Despite vast scientific interest, molecular mechanisms underlying neural stem cell self-renewal and neurogenesis remain elusive. Recently, microRNAs emerged as central players in neural stem cell self-renewal and differentiation. . This Research Topic will focus on how microRNAs, through their interactions with other components of the regulatory network, control the self-renewal and differentiation of neural stem cells.  Deciphering such a mechanism will not only provide insights into basic neurosciences but also facilitate clinical applications of stem cell-based therapies for neurodegenerative diseases.

Department of Stem Cell Biology and Regenerative Medicine, City of Hope Medical Center

University of New Mexico

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Frontiers in Neuroscience

Neurogenesis

microRNAs in neural stem cells and neurogenesis

There are many biological processes occurring in nature with dynamic complexity: neurogenesis is one such process. The special topic of &#x201c;microRNAs in Neural Stem Cells (NSCs) and Neurogenesis&#x201d; in Frontiers in Neuroscience discussed diverse roles microRNAs (miRNA) play in neurogenesis through a series of review articles and original research articles. This editorial serves to highlight these appealing articles, which build a comprehensive story about the many different roles miRNA molecules play in NSCs and neurogenesis.Neurogenesis involves the generation of newborn neuronal cells from NSCs and progenitor cells in the mammalian brain. The main steps of neurogenesis include NSC self-renewal, neural progenitor cell proliferation, neuronal commitment, migration, maturation, and integration (<a href="#B14">Shi et al., 2010</a>). Neurogenesis is highly active during development when new neurons are formed and integrated into the growing brain. Neurogenesis continues to occur in the brain throughout adulthood within two discrete adult neurogenic niches, the hippocampal dentate gyrus and the subventricular zone (<a href="#B10">Lois and Alvarez-Buylla, 1994</a>; <a href="#B5">Eriksson et al., 1998</a>).In recent years, much attention has been focused on the impact miRNAs have on the process of neurogenesis. miRNAs are single-stranded, non-coding molecules that typically range between 21 and 24 nucleotides in length. They are post-transcriptional regulators that are endogenously expressed and bind to complementary sequences of messenger RNA targets. miRNAs regulate multiple processes including: development, cell proliferation and differentiation, growth and neurogenesis (<a href="#B2">Ambros, 2004</a>; <a href="#B3">Bartel, 2004</a>). Though small in length, miRNAs control gene expression through targeting many downstream targets. In a recent review, miRNAs are introduced as an important player in the regulation of embryonic stem cells and neurogenesis (<a href="#B8">Kawahara et al., 2012</a>). Emphasis is placed on components of miRNA biogenesis, such as Dicer, Drosha, DGCR8, Lin-28, and other related proteins involved in neurogenesis; some of which have been linked to cancer, fragile × syndrome, and nervous system disorders (<a href="#B8">Kawahara et al., 2012</a>). The review written by Lang and Shi highlighted the dynamic roles that miRNAs play in multiple steps of neurogenesis including: NSC proliferation, self-renewal, neuronal differentiation, maturation, and dendritic spine morphogenesis (<a href="#B9">Lang and Shi, 2012</a>). This review covered key miRNA regulators in neural development and adult neurogenesis. It summarized the role of miR-9, miR-124, miR-137, miR-184, and let-7 in NSC proliferation and differentiation, miR-125b and miR-128 in neuronal differentiation and maturation, and miR-132, miR-134, miR-138 in dendritic spine morphogenesis (<a href="#B9">Lang and Shi, 2012</a>).Switching gears from neuronal differentiation, Barca-Mayo and Lu discussed miRNAs important for glial-lineage fate specification with an emphasis on the fine-tuning of oligodendrocyte development (<a href="#B11">Lu and Barca, 2012</a>). The authors elaborated on the roles of miRNAs, particularly miR-219, miR-338, and miR-138, in regulating oligodendrocyte differentiation and maturation. Also highlighted in this article is a discussion concerning the potential use of miRNAs as disease biomarkers for nervous system diseases, such as multiple sclerosis, owing to a greater stability of miRNAs in comparison to messenger RNAs (<a href="#B11">Lu and Barca, 2012</a>).As discussed by Schouten et al., miRNAs have been implicated in aging-associated cognitive decline, synapse formation, and the effects of circulating levels of