Abstract
Although the function of posttranscriptional processes in regulating the expression of genes involved in muscle fiber formation (myogenesis) is well accepted, the mechanisms by which these effects are mediated remain elusive. Here, we uncover such a mechanism and show that during myogenesis, a fraction of the posttranscriptional regulator human antigen R (HuR) is cleaved in a caspase-dependent manner in both cell culture and animal models. Disruption of caspase activity in cultured myoblasts or knocking out the caspase-3 gene in mice significantly reduced HuR cleavage and the cytoplasmic accumulation of HuR in muscle fibers. The non-cleavable isoform of HuR, HuRD226A, failed to reestablish the myogenic potential of HuR-depleted myoblasts. HuR cleavage generates two fragments: HuR-cleavage product 1 (HuR-CP1) (24 kDa) and HuR-CP2 (8 kDa). Here, we show that one of these fragments (HuR-CP1) binds to the HuR import factor transportin-2 (TRN2) allowing HuR to accumulate in the cytoplasm. As this cytoplasmic accumulation is required for the promyogenic function of HuR, our data support a model, whereby during the transition phase from myoblasts to myotubes, a proportion of HuR is cleaved to generate HuR-CP1. By interfering with the TRN2-mediated import of HuR, this CP helps non-cleaved HuR accumulate in the cytoplasm thus promoting myogenesis.
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Abbreviations
- HuR:
-
human antigen R
- TRN2:
-
transportin-2
- MRFs:
-
myogenic regulatory factors
- AREs:
-
AU-rich elements
- HuR-CP:
-
HuR cleavage product
- caspase-CP:
-
caspase cleavage product
- AP:
-
antennapedia
- GFP:
-
green fluorescence protein
- Rec:
-
recombinant
- GST:
-
gluthation S-transferase
- RRM:
-
RNA recognition motif
- ActD:
-
actinomycin D
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Acknowledgements
We thank Drs S Di Marco and V Dormoy-Raclet for their helpful suggestions throughout the progress of this work. We also thank Ms C Clow for her technical help with the in vivo experiments, and Mr N Punjani for his assistance with the production of cleavable GST-plasmids and proteins. This work was supported by a Cole Foundation fellowship for PB, a FRSQ Master's fellowship for CN, a CIHR Doctoral Research Award for CvR, and a CIHR (MOP-89798) operating grant to IG. IG is a recipient of a Tier II Canada Research Chair.
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Beauchamp, P., Nassif, C., Hillock, S. et al. The cleavage of HuR interferes with its transportin-2-mediated nuclear import and promotes muscle fiber formation. Cell Death Differ 17, 1588–1599 (2010). https://doi.org/10.1038/cdd.2010.34
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DOI: https://doi.org/10.1038/cdd.2010.34
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