DAB2IP is an important tumor suppressor in multiple key oncogenic pathways, including TNFα/NF-κB, WNT/β-catenin, PI3K/AKT, and androgen receptors, therefore, inactivated DAB2IP is widely reported to trigger tumor initiating and cancer progressing [13, 15]. As a scaffold protein, DAB2IP could function as a competitor, or scavenger, by binding with multiple signaling regulator and preventing their interaction with other up/downstream effectors, thus potentially modulating a remarkable array of cancer-related pathway [13]. Our previous studies first unveiled the tumor suppression effect and mechanism of DAB2IP in prostate cancer [20–22], and demonstrated DAB2IP could inhibit cell proliferation, epithelial-mesenchymal transition (EMT) and stem cell-like features in human colorectal cancer, especially in p53 wild type subgroup [16, 23]. Additionally, we also reported that DAB2IP could inhibit invasiveness and metastasis in breast cancer through inhibiting invadopodia formation [17]. Currently, reprogrammed glucose metabolism appears to be a hallmark of cancer [24], it could often increase the hypoxic adaptation of tumor cells thus leading a poor cancer survival [25]. Interesting, some studies have pointed out DAB2IP could involve in hypoxia-related regulation in breast and liver cancer [26, 27]. However, the concrete role of DAB2IP in metabolic reprograming of tumor under hypoxia condition is still uncertain.
In this research, we first revealed that DAB2IP could decrease glucose uptake under hypoxia condition, therefore inhibited glycolysis and intracellular ATP production in breast cancer. Considering the critical role of glucose in the energy supplement of cancer cells, intervention on DAB2IP could be a potential strategy to coordinately dampen tumor related metabolic reprogramming, thus limiting cancer progression on a wide spectrum. Our study also demonstrated DAB2IP could downregulate glucose transporter 1 (GLUT1) and its downstream PGK1 through suppressing HIF-1a signaling. Coincidently, Zhou J, et al. reported that loss of DAB2IP could induce HIF-2a expression through activating mTOR pathway in renal cell cancer [28], while Wang B, et al. found DAB2IP could regulate EMT through destabilizing HIF-1a protein in prostate cancer [29]. However, little was known on the detailed biological function of DAB2IP in the HIF1-a related signaling. Our study provided evidences that DAB2IP could induce HIF-1a degradation in an ubiquitylation dependent manner. Moreover, we have identified the ubiquitin E3 ligase, STUB1, which was essential in DAB2IP mediate HIF-1a ubiquitylation. Our results revealed a new possibility of DAB2IP mediate tumor suppression in breast cancer.
Ubiquitylation is a crucial component of post-translational modification and affected the function of target proteins under healthy or pathological events [30]. In HIF-1 signaling, ubiquitylation plays an essential role in determining the half-life and transcriptional activity of HIF-1a, thus controlling the hypoxic response of mammalian cells [31]. Multiple oncogenic pathways, including CDC20/PHD3 [32], SENP1/USP28 [33], and ATF4/pVHL [34] have been demonstrated to dysregulate HIF-1a ubiquitylation and induce tumor progression. In our current research, we found DAB2IP could induce HIF-1a ubiquitylation and degradation through STUB1. STUB1 (also known as CHIP) is an essential E3 ligase involved in protein quality control, and could degrade oncoproteins to exert tumor-suppressive functions [35]. Recently, STUB1 has emerged as an important player in regulating aging, autophagy, cancer immunity [35]. In 2010, Luo W, et al. and Bento CF, et al. observed that STUB1 could selectively mediate ubiquitylated degradation of HIF-1a [36, 37], however the deeply mechanism still needed to be clarified. Our finding showed that DAB2IP could act as a platform to recruit STUB1 and HIF-1a protein, and induced STUB1 mediated HIF-1a degradation through enhancing their interaction. These results partially unveiled the regulation details of STUB1 mediated ubiquitylation. Whether DAB2IP could involve in STUB1-driven ubiquitylated degradation of other oncogenic proteins should be explored in the future.