steroid hormones (<a href="#B13">Schouten et al., 2012</a>). Within this framework, the ability of newborn neurons to functionally integrate into hippocampal circuits could be partly attributed to the activity of miR-132. Furthermore, the authors gave a compelling account of miRNAs in adult hippocampal neurogenesis, along with their effects on target genes and the potential influence these interactions have on neural development disorders, such as Rett syndrome and autism.A discussion concerning neurogenesis would not be complete without taking a look at the epigenetic mechanisms regulating gene expression important for NSC maintenance and fate specification. In their review, Jobe et al. explored the ability of non-coding RNAs to “crosstalk” with other epigenetic mechanisms—namely DNA methylation and histone modification. The authors paid particular attention to the contributions of epigenetic mechanisms to NSC regulatory networks in adult neurogenesis (<a href="#B7">Jobe et al., 2012</a>). Furthermore, this review raises a discussion concerning how neuronal activities, inflammation, stress, and diseases lead to changes in epigenetic states.While there are many tools offering a “straight-forward” approach to miRNA transcriptional profiling, such as miRNA arrays, PCR, and Northern blotting, functional studies on miRNA remain technically challenging (<a href="#B1">Akerblom et al., 2012</a>). For instance, Dicer knockout studies suggest critical roles for miRNA in neurogenesis, but the results are difficult to interpret because knockout of Dicer affects all miRNAs, not just the miRNAs of interest. In addition, the stability of mature miRNAs makes conditional Dicer knockout studies difficult to control temporally. This being said, in their review, Akerblom et al. discussed examples taken from studies done on critical regulatory miRNAs in neurogenesis, including miR-124, miR-9, and Let-7 family members (<a href="#B1">Akerblom et al., 2012</a>).In an interesting original research article, Chen and Wichterle performed conditional knockout of Dicer in motor neuron progenitors in order to elucidate a role for miRNAs in the regulation of post-mitotic neurons and fate specification of different motor neuron subtypes in each segment of the developing spinal cord (<a href="#B4">Chen and Wichterle, 2012</a>). Their studies revealed that the requirement for Dicer function in motor neurons is subtype-specific and that miRNAs are important for proper motor pool specification and the maintenance of motor neurons (<a href="#B4">Chen and Wichterle, 2012</a>).In another original research article included in this series, a miRNA sponge was used to block the silencing activity of miR-9 to study the functional role of miR-9 (<a href="#B12">Otaegi et al., 2012</a>). The authors demonstrated that blocking endogenous miR-9 allows for stronger expression of FoxP1 and a mild reduction in Lhx3-expressing motor neurons, suggesting the involvement of miRNAs in the fine-tuning process of motor neuron subtype identity specification (<a href="#B12">Otaegi et al., 2012</a>).Using miRNA genome-wide array profiling combined with bioinformatics analysis, Gao et al. identified a collection of miRNAs dynamically regulated by the zinc-finger protein, REST, during neuronal differentiation of primary mouse NSCs. In this study, the authors found that REST is critical for the proper expression of certain miRNAs under varying differentiation conditions, which in turn, provides insight into the REST-regulated stage-specific expression of miRNAs during NSC-neuronal differentiation (<a href="#B6">Gao et al., 2012</a>). Studies such as these help us to unfold the role that miRNAs play in neurogenesis.In conclusion, the study of neurogenesis is a poignant area of study, particularly because of the difficulties associated with modeling human brain diseases and injuries. Studies of key molecules that regulate the dynamic interactions in the brain, particularly miRNAs, present an opportunity to control and fine-tune NSC populations and cell fate conversion, as they are capable of intricate balance and regulation. Together, these articles reflect a sophisticated and exciting story of the dynamic roles miRNAs play in NSCs and neurogenesis. Further elucidation of the roles of small non-coding RNAs in brain development will serve as a powerful tool in brain disease modeling and therapeutics.We thank Drs. C. Fitzsimons, Q. R. Liu, J. Hsieh, and T. Sun for comments, C. Roberts for editing the manuscript. This work was supported by NIH NINDS R01 NS059546, RC1 NS068370, and California Institute for Regenerative Medicine TR2-01832.

