Genoarchitecture of the Early Postmitotic Pretectum and the Role of Wnt Signaling in Shaping Pretectal Neurochemical Anatomy in Zebrafish.

The pretectum has a distinct nuclear arrangement and complex neurochemical anatomy. While previous genoarchitectural studies have described rostrocaudal and dorsoventral progenitor domains and subdomains in different species, the relationship between these early partitions and its later derivatives in the mature anatomy is less understood. The signals and transcription factors that control the establishment of pretectal anatomy are practically unknown. We investigated the possibility that some aspects of the development of pretectal divisions are controlled by Wnt signaling, focusing on the transitional stage between neurogenesis and histogenesis in zebrafish. Using several molecular markers and following the prosomeric model, we identified derivatives from each rostrocaudal pretectal progenitor domain and described the localization of gad1b-positive GABAergic and vglut2.2-positive glutamatergic cell clusters. We also attempted to relate these clusters to pretectal nuclei in the mature brain. Then, we examined the influence of Wnt signaling on the size of neurochemically distinctive pretectal areas, using a chemical inhibitor of the Wnt pathway and the CRISPR/Cas9 approach to knock out genes that encode the Wnt pathway mediators, Lef1 and Tcf7l2. The downregulation of the Wnt pathway led to a decrease in two GABAergic clusters and an expansion of a glutamatergic subregion in the maturing pretectum. This revealed an instructive role of the Wnt signal in the development of the pretectum during neurogenesis. The molecular anatomy presented here improves our understanding of pretectal development during early postmitotic stages and support the hypothesis that Wnt signaling is involved in shaping the neurochemical organization of the pretectum.

MLK4 regulates DNA damage response and promotes triple-negative breast cancer chemoresistance.

Chemoresistance constitutes a major challenge in the treatment of triple-negative breast cancer (TNBC). Mixed-Lineage Kinase 4 (MLK4) is frequently amplified or overexpressed in TNBC where it facilitates the aggressive growth and migratory potential of breast cancer cells. However, the functional role of MLK4 in resistance to chemotherapy has not been investigated so far. Here, we demonstrate that MLK4 promotes TNBC chemoresistance by regulating the pro-survival response to DNA-damaging therapies. We observed that MLK4 knock-down or inhibition sensitized TNBC cell lines to chemotherapeutic agents in vitro. Similarly, MLK4-deficient cells displayed enhanced sensitivity towards doxorubicin treatment in vivo. MLK4 silencing induced persistent DNA damage accumulation and apoptosis in TNBC cells upon treatment with chemotherapeutics. Using phosphoproteomic profiling and reporter assays, we demonstrated that loss of MLK4 reduced phosphorylation of key DNA damage response factors, including ATM and CHK2, and compromised DNA repair via non-homologous end-joining pathway. Moreover, our mRNA-seq analysis revealed that MLK4 is required for DNA damage-induced expression of several NF-кB-associated cytokines, which facilitate TNBC cells survival. Lastly, we found that high MLK4 expression is associated with worse overall survival of TNBC patients receiving anthracycline-based neoadjuvant chemotherapy. Collectively, these results identify a novel function of MLK4 in the regulation of DNA damage response signaling and indicate that inhibition of this kinase could be an effective strategy to overcome TNBC chemoresistance.

Spreading Depressions and Periinfarct Spreading Depolarizations in the Context of Cortical Plasticity.

Studies of cortical function-recovery require a comparison between normal and post-stroke conditions that lead to changes in cortical metaplasticity. Focal cortical stroke impairs experience-dependent plasticity in the neighboring somatosensory cortex and usually evokes periinfarct depolarizations (PiDs) - spreading depression-like waves. Experimentally induced spreading depressions (SDs) affect gene expression and some of these changes persist for at least 30 days. In this study we compare the effects of non-stroke depolarizations that impair cortical experience-dependent plasticity to the effects of stroke, by inducing experience-dependent plasticity in rats with SDs or PiDs by a month of contralateral partial whiskers deprivation. We found that whiskers' deprivation after SDs resulted in normal cortical representation enlargement suggesting that SDs and PiDs depolarization have no influence on experience-dependent plasticity cortical map reorganization. PiDs and the MMP-9, -3, -2 or COX-2 proteins, which are assumed to influence metaplasticity in rats after stroke were compared between SDs induced by high osmolarity KCl solution and the PiDs that followed cortical photothrombotic stroke (PtS). We found that none of these factors directly caused cortical post-stroke metaplasticity changes. The only significant difference between stoke and induced SD was a greater imbalance in interhemispheric activity equilibrium after stroke. The interhemispheric interactions that were modified by stroke may therefore be promising targets for future studies of post-stroke experience-dependent plasticity and of recuperation studies.

Wnt/ß-catenin signaling in brain development and mental disorders: keeping TCF7L2 in mind.

Canonical Wnt signaling, which is transduced by ß-catenin and lymphoid enhancer factor 1/T cell-specific transcription factors (LEF1/TCFs), regulates many aspects of metazoan development and tissue renewal. Although much evidence has associated canonical Wnt/ß-catenin signaling with mood disorders, the mechanistic links are still unknown. Many components of the canonical Wnt pathway are involved in cellular processes that are unrelated to classical canonical Wnt signaling, thus further blurring the picture. The present review critically evaluates the involvement of classical Wnt/ß-catenin signaling in developmental processes that putatively underlie the pathology of mental illnesses. Particular attention is given to the roles of LEF1/TCFs, which have been discussed surprisingly rarely in this context. Highlighting recent discoveries, we propose that alterations in the activity of LEF1/TCFs, and particularly of transcription factor 7-like 2 (TCF7L2), result in defects previously associated with neuropsychiatric disorders, including imbalances in neurogenesis and oligodendrogenesis, the functional disruption of thalamocortical circuitry and dysfunction of the habenula.

TCF7L2 mediates the cellular and behavioral response to chronic lithium treatment in animal models.

The mechanism of lithium's therapeutic action remains obscure, hindering the discovery of safer treatments for bipolar disorder. Lithium can act as an inhibitor of the kinase GSK3α/ß, which in turn negatively regulates ß-catenin, a co-activator of LEF1/TCF transcription factors. However, unclear is whether therapeutic levels of lithium activate ß-catenin in the brain, and whether this activation could have a therapeutic significance. To address this issue we chronically treated mice with lithium. Although the level of non-phospho-ß-catenin increased in all of the brain areas examined, ß-catenin translocated into cellular nuclei only in the thalamus. Similar results were obtained when thalamic and cortical neurons were treated with a therapeutically relevant concentration of lithium in vitro. We tested if TCF7L2, a member of LEF1/TCF family that is highly expressed in the thalamus, facilitated the activation of ß-catenin. Silencing of Tcf7l2 in thalamic neurons prevented ß-catenin from entering the nucleus, even when the cells were treated with lithium. Conversely, when Tcf7l2 was ectopically expressed in cortical neurons, ß-catenin shifted to the nucleus, and lithium augmented this process. Lastly, we silenced tcf7l2 in zebrafish and exposed them to lithium for 3 days, to evaluate whether TCF7L2 is involved in the behavioral response. Lithium decreased the dark-induced activity of control zebrafish, whereas the activity of zebrafish with tcf7l2 knockdown was unaltered. We conclude that therapeutic levels of lithium activate ß-catenin selectively in thalamic neurons. This effect is determined by the presence of TCF7L2, and potentially contributes to the therapeutic response.