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NDRG2 promotes myoblast proliferation and caspase 3/7 activities during differentiation, and attenuates hydrogen peroxide - But not palmitate-induced toxicity.

Anderson KJ, Russell AP, Foletta VC - FEBS Open Bio (2015)

Bottom Line: NDRG2 also attenuated apoptosis by reducing cleavage of PARP and caspase 3 and expression of pro-apoptotic Bax while enhancing the pro-survival Bcl-2 and Bcl-xL levels.In contrast, Mcl-1 was not altered, and NDRG2 did not protect against palmitate-induced lipotoxicity.Furthermore, NDRG2 attenuates H2O2-induced oxidative stress and specific serine and threonine amino acid residues appear to contribute to its function in muscle cells.

View Article: PubMed Central - PubMed

Affiliation: Centre for Physical Activity and Nutrition Research (C-PAN), School of Exercise and Nutrition Sciences, Faculty of Health, Deakin University, Melbourne, Australia.

ABSTRACT
The function of the stress-responsive N-myc downstream-regulated gene 2 (NDRG2) in the control of myoblast growth, and the amino acids contributing to its function, are not well characterized. Here, we investigated the effect of increased NDRG2 levels on the proliferation, differentiation and apoptosis in skeletal muscle cells under basal and stress conditions. NDRG2 overexpression increased C2C12 myoblast proliferation and the expression of positive cell cycle regulators, cdk2, cyclin B and cyclin D, and phosphorylation of Rb, while the serine/threonine-deficient NDRG2, 3A-NDRG2, had less effect. The onset of differentiation was enhanced by NDRG2 as determined through the myogenic regulatory factor expression profiles and myocyte fusion index. However, the overall level of differentiation in myotubes was not different. While NDRG2 up-regulated caspase 3/7 activities during differentiation, no increase in apoptosis was measured by TUNEL assay or through cleavage of caspase 3 and PARP proteins. During H2O2 treatment to induce oxidative stress, NDRG2 helped protect against the loss of proliferation and ER stress as measured by GRP78 expression with 3A-NDRG2 displaying less protection. NDRG2 also attenuated apoptosis by reducing cleavage of PARP and caspase 3 and expression of pro-apoptotic Bax while enhancing the pro-survival Bcl-2 and Bcl-xL levels. In contrast, Mcl-1 was not altered, and NDRG2 did not protect against palmitate-induced lipotoxicity. Our findings show that NDRG2 overexpression increases myoblast proliferation and caspase 3/7 activities without increasing overall differentiation. Furthermore, NDRG2 attenuates H2O2-induced oxidative stress and specific serine and threonine amino acid residues appear to contribute to its function in muscle cells.

No MeSH data available.


Related in: MedlinePlus

The expression of negative cell cycle regulators following NDRG2 and 3A-NDRG2 overexpression in myoblasts at days 2 and 3 of proliferation (P2 and P3) and at confluence (D0). (A) p27 mRNA (upper panel) and protein (lower panel) expression, and (B) p21 mRNA (upper panel) and protein (lower panel) expression with representative blots indicated. Data are mean of three independent experiments (n = 3 per treatment). HSP90 expression indicates protein levels loaded. ***P < 0.001 and *P < 0.05 compared to vector; ###P < 0.001 compared to NDRG2.
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f0015: The expression of negative cell cycle regulators following NDRG2 and 3A-NDRG2 overexpression in myoblasts at days 2 and 3 of proliferation (P2 and P3) and at confluence (D0). (A) p27 mRNA (upper panel) and protein (lower panel) expression, and (B) p21 mRNA (upper panel) and protein (lower panel) expression with representative blots indicated. Data are mean of three independent experiments (n = 3 per treatment). HSP90 expression indicates protein levels loaded. ***P < 0.001 and *P < 0.05 compared to vector; ###P < 0.001 compared to NDRG2.

Mentions: The expression of the cell cycle inhibitors p27 Kip1 (p27) and p21 were also investigated. Following NDRG2 overexpression p27 mRNA levels were reduced 1.4-fold (P < 0.05) at P2 with a corresponding 2-fold suppression of its protein levels at P3 (P < 0.05, Fig. 3A). In contrast, no difference in p21 mRNA was observed at P2 or P3. However, when myoblasts reached confluence (D0), p21 mRNA and protein levels increased 1.7-fold with NDRG2 treatment (P < 0.001; Fig. 3B). No effect of 3A-NDRG2 was observed for any of the mRNA or protein targets measured.