The little molecules that could: a story about microRNAs in neural stem cells and neurogenesis

MicroRNAs (miRNAs) are short non-coding RNAs that regulate gene expression at the post-transcriptional level by mediating mRNA degradation or translational inhibition. MiRNAs are implicated in many biological functions, including neurogenesis. It has been shown that miRNAs regulate multiple steps of neurogenesis, from neural stem cell proliferation to neuronal differentiation and maturation. MiRNAs execute their functions in a dynamic and context-dependent manner by targeting diverse downstream target genes, from transcriptional factors to epigenetic regulators. Identifying context-specific target genes is instrumental for understanding the roles that miRNAs play in neurogenesis. This review summarizes our current state of knowledge on the dynamic roles that miRNAs play in neural stem cells and neurogenesis.

MicroRNAs (miRNAs) are short non-coding RNAs that regulate gene expression at the post-transcriptional level by mediating mRNA degradation or translational inhibition. MiRNAs are implicated in many biological functions, including neurogenesis. It has been shown that miRNAs regulate multiple steps of neurogenesis, from neural stem cell proliferation to neuronal differentiation and maturation. MiRNAs execute their functions in a dynamic and context-dependent manner by targeting diverse downstream target genes, from transcriptional factors to epigenetic regulators. Identifying context-specific target genes is instrumental for understanding the roles that miRNAs play in neurogenesis. This review summarizes our current state of knowledge on the dynamic roles that miRNAs play in neural stem cells and neurogenesis.

Dynamic Roles of microRNAs in Neurogenesis

Diversification of mammalian spinal motor neurons into hundreds of subtypes is critical for the maintenance of body posture and coordination of complex movements. Motor neuron differentiation is controlled by extrinsic signals that regulate intrinsic genetic programs specifying and consolidating motor neuron subtype identity. While transcription factors have been recognized as principal regulators of the intrinsic program, the role of posttranscriptional regulations has not been systematically tested. MicroRNAs produced by Dicer mediated cleavage of RNA hairpins contribute to gene regulation by posttranscriptional silencing. Here we used Olig2-cre conditional deletion of Dicer gene in motor neuron progenitors to examine effects of miRNA biogenesis disruption on postmitotic spinal motor neurons. We report that despite the initial increase in the number of motor neuron progenitors, disruption of Dicer function results in a loss of many limb- and sympathetic ganglia-innervating spinal motor neurons. Furthermore, it leads to defects in motor pool identity specification. Thus, our results indicate that miRNAs are an integral part of the genetic program controlling motor neuron survival and acquisition of subtype specific properties.

Diversification of mammalian spinal motor neurons into hundreds of subtypes is critical for the maintenance of body posture and coordination of complex movements. Motor neuron differentiation is controlled by extrinsic signals that regulate intrinsic genetic programs specifying and consolidating motor neuron subtype identity. While transcription factors have been recognized as principal regulators of the intrinsic program, the role of posttranscriptional regulations has not been systematically tested. MicroRNAs produced by Dicer mediated cleavage of RNA hairpins contribute to gene regulation by posttranscriptional silencing. Here we used Olig2-cre conditional deletion of Dicer gene in motor neuron progenitors to examine effects of miRNA biogenesis disruption on postmitotic spinal motor neurons. We report that despite the initial increase in the number of motor neuron progenitors, disruption of Dicer function results in a loss of many limb- and sympathetic ganglia-innervating spinal motor neurons. Furthermore, it leads to defects in motor pool identity specification. Thus, our results indicate that miRNAs are an integral part of the genetic program controlling motor neuron survival and acquisition of subtype specific properties.

