| Through an RNAi-based screening for kinases involved in myogenic differentiation, we identified Stk24/MST3, a member of the mammalian Ste20 family, as a key regulator. Knockdown of Stk24 by siRNA in both primary and immortalized myoblasts resulted in remarkable suppression of differentiation. Consistently, overexpression of the wild-type or a cleavage-resistant form of Stk24 greatly accelerated myogenic differentiation, while the kinase-dead form of Stk24 barely had any effect. Our data suggested that the kinase activity of Stk24 is required, but the caspase 3-mediated proteolytic cleavage of stk24 is dispensable. Further in-depth studies revealed that the pro-myogenic effect of Stk24 partially lies in its ability to activate NDR1/2, which are required to activate p38 MAPK, a kinase indispensable for myogenic differentiation. To look for additional novel substrates of Stk24, we conducted a yeast two-hybrid screen using a kinase-dead Stk24 as bait. We found that the DEAD box RNA helicase Ddx5/p68, a known modulator of myogenic differentiation could physically interact with Stk24. Moreover, the wild type, but not the kinase dead Stk24 could enhance the binding between Ddx5 and MyoD, augment the stimulating effect of Ddx5 on MyoD, and increase the recruitment of both Ddx5 and MyoD to the MyoG promoter. In vitro kinase assays also confirmed that Stk24 could directly phosphorylate Ddx5. Collectively, our study reveals that Stk24 controls myogenic differentiation through multiple mechanisms. |
