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The novel protein MANI modulates neurogenesis and neurite-cone growth.

Mishra M, Akatsu H, Heese K - J. Cell. Mol. Med. (2011)

Bottom Line: To date, three myelin-associated proteins [Nogo or reticulon 4 (RTN4), myelin-associated glycoprotein (MAG) and oligodendrocyte myelin glycoprotein (OMG)] are known to inhibit axonal regeneration via activation of the neuronal glycosylphosphatidylinositol-anchored Nogo receptor [NgR, together with p75 neurotrophin receptor (p75NTR) and Lingo-1].We show that knockdown of Cdc27, a component of the anaphase-promoting complex (APC), leads to enhanced neurite outgrowth.Our finding describes the novel MANI-Cdc27-APC pathway as an important cascade that prevents neurons from extending axons, thus providing implications for the potential treatment of neurodegenerative diseases.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cell Biology, School of Biological Sciences, College of Science, Nanyang Technological University, Singapore.

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Mani inhibits neurite outgrowth. Expression of Mani in PC12 cells inhibits Ngf (100 ng/ml)-induced neurite outgrowth and modulates the expression of pivotal cell signalling molecules. Control (mock/GFP transfected; left panel) and Mani-transfected (right panel) PC12 cells were incubated in the presence of Ngf to induce neurite outgrowth. Pictures were taken on the 6th and 12th day of Ngf stimulation. Whereas the control cells showed a normal neurite network, Mani-transfected cells exhibited no or little response to Ngf, as demonstrated by the significantly inhibited neurite outgrowth, even after 12 days. Scale bar = 50 μm (A). PC12 cells were grown and differentiated with Ngf, and lysates were prepared on the 6th and 12th day of induction, followed by Western blot analyses (C = control [mock/GFP transfected], Mani = Mani transfected). Mani-transfected cells displayed altered expression and phosphorylation (p-) levels of various signalling molecules. * indicates short-term induction with Ngf, as Stat-3-Ser-727 is transiently activated first, prior to Stat-3-Tyr-705 [23]. Mani itself was down-regulated upon Ngf stimulation (B). PC12 cells were grown and differentiated via application of Ngf for 10 days in the presence of either the Mani antibody or a random unspecific antibody (xAb). As shown in (ii), the presence of the ManiAb enhanced Ngf-induced neurite outgrowth. Scale bars = 50 μm (C). Statistical evaluation of the neurite outgrowth described in (C, i–iii). Data represent mean ± S.D. of three independent experiments each done in duplicates (*P < 0.05 compared with controls (no antibody added [C, i]) (D).
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fig07: Mani inhibits neurite outgrowth. Expression of Mani in PC12 cells inhibits Ngf (100 ng/ml)-induced neurite outgrowth and modulates the expression of pivotal cell signalling molecules. Control (mock/GFP transfected; left panel) and Mani-transfected (right panel) PC12 cells were incubated in the presence of Ngf to induce neurite outgrowth. Pictures were taken on the 6th and 12th day of Ngf stimulation. Whereas the control cells showed a normal neurite network, Mani-transfected cells exhibited no or little response to Ngf, as demonstrated by the significantly inhibited neurite outgrowth, even after 12 days. Scale bar = 50 μm (A). PC12 cells were grown and differentiated with Ngf, and lysates were prepared on the 6th and 12th day of induction, followed by Western blot analyses (C = control [mock/GFP transfected], Mani = Mani transfected). Mani-transfected cells displayed altered expression and phosphorylation (p-) levels of various signalling molecules. * indicates short-term induction with Ngf, as Stat-3-Ser-727 is transiently activated first, prior to Stat-3-Tyr-705 [23]. Mani itself was down-regulated upon Ngf stimulation (B). PC12 cells were grown and differentiated via application of Ngf for 10 days in the presence of either the Mani antibody or a random unspecific antibody (xAb). As shown in (ii), the presence of the ManiAb enhanced Ngf-induced neurite outgrowth. Scale bars = 50 μm (C). Statistical evaluation of the neurite outgrowth described in (C, i–iii). Data represent mean ± S.D. of three independent experiments each done in duplicates (*P < 0.05 compared with controls (no antibody added [C, i]) (D).

Mentions: Data obtained thus far prompted us to explore the possible signalling pathways involved in Mani-mediated morphological changes during the differentiation of catecholaminergic (Th+) neurons. Thus, Ngf was applied for 2 weeks to catecholaminergic PC12 cells and, surprisingly, even after 1 week of Ngf stimulation, PC12 cells transfected with Mani did not show any significant observable neurite outgrowth – instead, only very short neurite extensions could be seen on day 12 of induction (Fig. 7A).