Apoptosis of Limb Innervating Motor Neurons and Erosion of Motor Pool Identity upon Lineage Specific Dicer Inactivation

Multipotent neural stem cells (NSCs) possess the ability to self-renew and differentiate into both neurons and glia. However, the detailed mechanisms underlying NSC fate decisions are not well understood. Recent work suggests that the interaction between cell type specific transcription factors and microRNAs (miRNAs) is important as resident neural stem/progenitor cells give rise to functionally mature neurons. Recently, we demonstrated that the transcriptional repressor REST (RE1-silencing transcription factor) is essential to prevent precocious neuronal differentiation and maintain NSC self-renewal in the adult hippocampus. Here we show that REST is required for orchestrating the expression of distinct subsets of miRNAs in primary mouse NSC cultures, a physiologically relevant cell type. Using miRNA array profiling, we identified known REST-regulated miRNA genes, as well as previously uncharacterized REST-dependent miRNAs. Interestingly, in response to proliferation and differentiation stimuli, REST-regulated miRNAs formed distinct clusters and displayed variable expression dynamics. These results suggest that REST functions in a context-dependent manner through its target miRNAs for mediating neuronal production.

Multipotent neural stem cells (NSCs) possess the ability to self-renew and differentiate into both neurons and glia. However, the detailed mechanisms underlying NSC fate decisions are not well understood. Recent work suggests that the interaction between cell type specific transcription factors and microRNAs (miRNAs) is important as resident neural stem/progenitor cells give rise to functionally mature neurons. Recently, we demonstrated that the transcriptional repressor REST (RE1-silencing transcription factor) is essential to prevent precocious neuronal differentiation and maintain NSC self-renewal in the adult hippocampus. Here we show that REST is required for orchestrating the expression of distinct subsets of miRNAs in primary mouse NSC cultures, a physiologically relevant cell type. Using miRNA array profiling, we identified known REST-regulated miRNA genes, as well as previously uncharacterized REST-dependent miRNAs. Interestingly, in response to proliferation and differentiation stimuli, REST-regulated miRNAs formed distinct clusters and displayed variable expression dynamics. These results suggest that REST functions in a context-dependent manner through its target miRNAs for mediating neuronal production.

Profiling of REST-Dependent microRNAs Reveals Dynamic Modes of Expression

The process of neurogenesis includes neural stem cell proliferation, fate specification, young neuron migration, neuronal maturation, and functional integration into existing circuits. Although neurogenesis occurs largely during embryonic development, low levels but functionally important neurogenesis persists in restricted regions of the postnatal brain, including the subgranular zone of the dentate gyrus in the hippocampus and the subventricular zone of the lateral ventricles. This review will cover both embryonic and adult neurogenesis with an emphasis on the latter. Of the many endogenous mediators of postnatal neurogenesis, epigenetic pathways, such as mediators of DNA methylation, chromatin remodeling systems, and non-coding RNA modulators, appear to play an integral role. Mounting evidence shows that such epigenetic factors form regulatory networks, which govern each step of postnatal neurogenesis. In this review, we explore the emerging roles of epigenetic mechanisms particularly microRNAs, element-1 silencing transcription factor/neuron-restrictive silencing factor (REST/NRSF), polycomb proteins, and methyl-CpG bindings proteins, in regulating the entire process of postnatal and adult neurogenesis. We further summarize recent data regarding how the crosstalk among these different epigenetic proteins forms the critical regulatory network that regulates neuronal development. We finally discuss how crosstalk between these pathways may serve to translate environmental cues into control of the neurogenic process.

