Abstract
Although ASXL1 mutations are frequently found in human diseases, including myeloid leukemia, the cell proliferation–associated function of ASXL1 is largely unknown. Here, we explored the molecular mechanism underlying the growth defect found in Asxl1-deficient mouse embryonic fibroblasts (MEFs). We found that Asxl1, through amino acids 371 to 655, interacts with the kinase domain of AKT1. In Asxl1-null MEFs, IGF-1 was unable to induce AKT1 phosphorylation and activation; p27Kip1, which forms a ternary complex with ASXL1 and AKT1, therefore remained unphosphorylated. Hypophosphorylated p27Kip1 is able to enter the nucleus, where it prevents the phosphorylation of Rb; this ultimately leads to the down-regulation of E2F target genes as confirmed by microarray analysis. We also found that senescence-associated (SA) genes were upregulated and that SA β-gal staining was increased in Asxl1−/− MEFs. Further, the treatment of an AKT inhibitor not only stimulated nuclear accumulation of p27Kip1 leading to E2F inactivation, but also promoted senescence. Finally, Asxl1 disruption augmented the expression of p16Ink4a as result of the defect in Asxl1-Ezh2 cooperation. Overall, our study provides the first evidence that Asxl1 both activates the AKT-E2F pathway and cooperates with Ezh2 through direct interactions at early embryonic stages, reflecting that Asxl1 disruption causes cellular senescence.
Highlights
The additional sex comb (Asx) gene was originally identified in Drosophila as an enhancer of trithorax group (TrxG) and Polycomb group (PcG) proteins[1]
The higher serum concentration significantly increased the proliferation of both WT and heterozygote mouse embryonic fibroblasts (MEFs), but no serum effect was observed in Asxl1−/− MEFs (Fig. 1d)
Further FACS analysis indicated that the G0/G1 population was significantly higher in Asxl1-null MEFs (67.11%) than in WT (48.94%) and heterozygote (51.33%) MEFs (Supplementary Figure 1)
Summary
The additional sex comb (Asx) gene was originally identified in Drosophila as an enhancer of trithorax group (TrxG) and Polycomb group (PcG) proteins[1]. Proliferation and survival[15], but it plays a role in the deregulation of cell cycle control by phosphorylating various target proteins[16]. Specific regulation of the Rb-E2F-p27Kip1-AKT network could be critical for the control of cell proliferation and senescence. The expression of p16Ink4a increased due to defective cooperation with Ezh[2] in Asxl1-null MEFs. In addition, the expression of p16Ink4a increased due to defective cooperation with Ezh[2] in Asxl1-null MEFs Overall, these data suggest that Asxl[1] plays a critical role in the proliferation of embryonic cells by cooperating with both the AKT-E2F axis and Ezh[2], the disruption of which leads to senescence
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