GATA4 in satellite cells is involved in skeletal muscle regeneration
Tomohide Takaya, Atsushi Asakura.
Stem Cell institute, Paul & Sheila Wellstone Muscular Dystrophy Center, Department of Neurology, University of Minnesota Medical School, Minneapolis, MN, USA.
The Greg Marzolf Jr Symposium 2014 (Minneapolis, USA), 2014/12/04 (Poster).
Abstract
GATA factors are indispensable transcription factors for proliferation and differentiation in various stem cells. GATA1/2/3 determine the cell fate of hematopoietic stem cells, and GATA4/5/6 have been well studied in cardiovascular development. However, the roles of GATA factors in satellite cells during skeletal muscle regeneration have not been clarified.
GATA4 was not expressed in Pax7+ quiescent satellite cells but appeared in Pax7+/MyoD+ activated satellite cells on myofibers. In cultured myoblasts, GATA4 was colocalized with Pax7 and MyoD in growth condition then eventually disappeared in MyoD+/myogenin+ differentiated myotubes. Overexpression and knockout experiments showed that GATA4 promotes proliferation and inhibits differentiation of myoblasts. Mutational and pharmacological assays indicated that p300-induced acetylation is required for GATA4 function. Acetylation of GATA4 improves its DNA-binding activity. qPCR and microarray data demonstrated that GATA4 induces the expression of cell cycle-related genes and suppresses those of myogenic and muscle contractile genes. These characteristics of GATA4-dependent pathways are opposite to the downstream of MyoD. To investigate whether GATA4 and MyoD competitively regulates transcription, activities of cyclin D2 promoter and myosin light chain 1 (MLC1) enhancer were examined. Luciferase assays revealed that GATA4 enhances cyclin D2 promoter but this activation is suppressed by MyoD. In contrast, MyoD-induced MLC1 enhancer activity was attenuated by GATA4. These results suggest that GATA4 and MyoD cooperatively switch gene expression program during myoblasts differentiation.
Gata4fl/fl;Pax7CreER mice were generated to knockout GATA4 conditionally in Pax7+ satellite cells by tamoxifen. After fulfillment of tamoxifen treatment, cardiotoxin was injected into leg muscle to induce muscle injury and regeneration. At day 7 after the injection, the diameter of regenerating myofibers was significantly smaller in tamoxifen-treated group. Although there were no difference in fiber diameter at day 14, the number of Pax7+ satellite cells were strikingly decreased at day 28 in tamoxifen group. Thus cardiotoxin was injected again at day 28 then muscles were harvested at day 14 after the second injection. Myofibers of control mice were regenerated as well as after the first injection, however, those of tamoxifen group were markedly small. These findings demonstrate that GATA4 activated in satellite cells is necessary for skeletal muscle regeneration with maintaining satellite cell population.