The process of neurogenesis includes neural stem cell proliferation, fate specification, young neuron migration, neuronal maturation, and functional integration into existing circuits. Although neurogenesis occurs largely during embryonic development, low levels but functionally important neurogenesis persists in restricted regions of the postnatal brain, including the subgranular zone of the dentate gyrus in the hippocampus and the subventricular zone of the lateral ventricles. This review will cover both embryonic and adult neurogenesis with an emphasis on the latter. Of the many endogenous mediators of postnatal neurogenesis, epigenetic pathways, such as mediators of DNA methylation, chromatin remodeling systems, and non-coding RNA modulators, appear to play an integral role. Mounting evidence shows that such epigenetic factors form regulatory networks, which govern each step of postnatal neurogenesis. In this review, we explore the emerging roles of epigenetic mechanisms particularly microRNAs, element-1 silencing transcription factor/neuron-restrictive silencing factor (REST/NRSF), polycomb proteins, and methyl-CpG bindings proteins, in regulating the entire process of postnatal and adult neurogenesis. We further summarize recent data regarding how the crosstalk among these different epigenetic proteins forms the critical regulatory network that regulates neuronal development. We finally discuss how crosstalk between these pathways may serve to translate environmental cues into control of the neurogenic process.

Crosstalk among Epigenetic Pathways Regulates Neurogenesis

MicroRNA (miRNA) is a type of short-length (~22 nt) non-coding RNA. Most miRNAs are transcribed by RNA polymerase II and processed by Drosha-DGCR8 and Dicer complexes in the cropping and dicing steps, respectively. miRNAs are exported by exportin-5 from the nucleus to the cytoplasm after cropping. Trimmed mature miRNA is loaded and targets mRNA at the 3′ or 5′ untranslated region (UTR) by recognition of base-pairing in the miRNA-loaded RISC, where it is involved in gene silencing including translational repression and/or degradation along with deadenylation. Recent studies have shown that miRNA participates in various biological functions including cell fate decision, developmental timing regulation, apoptosis, and tumorigenesis. Analyses of miRNA expression profiles have demonstrated tissue- and stage-specific miRNAs including the let-7 family, miR-124, and miR-9, which regulate the differentiation of embryonic stem cells and/or neurogenesis. This review focuses on RNA-binding protein-mediated miRNA biogenesis during neurogenesis. These miRNA biogenesis-relating proteins have also been linked to human diseases because their mutations can cause several nervous system disorders. Moreover, defects in core proteins involved in miRNA biogenesis including Drosha, DGCR8, and Dicer promote tumorigenesis. Thus, the study of not only mature miRNA function but also miRNA biogenesis steps is likely to be important.

MicroRNA (miRNA) is a type of short-length (~22 nt) non-coding RNA. Most miRNAs are transcribed by RNA polymerase II and processed by Drosha-DGCR8 and Dicer complexes in the cropping and dicing steps, respectively. miRNAs are exported by exportin-5 from the nucleus to the cytoplasm after cropping. Trimmed mature miRNA is loaded and targets mRNA at the 3′ or 5′ untranslated region (UTR) by recognition of base-pairing in the miRNA-loaded RISC, where it is involved in gene silencing including translational repression and/or degradation along with deadenylation. Recent studies have shown that miRNA participates in various biological functions including cell fate decision, developmental timing regulation, apoptosis, and tumorigenesis. Analyses of miRNA expression profiles have demonstrated tissue- and stage-specific miRNAs including the let-7 family, miR-124, and miR-9, which regulate the differentiation of embryonic stem cells and/or neurogenesis. This review focuses on RNA-binding protein-mediated miRNA biogenesis during neurogenesis. These miRNA biogenesis-relating proteins have also been linked to human diseases because their mutations can cause several nervous system disorders. Moreover, defects in core proteins involved in miRNA biogenesis including Drosha, DGCR8, and Dicer promote tumorigenesis. Thus, the study of not only mature miRNA function but also miRNA biogenesis steps is likely to be important.

