Limits...
A feedback regulatory loop involving microRNA-9 and nuclear receptor TLX in neural stem cell fate determination.

Zhao C, Sun G, Li S, Shi Y - Nat. Struct. Mol. Biol. (2009)

Bottom Line: Here we show that miR-9 suppresses TLX expression to negatively regulate neural stem cell proliferation and accelerate neural differentiation.Moreover, TLX represses expression of the miR-9 pri-miRNA.By forming a negative regulatory loop with TLX, miR-9 provides a model for controlling the balance between neural stem cell proliferation and differentiation.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurosciences, Center for Gene Expression and Drug Discovery, Beckman Research Institute of City of Hope, Duarte, California, USA.

ABSTRACT
MicroRNAs have been implicated as having important roles in stem cell biology. MicroRNA-9 (miR-9) is expressed specifically in neurogenic areas of the brain and may be involved in neural stem cell self-renewal and differentiation. We showed previously that the nuclear receptor TLX is an essential regulator of neural stem cell self-renewal. Here we show that miR-9 suppresses TLX expression to negatively regulate neural stem cell proliferation and accelerate neural differentiation. Introducing a TLX expression vector that is not prone to miR-9 regulation rescued miR-9-induced proliferation deficiency and inhibited precocious differentiation. In utero electroporation of miR-9 in embryonic brains led to premature differentiation and outward migration of the transfected neural stem cells. Moreover, TLX represses expression of the miR-9 pri-miRNA. By forming a negative regulatory loop with TLX, miR-9 provides a model for controlling the balance between neural stem cell proliferation and differentiation.

Show MeSH
TLXΔ3′ UTR rescues miR-9-induced neural stem cell proliferation deficiencya. RT-PCR analysis of TLXΔ3′ UTR and total TLX expression in control neural stem cells (C) and TLXΔ3′ UTR-expressing cells treated with control RNA (- miR-9) or miR-9 (+ miR-9). Actin was included as a loading control. b. Control (C) or TLXΔ3′ UTR-expressing cells were transfected with control RNA (-miR-9) or miR-9 followed by BrdU labeling (green). Merged panels show BrdU staining along with phase contrast images. c. Quantification of BrdU-positive (BrdU+) cells in control (C) and TLXΔ3′ UTR-expressing cells treated with control RNA (-miR-9) or miR-9. Error bars are s.d. of the mean. * p=0.003 by Student's t-test. ** p=0.04 by Student's t-test. d. Quantitation of GFAP-positive (GFAP+) cells in control (C) and TLXΔ3′ UTR-expressing cells treated with control RNA (-miR-9) or miR-9. Error bars are s.d. of the mean. * p=0.02 by Student's t-test. GFP siRNA was included as the control RNA.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2667220&req=5

Figure 3: TLXΔ3′ UTR rescues miR-9-induced neural stem cell proliferation deficiencya. RT-PCR analysis of TLXΔ3′ UTR and total TLX expression in control neural stem cells (C) and TLXΔ3′ UTR-expressing cells treated with control RNA (- miR-9) or miR-9 (+ miR-9). Actin was included as a loading control. b. Control (C) or TLXΔ3′ UTR-expressing cells were transfected with control RNA (-miR-9) or miR-9 followed by BrdU labeling (green). Merged panels show BrdU staining along with phase contrast images. c. Quantification of BrdU-positive (BrdU+) cells in control (C) and TLXΔ3′ UTR-expressing cells treated with control RNA (-miR-9) or miR-9. Error bars are s.d. of the mean. * p=0.003 by Student's t-test. ** p=0.04 by Student's t-test. d. Quantitation of GFAP-positive (GFAP+) cells in control (C) and TLXΔ3′ UTR-expressing cells treated with control RNA (-miR-9) or miR-9. Error bars are s.d. of the mean. * p=0.02 by Student's t-test. GFP siRNA was included as the control RNA.

Mentions: To determine whether the effect of miR-9 transfection on neural stem cell proliferation and differentiation is mediated through TLX, neural stem cells were stably transduced with a TLX-expressing vector (TLXΔ3′ UTR), which lacks the TLX 3′ UTR. Transfection of miR-9 had no effect on the expression of TLXΔ3′ UTR, although miR-9 downregulated endogenous TLX expression levels (Fig. 3a). Expression of TLXΔ3′ UTR led to 1.24-fold increase in neural stem cell proliferation. Co-transfection of TLXΔ3′ UTR and miR-9 reversed the proliferative deficiency induced by miR-9 substantially (Fig. 3b, c). Furthermore, while transfection of miR-9 increased astroglial differentiation in control neural stem cells, no appreciable increase in astrocyte differentiation was detected in TLXΔ3′ UTR-transduced cells upon miR-9 treatment (Fig. 3d). These results strongly suggest that miR-9 regulates neural stem cell proliferation and differentiation, at least in part, by inhibiting TLX expression through its 3′ UTR.