The novel protein MANI modulates neurogenesis and neurite-cone growth.

Mishra M, Akatsu H, Heese K - J. Cell. Mol. Med. (2011)

Mani inhibits neurite outgrowth. Expression of Mani in PC12 cells inhibits Ngf (100 ng/ml)-induced neurite outgrowth and modulates the expression of pivotal cell signalling molecules. Control (mock/GFP transfected; left panel) and Mani-transfected (right panel) PC12 cells were incubated in the presence of Ngf to induce neurite outgrowth. Pictures were taken on the 6th and 12th day of Ngf stimulation. Whereas the control cells showed a normal neurite network, Mani-transfected cells exhibited no or little response to Ngf, as demonstrated by the significantly inhibited neurite outgrowth, even after 12 days. Scale bar = 50 μm (A). PC12 cells were grown and differentiated with Ngf, and lysates were prepared on the 6th and 12th day of induction, followed by Western blot analyses (C = control [mock/GFP transfected], Mani = Mani transfected). Mani-transfected cells displayed altered expression and phosphorylation (p-) levels of various signalling molecules. * indicates short-term induction with Ngf, as Stat-3-Ser-727 is transiently activated first, prior to Stat-3-Tyr-705 [23]. Mani itself was down-regulated upon Ngf stimulation (B). PC12 cells were grown and differentiated via application of Ngf for 10 days in the presence of either the Mani antibody or a random unspecific antibody (xAb). As shown in (ii), the presence of the ManiAb enhanced Ngf-induced neurite outgrowth. Scale bars = 50 μm (C). Statistical evaluation of the neurite outgrowth described in (C, i–iii). Data represent mean ± S.D. of three independent experiments each done in duplicates (*P < 0.05 compared with controls (no antibody added [C, i]) (D).
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fig07: Mani inhibits neurite outgrowth. Expression of Mani in PC12 cells inhibits Ngf (100 ng/ml)-induced neurite outgrowth and modulates the expression of pivotal cell signalling molecules. Control (mock/GFP transfected; left panel) and Mani-transfected (right panel) PC12 cells were incubated in the presence of Ngf to induce neurite outgrowth. Pictures were taken on the 6th and 12th day of Ngf stimulation. Whereas the control cells showed a normal neurite network, Mani-transfected cells exhibited no or little response to Ngf, as demonstrated by the significantly inhibited neurite outgrowth, even after 12 days. Scale bar = 50 μm (A). PC12 cells were grown and differentiated with Ngf, and lysates were prepared on the 6th and 12th day of induction, followed by Western blot analyses (C = control [mock/GFP transfected], Mani = Mani transfected). Mani-transfected cells displayed altered expression and phosphorylation (p-) levels of various signalling molecules. * indicates short-term induction with Ngf, as Stat-3-Ser-727 is transiently activated first, prior to Stat-3-Tyr-705 [23]. Mani itself was down-regulated upon Ngf stimulation (B). PC12 cells were grown and differentiated via application of Ngf for 10 days in the presence of either the Mani antibody or a random unspecific antibody (xAb). As shown in (ii), the presence of the ManiAb enhanced Ngf-induced neurite outgrowth. Scale bars = 50 μm (C). Statistical evaluation of the neurite outgrowth described in (C, i–iii). Data represent mean ± S.D. of three independent experiments each done in duplicates (*P < 0.05 compared with controls (no antibody added [C, i]) (D).
Mentions: Data obtained thus far prompted us to explore the possible signalling pathways involved in Mani-mediated morphological changes during the differentiation of catecholaminergic (Th+) neurons. Thus, Ngf was applied for 2 weeks to catecholaminergic PC12 cells and, surprisingly, even after 1 week of Ngf stimulation, PC12 cells transfected with Mani did not show any significant observable neurite outgrowth – instead, only very short neurite extensions could be seen on day 12 of induction (Fig. 7A).

Bottom Line: To date, three myelin-associated proteins [Nogo or reticulon 4 (RTN4), myelin-associated glycoprotein (MAG) and oligodendrocyte myelin glycoprotein (OMG)] are known to inhibit axonal regeneration via activation of the neuronal glycosylphosphatidylinositol-anchored Nogo receptor [NgR, together with p75 neurotrophin receptor (p75NTR) and Lingo-1].We show that knockdown of Cdc27, a component of the anaphase-promoting complex (APC), leads to enhanced neurite outgrowth.Our finding describes the novel MANI-Cdc27-APC pathway as an important cascade that prevents neurons from extending axons, thus providing implications for the potential treatment of neurodegenerative diseases.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cell Biology, School of Biological Sciences, College of Science, Nanyang Technological University, Singapore.

Show MeSH
Related in: MedlinePlus