Adult neural stem cell (aNSC) activity is tuned by external stimuli through the recruitment of transcription factors. This study examines the RE1 silencing transcription factor (REST) in neural stem/progenitor cells isolated from the subventricular zone of adult mouse brain and provides the first extensive characterization of REST‐mediated control of the cellular and molecular properties. This study shows that REST knockdown affects the capacity of progenitor cells to generate neurospheres, reduces cell proliferation, and triggers cell differentiation despite the presence of growth factors. Genome‐ and transcriptome‐wide analyses show that REST binding sites are significantly enriched in genes associated with synaptic transmission and nervous system development and function. Seeking candidate regulators of aNSC function, this study identifies a member of the bone morphogenetic protein (BMP) family, BMP6, the mRNA and protein of which increased after REST knockdown. The results of this study extend previous findings, demonstrating a reciprocal control of REST expression by BMPs. Administration of exogenous BMP6 inhibits aNSC proliferation and induces the expression of the astrocytic marker glial fibrillary acidic protein, highlighting its antimitogenic and prodifferentiative effects. This study suggests that BMP6 produced in a REST‐regulated manner together with other signals can contribute to regulation of NSC maintenance and fate. © 2015 Wiley Periodicals, Inc.
We investigated the level and characteristics of "low Km" 3'-5' cyclic nucleotide phosphodiesterase (PDE) activity in adult and embryo chick spinal cord. The DEAE cellulose chromatography elution profile of Triton X-100 extracts showed a single peak of calmodulin-dependent cAMP/cGMP PDE activity. After two additional purification steps, this activity showed a five-fold activation by calmodulin (Ka = 1.5 nM) for cGMP hydrolysis, and a linear kinetic behaviour with a Km of 1.3 microM. Conversely, the activity showed a biphasic behaviour for cAMP hydrolysis, with Km values of 3.1 and 18.5 microM. The enzyme showed a Stokes radius of 4.5 nm. Western blot analysis of the purified enzyme revealed two immunoreactive bands with molecular mass of 59 and 65 kDa, respectively. Immunohistochemical staining showed motoneuron decoration both on cell soma and fibres. The developmental pattern of Ca2+-calmodulin-dependent PDE expression in spinal cord was also studied; the hydrolytic activity for both substrates has been found to increase constantly from E5 to post-hatching stages, when it appears 5.6-fold higher as compared to the early embryo levels. Furthermore, in cultured spinal cord neurons from E8 embryos, muscle extract has been shown to induce a two-fold increase of Ca2+-calmodulin-dependent cGMP activity. In conclusion, the studies reported here present three relevant findings: (1) the presence in adult and embryo chick spinal cord of PDE activities with characteristics similar to those of the mammalian PDE I enzyme; (2) its localization in the ventral horn motoneurons; (3) its regulated expression during embryogenesis that is possibly related to soluble epigenetic factors produced by the target cells.
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