AKT/PKB is a ubiquitous serine-threonine kinase known to mediate the action of a wide range of stimuli thus regulating a plethora of processes in diverse cell types (Manning and Toker, 2017). In particular, AKT plays a central role in the maintenance of mammalian-derived pluripotent stem cells (PSCs) (Kimura and Nakano, 2009, Watanabe et al., 2006). These cells have the capacity to indefinitely self-renew and to potentially differentiate into all the cell types derived from the three germ layers (Evans and Kaufman, 1981, Thomson et al., 1998), and constitute an excellent system for research in developmental biology, disease modeling and drug discovery (Yamanaka, 2020). These capacities are mainly maintained by the pluripotency transcription factors (TFs) NANOG, OCT4 and SOX2, which together constitute the core network of pluripotency that induces the expression of crucial pluripotency genes and inhibits those that promote cell differentiation (Boyer et al., 2005, Loh et al., 2006).

The activation of AKT relies on its recruitment to the plasma membrane, a crucial step for its phosphorylation by PDK1 and mTORC2 (Alessi et al., 1996, Nicholson and Anderson, 2002). Subsequently, AKT activity and target specificity are finely regulated by multiple post-translational modifications (PTMs) (Risso et al., 2015) including the conjugation to the small ubiquitin-related modifier (SUMO) known as SUMOylation (Vidal et al., 2021). This PTM requires a multiple-step activation process of the SUMO peptide that involves different specific enzymes, followed by its ligation to the target lysine residues by the SUMO conjugase enzyme UBC9 (Flotho and Melchior, 2013). Besides, SUMOylation plays an important role in the regulation and function of a large number of proteins, and the deregulation of this pathway has been even associated with disease and cancer (Lara-Ureña et al., 2022). This PTM also affects the localization and nuclear accumulation of other proteins (Knittle et al., 2017, Lara-Ureña et al., 2022), thus impacting on protein-protein interactions (Han et al., 2018). For example, SUMOylation is associated with the nuclear localization of Caspase-8 (Besnault-Mascard et al., 2005), determines the localization at the plasma membrane of the transmembrane adhesion protein Nephrin (Tossidou et al., 2014), and is required for the proper nuclear and subnuclear localization of Dyskerin (MacNeil et al., 2021), among others (Gareau and Lima, 2010).

The SUMOylation of AKT1 affects diverse cellular processes such as cell survival, proliferation and the oncogenic potential of established cell lines (de La Cruz-Herrera et al., 2015, Li et al., 2013, Lin et al., 2016, Risso et al., 2013). We have previously reported that AKT1 SUMOylation is required to induce the expression of the pluripotency TF Nanog in mouse embryonic stem cells (ESCs) (Francia et al., 2021), PSCs derived from the inner cell mass of mouse blastocysts.

In this work, we aimed to deepen into the SUMOylation effects on AKT1 in mouse ESCs. Specifically, we hypothesized that SUMOylation could affect the subcellular distribution of AKT and its affinity for its targets ultimately impacting in its function and specificity.

We show that SUMOylation modulates AKT1 subcellular compartmentalization and distribution, specially within the nuclear space and also affects the chromatin-binding dynamics of Nanog in embryonic stem cells. Interestingly, the E17K mutation of AKT1, found in several types of cancer (Chen et al., 2020), affects these relevant properties of AKT1 probably linked to its oncogenic role.