Meis2 is a Pax6 co-factor in neurogenesis and dopaminergic periglomerular fate specification in the adult olfactory bulb.

Meis homeodomain transcription factors control cell proliferation, cell fate specification and differentiation in development and disease. Previous studies have largely focused on Meis contribution to the development of non-neuronal tissues. By contrast, Meis function in the brain is not well understood. Here, we provide evidence for a dual role of the Meis family protein Meis2 in adult olfactory bulb (OB) neurogenesis. Meis2 is strongly expressed in neuroblasts of the subventricular zone (SVZ) and rostral migratory stream (RMS) and in some of the OB interneurons that are continuously replaced during adult life. Targeted manipulations with retroviral vectors expressing function-blocking forms or with small interfering RNAs demonstrated that Meis activity is cell-autonomously required for the acquisition of a general neuronal fate by SVZ-derived progenitors in vivo and in vitro. Additionally, Meis2 activity in the RMS is important for the generation of dopaminergic periglomerular neurons in the OB. Chromatin immunoprecipitation identified doublecortin and tyrosine hydroxylase as direct Meis targets in newly generated neurons and the OB, respectively. Furthermore, biochemical analyses revealed a previously unrecognized complex of Meis2 with Pax6 and Dlx2, two transcription factors involved in OB neurogenesis. The full pro-neurogenic activity of Pax6 in SVZ derived neural stem and progenitor cells requires the presence of Meis. Collectively, these results show that Meis2 cooperates with Pax6 in generic neurogenesis and dopaminergic fate specification in the adult SVZ-OB system.

Genetic and physical interaction of Meis2, Pax3 and Pax7 during dorsal midbrain development.

BACKGROUND: During early stages of brain development, secreted molecules, components of intracellular signaling pathways and transcriptional regulators act in positive and negative feed-back or feed-forward loops at the mid-hindbrain boundary. These genetic interactions are of central importance for the specification and subsequent development of the adjacent mid- and hindbrain. Much less, however, is known about the regulatory relationship and functional interaction of molecules that are expressed in the tectal anlage after tectal fate specification has taken place and tectal development has commenced. RESULTS: Here, we provide experimental evidence for reciprocal regulation and subsequent cooperation of the paired-type transcription factors Pax3, Pax7 and the TALE-homeodomain protein Meis2 in the tectal anlage. Using in ovo electroporation of the mesencephalic vesicle of chick embryos we show that (i) Pax3 and Pax7 mutually regulate each other's expression in the mesencephalic vesicle, (ii) Meis2 acts downstream of Pax3/7 and requires balanced expression levels of both proteins, and (iii) Meis2 physically interacts with Pax3 and Pax7. These results extend our previous observation that Meis2 cooperates with Otx2 in tectal development to include Pax3 and Pax7 as Meis2 interacting proteins in the tectal anlage. CONCLUSION: The results described here suggest a model in which interdependent regulatory loops involving Pax3 and Pax7 in the dorsal mesencephalic vesicle modulate Meis2 expression. Physical interaction with Meis2 may then confer tectal specificity to a wide range of otherwise broadly expressed transcriptional regulators, including Otx2, Pax3 and Pax7.

Meis2 competes with the Groucho co-repressor Tle4 for binding to Otx2 and specifies tectal fate without induction of a secondary midbrain-hindbrain boundary organizer.

The transcription factor Otx2 is expressed throughout the anterior neuroectoderm and is required for the formation of all forebrain- and midbrain-derived structures. The molecular determinants that cooperate with Otx2 to subdivide its expression domain into distinct functional units are, however, poorly understood at present. We show here that the TALE-homeodomain protein Meis2 is expressed in the chick tectal anlage and is both necessary and sufficient for tectal development. Unlike known tectum-inducing genes, the ability of Meis2 to initiate tectal development does not involve the formation of a secondary midbrain-hindbrain boundary organizer, but instead requires direct interaction with Otx2. Using an Otx2-dependent reporter assay we demonstrate that Meis2 competes with the Groucho co-repressor Tle4 (Grg4) for binding to Otx2 and thereby restores Otx2 transcriptional activator function. Together, our data suggest a model in which the balance between a co-repressor and a co-activator, which compete for binding to Otx2 in the mesencephalic vesicle, provides spatial and temporal control over tectal development. Controlled formation of Meis2-containing higher order protein complexes might thus serve as a general mechanism to achieve subdivision of the anterior neuroectoderm into distinct functional units during embryogenesis.