Another question raised from this research is the paradox role of DAB2IP in regulating protein ubiquitylation. In this study, DAB2IP functioned as an inducer of STUB1-driven HIF-1a ubiquitylation. However, our previous study showed that DAB2IP could act as an inhibitor of wild-type p53 ubiquitylation through antagonizing GRP75 in colorectal cancer [16]. Meanwhile, DAB2IP could induce the deubiquitylation of ALK through USP10 in breast cancer [17]. Similarly, other researches also reported that DAB2IP could induce PARP-1 protein ubiquitylation [38], while in contrast inhibited p27 protein ubiquitylation [39] in renal carcinoma. The biological function of DAB2IP in regulating ubiquitylation and protein stabilization still needed to be evaluated in the future.
As a scaffold protein, DAB2IP consists of several domains, including the pleckstrin homology (PH) domain, PKC-conserved region 2 (C2) domain, Ras-GTPase activating protein (Ras-GAP) domain, C-terminal period-like (PER) domain, proline-rich (PR) domain, and leucine-zipper motif (LZ) [15]. In this research, we identified the PER domain of DAB2IP was essential for STUB1 mediated HIF-1a ubiquitination. PER domain was a non-described region involved in protein-protein interaction [13]. The biological role of PER domain in DAB2IP has still no certainty. Zhang HZ et al. reported that it could interact with TRAF2, thus inhibiting NF-kB activity [40], while Zhang H et al. observed that PER of DAB2IP could displace the binding between 14-3-3 and ASK1 protein, then enhanced ASK1 auto-phosphorylation and pro-apoptotic signals [41]. Furthermore, PER domain could also be important for modulating PI3K-Akt activity, therefore connecting both survival and death signals and maintaining cell homeostasis in prostate cancer cells [20]. This study unveiled that DAB2IP could interact with STUB1 and induced HIF-1a ubiquitylated degradation via the PER-domain. Interesting, we have previously found that DAB2IP could bind to GRP75 and enhance wild-type p53 stability via its Ras-GAP domain [16]. The concrete functions of different domain in DAB2IP-mediated ubiquitylation regulation need further investigation.
There are still some limitations in this study. First, we found loss of DAB2IP could significantly induce glucose uptake, then subsequently increase lactate and intracellular ATP production in breast cancer cells under hypoxic condition. However, the influence of DAB2IP expression on glucose metabolism under normoxia was not obvious. The reason might be that DAB2IP regulate glucose metabolism was dependent on HIF-1a signal, which was significantly induced by hypoxia. Other potential pathways involved in metabolic reprogramming should be validated in the further research. Second, although we found DAB2IP could inhibit GLUT1 and PGK1 expression thereby affect ECAR in breast cancer cells, however, other key enzymes involved in glucose metabolism, such as hexokinase (HK), glucose-6-phosphate isomerase (GPI), phosphofructokinase (PEK), pyruvate dehydrogenase (PDH) and lactate dehydrogenase A/B (LDHA/B) [42], were not well investigated. Third, tumor cells were found to derived the energy required for uncontrolled replication through rapidly consuming glucose and converting it into lactate (Warburg effect) [43]. Although loss of DAB2IP could induce lactate and ATP production, whether DAB2IP could subsequently regulate glycolysis or Warburg effect in breast cancer cells is still uncertain. Finally, PER domain has been identified as the key region and potential target for DAB2IP-mediated glucose metabolism, therefore, additional studies are required to synthesized short peptides or small molecular compounds targetting this motif, and to prove its therapeutic value.
In conclusion, our research exhibited a novel function of DAB2IP on STUB1-driven HIF-1a ubiquitylated degradation and glucose metabolic reprogramming in breast cancer. These findings enriched our understanding of the crosstalk between DAB2IP and hypoxia signaling pathway, and provided new potential target for cancer therapy.