A feedback regulatory loop involving microRNA-9 and nuclear receptor TLX in neural stem cell fate determination.

Zhao C, Sun G, Li S, Shi Y - Nat. Struct. Mol. Biol. (2009)

TLXΔ3′ UTR rescues miR-9-induced neural stem cell proliferation deficiencya. RT-PCR analysis of TLXΔ3′ UTR and total TLX expression in control neural stem cells (C) and TLXΔ3′ UTR-expressing cells treated with control RNA (- miR-9) or miR-9 (+ miR-9). Actin was included as a loading control. b. Control (C) or TLXΔ3′ UTR-expressing cells were transfected with control RNA (-miR-9) or miR-9 followed by BrdU labeling (green). Merged panels show BrdU staining along with phase contrast images. c. Quantification of BrdU-positive (BrdU+) cells in control (C) and TLXΔ3′ UTR-expressing cells treated with control RNA (-miR-9) or miR-9. Error bars are s.d. of the mean. * p=0.003 by Student's t-test. ** p=0.04 by Student's t-test. d. Quantitation of GFAP-positive (GFAP+) cells in control (C) and TLXΔ3′ UTR-expressing cells treated with control RNA (-miR-9) or miR-9. Error bars are s.d. of the mean. * p=0.02 by Student's t-test. GFP siRNA was included as the control RNA.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2667220&req=5

Figure 3: TLXΔ3′ UTR rescues miR-9-induced neural stem cell proliferation deficiencya. RT-PCR analysis of TLXΔ3′ UTR and total TLX expression in control neural stem cells (C) and TLXΔ3′ UTR-expressing cells treated with control RNA (- miR-9) or miR-9 (+ miR-9). Actin was included as a loading control. b. Control (C) or TLXΔ3′ UTR-expressing cells were transfected with control RNA (-miR-9) or miR-9 followed by BrdU labeling (green). Merged panels show BrdU staining along with phase contrast images. c. Quantification of BrdU-positive (BrdU+) cells in control (C) and TLXΔ3′ UTR-expressing cells treated with control RNA (-miR-9) or miR-9. Error bars are s.d. of the mean. * p=0.003 by Student's t-test. ** p=0.04 by Student's t-test. d. Quantitation of GFAP-positive (GFAP+) cells in control (C) and TLXΔ3′ UTR-expressing cells treated with control RNA (-miR-9) or miR-9. Error bars are s.d. of the mean. * p=0.02 by Student's t-test. GFP siRNA was included as the control RNA.
Mentions: To determine whether the effect of miR-9 transfection on neural stem cell proliferation and differentiation is mediated through TLX, neural stem cells were stably transduced with a TLX-expressing vector (TLXΔ3′ UTR), which lacks the TLX 3′ UTR. Transfection of miR-9 had no effect on the expression of TLXΔ3′ UTR, although miR-9 downregulated endogenous TLX expression levels (Fig. 3a). Expression of TLXΔ3′ UTR led to 1.24-fold increase in neural stem cell proliferation. Co-transfection of TLXΔ3′ UTR and miR-9 reversed the proliferative deficiency induced by miR-9 substantially (Fig. 3b, c). Furthermore, while transfection of miR-9 increased astroglial differentiation in control neural stem cells, no appreciable increase in astrocyte differentiation was detected in TLXΔ3′ UTR-transduced cells upon miR-9 treatment (Fig. 3d). These results strongly suggest that miR-9 regulates neural stem cell proliferation and differentiation, at least in part, by inhibiting TLX expression through its 3′ UTR.

Bottom Line: Here we show that miR-9 suppresses TLX expression to negatively regulate neural stem cell proliferation and accelerate neural differentiation.Moreover, TLX represses expression of the miR-9 pri-miRNA.By forming a negative regulatory loop with TLX, miR-9 provides a model for controlling the balance between neural stem cell proliferation and differentiation.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurosciences, Center for Gene Expression and Drug Discovery, Beckman Research Institute of City of Hope, Duarte, California, USA.

ABSTRACT
MicroRNAs have been implicated as having important roles in stem cell biology. MicroRNA-9 (miR-9) is expressed specifically in neurogenic areas of the brain and may be involved in neural stem cell self-renewal and differentiation. We showed previously that the nuclear receptor TLX is an essential regulator of neural stem cell self-renewal. Here we show that miR-9 suppresses TLX expression to negatively regulate neural stem cell proliferation and accelerate neural differentiation. Introducing a TLX expression vector that is not prone to miR-9 regulation rescued miR-9-induced proliferation deficiency and inhibited precocious differentiation. In utero electroporation of miR-9 in embryonic brains led to premature differentiation and outward migration of the transfected neural stem cells. Moreover, TLX represses expression of the miR-9 pri-miRNA. By forming a negative regulatory loop with TLX, miR-9 provides a model for controlling the balance between neural stem cell proliferation and differentiation.

Show MeSH