that its normal function in the V-SVZ also involves non-autonomous mechanisms. Additionally, we demonstrate distinct roles for Zeb2 protein-binding domains, suggesting that Zeb2 partners co-determine neuronal output from the mouse V-SVZ in both quantitative and qualitative ways in early postnatal life.

Aptamers and their potential to selectively target aspects of EGF, Wnt/ß-catenin and TGFß-smad family signaling.

The smooth identification and low-cost production of highly specific agents that interfere with signaling cascades by targeting an active domain in surface receptors, cytoplasmic and nuclear effector proteins, remain important challenges in biomedical research. We propose that peptide aptamers can provide a very useful and new alternative for interfering with protein-protein interactions in intracellular signal transduction cascades, including those emanating from activated receptors for growth factors. By their targeting of short, linear motif type of interactions, peptide aptamers have joined nucleic acid aptamers for use in signaling studies because of their ease of production, their stability, their high specificity and affinity for individual target proteins, and their use in high-throughput screening protocols. Furthermore, they are entering clinical trials for treatment of several complex, pathological conditions. Here, we present a brief survey of the use of aptamers in signaling pathways, in particular of polypeptide growth factors, starting with the published as well as potential applications of aptamers targeting Epidermal Growth Factor Receptor signaling. We then discuss the opportunities for using aptamers in other complex pathways, including Wnt/ß-catenin, and focus on Transforming Growth Factor-ß/Smad family signaling.

How cell-autonomous is neuronal migration in the forebrain? Molecular cross-talk at the cell membrane.

In the adult brain, different cell types communicate with each other through cell-cell contacts and brain activity is regulated at the cell membrane. But long before the brain is fully functional, different excitatory and inhibitory cell types generated at distinct places migrate through the developing brain to their final position. The elements guiding these migrating neurons, either structural axonal scaffolds or chemical guidance factors, are relatively well described. However, the molecules involved in the individual short-timed membrane contacts migrating cells make with other cells during their migration process are less well understood. This update focuses on recent novel insights into the molecular nature of these cell-cell contacts and the cross-talk taking place at the cell membrane.

Four amino acids within a tandem QxVx repeat in a predicted extended α-helix of the Smad-binding domain of Sip1 are necessary for binding to activated Smad proteins.

The zinc finger transcription factor Smad-interacting protein-1 (Sip1; Zeb2, Zfhx1b) plays an important role during vertebrate embryogenesis in various tissues and differentiating cell types, and during tumorigenesis. Previous biochemical analysis suggests that interactions with several partner proteins, including TGFß family receptor-activated Smads, regulate the activities of Sip1 in the nucleus both as a DNA-binding transcriptional repressor and activator. Using a peptide aptamer approach we mapped in Sip1 its Smad-binding domain (SBD), initially defined as a segment of 51 amino acids, to a shorter stretch of 14 amino acids within this SBD. Modelling suggests that this short SBD stretch is part of an extended α-helix that may fit the binding to a hydrophobic corridor within the MH2 domain of activated Smads. Four amino acids (two polar Q residues and two non-polar V residues) that form the tandem repeat (QxVx)2 in this 14-residue stretch were found to be crucial for binding to both TGFß/Nodal/Activin-Smads and BMP-Smads. A full-length Sip1 with collective mutation of these Q and V residues (to A) no longer binds to Smads, while it retains its binding activity to its cognate bipartite target DNA sequence. This missense mutant Sip1(AxAx)2 provides a new molecular tool to identify SBD (in)dependent target genes in Sip1-controlled TGFß and/or BMP (de)regulated cellular, developmental and pathological processes.

Directed migration of cortical interneurons depends on the cell-autonomous action of Sip1.

GABAergic interneurons mainly originate in the medial ganglionic eminence (MGE) of the embryonic ventral telencephalon (VT) and migrate tangentially to the cortex, guided by membrane-bound and secreted factors. We found that Sip1 (Zfhx1b, Zeb2), a transcription factor enriched in migrating cortical interneurons, is required for their proper differentiation and correct guidance. The majority of Sip1 knockout interneurons fail to migrate to the neocortex and stall in the VT. RNA sequencing reveals that Sip1 knockout interneurons do not acquire a fully mature cortical interneuron identity and contain increased levels of the repulsive receptor Unc5b. Focal electroporation of Unc5b-encoding vectors in the MGE of wild-type brain slices disturbs migration to the neocortex, whereas reducing Unc5b levels in Sip1 knockout slices and brains rescues the migration defect. Our results reveal that Sip1, through tuning of Unc5b levels, is essential for cortical interneuron guidance.

Dual-mode modulation of Smad signaling by Smad-interacting protein Sip1 is required for myelination in the central nervous system.

Myelination by oligodendrocytes in the central nervous system (CNS) is essential for proper brain function, yet the molecular determinants that control this process remain poorly understood. The basic helix-loop-helix transcription factors Olig1 and Olig2 promote myelination, whereas bone morphogenetic protein (BMP) and Wnt/ß-catenin signaling inhibit myelination. Here we show that these opposing regulators of myelination are functionally linked by the Olig1/2 common target Smad-interacting protein-1 (Sip1). We demonstrate that Sip1 is an essential modulator of CNS myelination. Sip1 represses differentiation inhibitory signals by antagonizing BMP receptor-activated Smad activity while activating crucial oligodendrocyte-promoting factors. Importantly, a key Sip1-activated target, Smad7, is required for oligodendrocyte differentiation and partially rescues differentiation defects caused by Sip1 loss. Smad7 promotes myelination by blocking the BMP- and ß-catenin-negative regulatory pathways. Thus, our findings reveal that Sip1-mediated antagonism of inhibitory signaling is critical for promoting CNS myelination and point to new mediators for myelin repair.

Few Smad proteins and many Smad-interacting proteins yield multiple functions and action modes in TGFß/BMP signaling in vivo.

Signaling by the many ligands of the TGFß family strongly converges towards only five receptor-activated, intracellular Smad proteins, which fall into two classes i.e. Smad2/3 and Smad1/5/8, respectively. These Smads bind to a surprisingly high number of Smad-interacting proteins (SIPs), many of which are transcription factors (TFs) that co-operate in Smad-controlled target gene transcription in a cell type and context specific manner. A combination of functional analyses in vivo as well as in cell cultures and biochemical studies has revealed the enormous versatility of the Smad proteins. Smads and their SIPs regulate diverse molecular and cellular processes and are also directly relevant to development and disease. In this survey, we selected appropriate examples on the BMP-Smads, with emphasis on Smad1 and Smad5, and on a number of SIPs, i.e. the CPSF subunit Smicl, Ttrap (Tdp2) and Sip1 (Zeb2, Zfhx1b) from our own research carried out in three different vertebrate models.