Differential and dose-dependent regulation of gene expression at the mid-hindbrain boundary by Ras-MAP kinase signaling.

Recent experiments suggest that activation of the Ras-MAP kinase pathway at the mid-hindbrain boundary (MHB) induces cerebellar development, whereas tectal development occurs in the absence of Ras-MAP kinase activity. To test this model we have stimulated or inhibited Ras-MAP kinase signaling in chick embryos through targeted misexpression of a constitutive active (Ras(V12)) or dominant negative (Ras(N17)) form of Ras. The consequence of these manipulations on the expression of several genes that are expressed in distinct patterns at or around the MHB organizer, including En1, Pax2, Pax3, Pax5, Wnt1, Meis2, and ephrin-A2, -A5, and -B1, was assessed. Extending previous findings we show that inhibition of Ras-MAP kinase signaling differently affects Pax3 expression in different regions of the mid-hindbrain territory, inhibiting its expression in the midbrain but inducing it in the MHB region. Expression of the midbrain specific marker gene Meis2 was not affected by Ras(N17) at first but later upregulated concomitantly with the morphological transformation of hindbrain to midbrain. In addition, we show that different dosages of Ras-MAP kinase activity are required for transcriptional activation of Wnt1 and En1 at the MHB. Collectively, these results validate and extend previous findings on the molecular changes associated with Fgf8 loss-of-function or gain-of-function phenotypes at the MHB, demonstrate that gene expression at the MHB is regulated by Ras-MAP kinase signaling in a spatially and temporally distinct manner and provide evidence for a dosage dependent function of Fgf8 signaling at the MHB.

MEIS homeodomain proteins facilitate PARP1/ARTD1-mediated eviction of histone H1.

Pre-B-cell leukemia homeobox (PBX) and myeloid ecotropic viral integration site (MEIS) proteins control cell fate decisions in many physiological and pathophysiological contexts, but how these proteins function mechanistically remains poorly defined. Focusing on the first hours of neuronal differentiation of adult subventricular zone-derived stem/progenitor cells, we describe a sequence of events by which PBX-MEIS facilitates chromatin accessibility of transcriptionally inactive genes: In undifferentiated cells, PBX1 is bound to the H1-compacted promoter/proximal enhancer of the neuron-specific gene doublecortin (Dcx) Once differentiation is induced, MEIS associates with chromatin-bound PBX1, recruits PARP1/ARTD1, and initiates PARP1-mediated eviction of H1 from the chromatin fiber. These results for the first time link MEIS proteins to PARP-regulated chromatin dynamics and provide a mechanistic basis to explain the profound cellular changes elicited by these proteins.

Simvastatin promotes adult hippocampal neurogenesis by enhancing Wnt/ß-catenin signaling.

Statins improve recovery from traumatic brain injury and show promise in preventing Alzheimer disease. However, the mechanisms by which statins may be therapeutic for neurological conditions are not fully understood. In this study, we present the initial evidence that oral administration of simvastatin in mice enhances Wnt signaling in vivo. Concomitantly, simvastatin enhances neurogenesis in cultured adult neural progenitor cells as well as in the dentate gyrus of adult mice. Finally, we find that statins enhance Wnt signaling through regulation of isoprenoid synthesis and not through cholesterol. These findings provide direct evidence that Wnt signaling is enhanced in vivo by simvastatin and that this elevation of Wnt signaling is required for the neurogenic effects of simvastatin. Collectively, these data add to the growing body of evidence that statins may have therapeutic value for treating certain neurological disorders.