MicroRNAs in Neural Stem Cells and Neurogenesis

Adult neurogenesis generates functional neurons from neural stem cells present in specific brain regions. It is largely confined to two main regions: the subventricular zone of the lateral ventricle, and the subgranular zone of the dentate gyrus (DG), in the hippocampus. With age, the function of the hippocampus and particularly the DG is impaired. For instance, adult neurogenesis is decreased with aging, in both proliferating and differentiation of newborn cells, while in parallel an age-associated decline in cognitive performance is often seen. Surprisingly, the synaptogenic potential of adult-born neurons is only marginally influenced by aging. Therefore, although proliferation, differentiation, and synaptogenesis of adult-born new neurons in the DG are closely related to each other, they are differentially affected by aging. In this review we discuss the crucial roles of a novel class of recently discovered modulators of gene expression, the small non-coding RNAs, in the regulation of adult neurogenesis. Multiple small non-coding RNAs are differentially expressed in the hippocampus. In particular a subgroup of the small non-coding RNAs, the microRNAs, fine-tune the progression of adult neurogenesis. This makes small non-coding RNAs appealing candidates to orchestrate the functional alterations in adult neurogenesis and cognition associated with aging. Finally, we summarize observations that link changes in circulating levels of steroid hormones with alterations in adult neurogenesis, cognitive decline, and vulnerability to psychopathology in advanced age, and discuss a potential interplay between steroid hormone receptors and microRNAs in cognitive decline in aging individuals.

Adult neurogenesis generates functional neurons from neural stem cells present in specific brain regions. It is largely confined to two main regions: the subventricular zone of the lateral ventricle, and the subgranular zone of the dentate gyrus (DG), in the hippocampus. With age, the function of the hippocampus and particularly the DG is impaired. For instance, adult neurogenesis is decreased with aging, in both proliferating and differentiation of newborn cells, while in parallel an age-associated decline in cognitive performance is often seen. Surprisingly, the synaptogenic potential of adult-born neurons is only marginally influenced by aging. Therefore, although proliferation, differentiation, and synaptogenesis of adult-born new neurons in the DG are closely related to each other, they are differentially affected by aging. In this review we discuss the crucial roles of a novel class of recently discovered modulators of gene expression, the small non-coding RNAs, in the regulation of adult neurogenesis. Multiple small non-coding RNAs are differentially expressed in the hippocampus. In particular a subgroup of the small non-coding RNAs, the microRNAs, fine-tune the progression of adult neurogenesis. This makes small non-coding RNAs appealing candidates to orchestrate the functional alterations in adult neurogenesis and cognition associated with aging. Finally, we summarize observations that link changes in circulating levels of steroid hormones with alterations in adult neurogenesis, cognitive decline, and vulnerability to psychopathology in advanced age, and discuss a potential interplay between steroid hormone receptors and microRNAs in cognitive decline in aging individuals.

New Neurons in Aging Brains: Molecular Control by Small Non-Coding RNAs

In adult mammals, neural stem cells (NSCs) are found in two niches of the brain; the subventricular zone by the lateral ventricles and the subgranular zone of the dentate gyrus in the hippocampus. Neurogenesis is a complex process that is tightly controlled on a molecular level. Recently, microRNAs (miRNAs) have been implicated to play a central role in the regulation of NCSs. miRNAs are small, endogenously expressed RNAs that regulate gene expression at the post-transcriptional level. However, functional studies of miRNAs are complicated due to current technical limitations. In this review we describe recent findings about miRNAs in NSCs looking closely at miR-124, miR-9, and let-7. In addition, we highlight technical strategies used to investigate miRNA function, accentuating limitations, and potentials.

In adult mammals, neural stem cells (NSCs) are found in two niches of the brain; the subventricular zone by the lateral ventricles and the subgranular zone of the dentate gyrus in the hippocampus. Neurogenesis is a complex process that is tightly controlled on a molecular level. Recently, microRNAs (miRNAs) have been implicated to play a central role in the regulation of NCSs. miRNAs are small, endogenously expressed RNAs that regulate gene expression at the post-transcriptional level. However, functional studies of miRNAs are complicated due to current technical limitations. In this review we describe recent findings about miRNAs in NSCs looking closely at miR-124, miR-9, and let-7. In addition, we highlight technical strategies used to investigate miRNA function, accentuating limitations, and potentials.