NDRG2 promotes myoblast proliferation and caspase 3/7 activities during differentiation, and attenuates hydrogen peroxide - But not palmitate-induced toxicity.

Anderson KJ, Russell AP, Foletta VC - FEBS Open Bio (2015)

The expression of negative cell cycle regulators following NDRG2 and 3A-NDRG2 overexpression in myoblasts at days 2 and 3 of proliferation (P2 and P3) and at confluence (D0). (A) p27 mRNA (upper panel) and protein (lower panel) expression, and (B) p21 mRNA (upper panel) and protein (lower panel) expression with representative blots indicated. Data are mean of three independent experiments (n = 3 per treatment). HSP90 expression indicates protein levels loaded. ***P < 0.001 and *P < 0.05 compared to vector; ###P < 0.001 compared to NDRG2.
© Copyright Policy - CC BY-NC-ND
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4556729&req=5

f0015: The expression of negative cell cycle regulators following NDRG2 and 3A-NDRG2 overexpression in myoblasts at days 2 and 3 of proliferation (P2 and P3) and at confluence (D0). (A) p27 mRNA (upper panel) and protein (lower panel) expression, and (B) p21 mRNA (upper panel) and protein (lower panel) expression with representative blots indicated. Data are mean of three independent experiments (n = 3 per treatment). HSP90 expression indicates protein levels loaded. ***P < 0.001 and *P < 0.05 compared to vector; ###P < 0.001 compared to NDRG2.
Mentions: The expression of the cell cycle inhibitors p27 Kip1 (p27) and p21 were also investigated. Following NDRG2 overexpression p27 mRNA levels were reduced 1.4-fold (P < 0.05) at P2 with a corresponding 2-fold suppression of its protein levels at P3 (P < 0.05, Fig. 3A). In contrast, no difference in p21 mRNA was observed at P2 or P3. However, when myoblasts reached confluence (D0), p21 mRNA and protein levels increased 1.7-fold with NDRG2 treatment (P < 0.001; Fig. 3B). No effect of 3A-NDRG2 was observed for any of the mRNA or protein targets measured.

Bottom Line: NDRG2 also attenuated apoptosis by reducing cleavage of PARP and caspase 3 and expression of pro-apoptotic Bax while enhancing the pro-survival Bcl-2 and Bcl-xL levels.In contrast, Mcl-1 was not altered, and NDRG2 did not protect against palmitate-induced lipotoxicity.Furthermore, NDRG2 attenuates H2O2-induced oxidative stress and specific serine and threonine amino acid residues appear to contribute to its function in muscle cells.

View Article: PubMed Central - PubMed

Affiliation: Centre for Physical Activity and Nutrition Research (C-PAN), School of Exercise and Nutrition Sciences, Faculty of Health, Deakin University, Melbourne, Australia.

ABSTRACT
The function of the stress-responsive N-myc downstream-regulated gene 2 (NDRG2) in the control of myoblast growth, and the amino acids contributing to its function, are not well characterized. Here, we investigated the effect of increased NDRG2 levels on the proliferation, differentiation and apoptosis in skeletal muscle cells under basal and stress conditions. NDRG2 overexpression increased C2C12 myoblast proliferation and the expression of positive cell cycle regulators, cdk2, cyclin B and cyclin D, and phosphorylation of Rb, while the serine/threonine-deficient NDRG2, 3A-NDRG2, had less effect. The onset of differentiation was enhanced by NDRG2 as determined through the myogenic regulatory factor expression profiles and myocyte fusion index. However, the overall level of differentiation in myotubes was not different. While NDRG2 up-regulated caspase 3/7 activities during differentiation, no increase in apoptosis was measured by TUNEL assay or through cleavage of caspase 3 and PARP proteins. During H2O2 treatment to induce oxidative stress, NDRG2 helped protect against the loss of proliferation and ER stress as measured by GRP78 expression with 3A-NDRG2 displaying less protection. NDRG2 also attenuated apoptosis by reducing cleavage of PARP and caspase 3 and expression of pro-apoptotic Bax while enhancing the pro-survival Bcl-2 and Bcl-xL levels. In contrast, Mcl-1 was not altered, and NDRG2 did not protect against palmitate-induced lipotoxicity. Our findings show that NDRG2 overexpression increases myoblast proliferation and caspase 3/7 activities without increasing overall differentiation. Furthermore, NDRG2 attenuates H2O2-induced oxidative stress and specific serine and threonine amino acid residues appear to contribute to its function in muscle cells.

No MeSH data available.


Related in: MedlinePlus