Hypoxia-inducible factors have distinct and stage-specific roles during reprogramming of human cells to pluripotency.

Pluripotent stem cells have distinct metabolic requirements, and reprogramming cells to pluripotency requires a shift from oxidative to glycolytic metabolism. Here, we show that this shift occurs early during reprogramming of human cells and requires hypoxia-inducible factors (HIFs) in a stage-specific manner. HIF1α and HIF2α are both necessary to initiate this metabolic switch and for the acquisition of pluripotency, and the stabilization of either protein during early phases of reprogramming is sufficient to induce the switch to glycolytic metabolism. In contrast, stabilization of HIF2α during later stages represses reprogramming, partly because of the upregulation of TNF-related apoptosis-inducing ligand (TRAIL). TRAIL inhibits induced pluripotent stem cell (iPSC) generation by repressing apoptotic caspase 3 activity specifically in cells undergoing reprogramming but not human embryonic stem cells (hESCs), and inhibiting TRAIL activity enhances human iPSC generation. These results shed light on the mechanisms underlying the metabolic shifts associated with the acquisition of a pluripotent identity during reprogramming.

Evidence against involvement of Bmp receptor 1b signaling in fate specification of the chick mesencephalic alar plate at HH16.

Studies in the developing spinal cord have established that morphogenes secreted from the roof- and floor plate influence pattern formation along the dorsal-ventral axis of the neural tube. Bone morphogenetic proteins (Bmps), secreted from the roof plate, act on the more laterally located alar plates to induce position dependent gene expression and cell fate changes. The dorsalizing activity of Bmps is counteracted by Sonic hedgehog (Shh), which is secreted from the floor plate and underlying notochord. Bmps are also expressed in the roof plate of the mesencephalic vesicle, yet it is unclear at present if they also provide patterning information to the mesencephalic alar plates. We have experimentally tested the hypothesis that Bmp signaling is required for fate specification of the mesencephalic alar plate by manipulating Bmp receptor signaling in the early chick embryo through ectopic expression of mutated forms of Bmp receptor 1b (BmpR1b), which render the receptor constitutively active or dominant negative, respectively. In contrast to published data on the embryonic spinal cord, neither activation nor blockage of BmpR1b signaling in stage 16 embryos altered expression of markers of the mesencephalic alar plates including Pax3, Pax7, Meis2 and efnb1. Moreover, simultaneous activation of BmpR1b signaling and blockage of Shh signaling was not sufficient to induce Meis2 expression in the ventral mesencephalon. Therefore, whereas the importance of Bmp signaling for dorsal differentiation in the spinal cord is well established, it appears to play a less prominent role in the dorsal specification of the developing mesencephalon during the same developmental stages.

Retroviral misexpression of cVax disturbs retinal ganglion cell axon fasciculation and intraretinal pathfinding in vivo and guidance of nasal ganglion cell axons in vivo.

The transcription factor cVax (Vax2) is expressed in the ventral neural retina and restricted expression is a prerequisite for at least three prominent aspects of retinal dorsal-ventral patterning: polarized expression of EphB/B-ephrin molecules, the retinotectal projection and the distribution of rod photoreceptors across the retina. In the chick retina, the fasciculation pattern of ganglion cell axons also differs between the dorsal and ventral eye. To investigate the molecular mechanisms involved, the nerve fiber layer was analyzed after retroviral misexpression of several factors known to regulate the positional specification of retinal ganglion cells. Forced cVax expression ventralized the fasciculation pattern and caused axon pathfinding errors near the optic disc. Ectopic expression of different ephrin molecules indicated that axon fasciculation is, at least in part, mediated by the EphB system. Finally, we report that retroviral misexpression of cVax increased the pool of EphA4 receptors phosphorylated on tyrosine residues and altered the guidance preference of nasal axons in vitro. These results identify novel functions for cVax in intraretinal axon fasciculation and pathfinding as well as suggest a mechanism to explain how restricted cVax expression may influence map formation along the dorso-ventral and antero-posterior axes of the optic tectum.