Functional Studies of microRNAs in Neural Stem Cells: Problems and Perspectives

Myelination of axons by oligodendrocytes in the central nervous system is essential for normal neuronal functions. The failure of remyelination due to injury or pathological insults results in devastating demyelinating diseases. Oligodendrocytes originate in restricted regions of the embryonic ventral neural tube. After migration to populate all areas of the brain and spinal cord, oligodendrocyte precursors undergo a temporally well-defined series of molecular and structural changes, ultimately culminating in the cessation of proliferation, and the elaboration of a highly complex myelin sheath. The emergence of microRNAs (miRNAs) as potent regulators of gene expression at the posttranscriptional level has broad implications in all facets of cell biology. Recent studies have demonstrated a critical role of miRNAs in oligodendrocyte development, including cell proliferation, differentiation, and myelin formation. In this review, we will highlight and discuss the recent understanding of functional links of miRNAs to regulatory networks for central myelination, as well as perspectives on the role of miRNAs in demyelinating diseases.

Myelination of axons by oligodendrocytes in the central nervous system is essential for normal neuronal functions. The failure of remyelination due to injury or pathological insults results in devastating demyelinating diseases. Oligodendrocytes originate in restricted regions of the embryonic ventral neural tube. After migration to populate all areas of the brain and spinal cord, oligodendrocyte precursors undergo a temporally well-defined series of molecular and structural changes, ultimately culminating in the cessation of proliferation, and the elaboration of a highly complex myelin sheath. The emergence of microRNAs (miRNAs) as potent regulators of gene expression at the posttranscriptional level has broad implications in all facets of cell biology. Recent studies have demonstrated a critical role of miRNAs in oligodendrocyte development, including cell proliferation, differentiation, and myelin formation. In this review, we will highlight and discuss the recent understanding of functional links of miRNAs to regulatory networks for central myelination, as well as perspectives on the role of miRNAs in demyelinating diseases.

Fine-Tuning Oligodendrocyte Development by microRNAs

The specification of motor neuron (MN) subtypes and columnar organization in developing spinal cord is controlled by multiple transcription factors. FoxP1 drives specification of lateral motor neuron (LMN) subtypes, and we demonstrated in our previous work that FoxP1 expression levels are regulated by the microRNA miR-9. Here we show that ectopic FoxP1 expression in the chick spinal cord can rescue Lhx3 and Hb9 expression in MNs altered by miR-9 over-expression, demonstrating that FoxP1 is a critical functional interaction partner for miR-9 in LMN development. Moreover, we have optimized a technique called a miRNA sponge in vitro, to permit easy discovery of the role of individual miRNA in vivo using a loss-of-function approach. We here show that narrow spacing between binding sites, inclusion of a coding gene, and optimizing the number of miRNA binding sites can significantly increase the blocking ability of a sponge. We go on to show that a miR-9 sponge reduces detectable miR-9 in the ventral horn, preventing miR-9 silencing of FoxP1 in vivo, and in turn modifies MN subtypes in the spinal cord. Our designs for optimized sponges provide a knockdown tool that is ready to be used to study the function of miRNA in vivo, and in particular for generating transgenic animal models.

The specification of motor neuron (MN) subtypes and columnar organization in developing spinal cord is controlled by multiple transcription factors. FoxP1 drives specification of lateral motor neuron (LMN) subtypes, and we demonstrated in our previous work that FoxP1 expression levels are regulated by the microRNA miR-9. Here we show that ectopic FoxP1 expression in the chick spinal cord can rescue Lhx3 and Hb9 expression in MNs altered by miR-9 over-expression, demonstrating that FoxP1 is a critical functional interaction partner for miR-9 in LMN development. Moreover, we have optimized a technique called a miRNA sponge in vitro, to permit easy discovery of the role of individual miRNA in vivo using a loss-of-function approach. We here show that narrow spacing between binding sites, inclusion of a coding gene, and optimizing the number of miRNA binding sites can significantly increase the blocking ability of a sponge. We go on to show that a miR-9 sponge reduces detectable miR-9 in the ventral horn, preventing miR-9 silencing of FoxP1 in vivo, and in turn modifies MN subtypes in the spinal cord. Our designs for optimized sponges provide a knockdown tool that is ready to be used to study the function of miRNA in vivo, and in particular for generating transgenic animal models.