Ai-ng Li

Cdk1-phosphorylated CUEDC2 promotes spindle checkpoint inactivation and chromosomal instability

Aneuploidy and chromosomal instability are major characteristics of human cancer. These abnormalities can result from defects in the spindle assembly checkpoint (SAC), which is a surveillance mechanism for accurate chromosome segregation through restraint of the activity of the anaphase-promoting complex/cyclosome (APC/C). Here, we show that a CUE-domain-containing protein, CUEDC2, is a cell-cycle regulator that promotes spindle checkpoint inactivation and releases APC/C from checkpoint inhibition. CUEDC2 is phosphorylated by Cdk1 during mitosis. Depletion of CUEDC2 causes a checkpoint-dependent delay of the metaphase–anaphase transition. Phosphorylated CUEDC2 binds to Cdc20, an activator of APC/C, and promotes the release of Mad2 from APC/C–Cdc20 and subsequent APC/C activation. CUEDC2 overexpression causes earlier activation of APC/C, leading to chromosome missegregation and aneuploidy. Interestingly, CUEDC2 is highly expressed in many types of tumours. These results suggest that CUEDC2 is a key regulator of mitosis progression, and that CUEDC2 dysregulation might contribute to tumour development by causing chromosomal instability.

NEDD4 ubiquitinates TRAF3 to promote CD40-mediated AKT activation

CD40, a member of tumour necrosis factor receptor (TNFR) superfamily, has a pivotal role in B-cell-mediated immunity through various effector pathways including AKT kinase, but the signal transduction of CD40-meidated AKT activation is poorly understood. Here we report that the neural precursor cell expressed developmentally downregulated protein 4 (NEDD4), homologous to E6-AP Carboxyl Terminus family E3 ubiquitin ligase, is a novel component of the CD40 signalling complex. It has a key role in CD40-mediated AKT activation and is involved in modulating immunoglobulin class switch through regulating the expression of activation-induced cytidine deaminase. NEDD4 constitutively interacts with CD40 and mediates K63-linked ubiquitination of TNFR-associated factor3 (TRAF3). The ubiquitination of TRAF3 by NEDD4 is critical for CD40-mediated AKT activation. Thus, NEDD4 is a previously unknown component of the CD40 signalling complex necessary for AKT activation.

CUEDC2 (CUE Domain-containing 2) and SOCS3 (Suppressors of Cytokine Signaling 3) Cooperate to Negatively Regulate Janus Kinase 1/Signal Transducers and Activators of Transcription 3 Signaling

Background: Deactivation of the JAK1/STAT3 pathway is tightly controlled in cells. Results: CUEDC2 inhibits JAK1/STAT3 signaling through binding to SOCS3. Conclusion: CUEDC2 is a novel regulator of JAK1/STAT3 signaling. Significance: Our study identified a novel potential mechanism of SOCS3-mediated suppression on the JAK/STAT signaling pathway and provided important insight into the critical roles of CUEDC2 in the complex signal transduction network. Janus kinase 1/signal transducers and activators of transcription 3 (JAK1/STAT3) pathway is one of the recognized oncogenic signaling pathways that frequently overactivated in a variety of human tumors. Despite rapid progress in elucidating the molecular mechanisms of activation of JAK/STAT pathway, the processes that regulate JAK/STAT deactivation need to be further clarified. Here we demonstrate that CUE domain-containing 2 (CUEDC2) inhibits cytokine-induced phosphorylation of JAK1 and STAT3 and the subsequent STAT3 transcriptional activity. Further analysis by a yeast two-hybrid assay showed that CUEDC2 could engage in a specific interaction with a key JAK/STAT inhibitor, SOCS3 (suppressors of cytokine signaling 3). The interaction between CUEDC2 and SOCS3 is required for the inhibitory effect of CUEDC2 on JAK1 and STAT3 activity. Additionally, we found CUEDC2 functions collaboratively with SOCS3 to inhibit JAK1/STAT3 signaling by increasing SOCS3 stability via enhancing its association with Elongin C. Therefore, our findings revealed a new biological activity for CUEDC2 as the regulator of JAK1/STAT3 signaling and paved the way to a better understanding of the mechanisms by which SOCS3 has been linked to suppression of the JAK/STAT pathway.

Serum Peptidome Variations in a Healthy Population: Reference to Identify Cancer-Specific Peptides

The emergence of mass spectrometry (MS)-based signatures as biomarkers has generated considerable enthusiasm among oncologists. However, variations in normal individuals also exist, and a better understanding of serum peptide patterns of healthy individuals will be important for further exploring disease-specific serum peptide patterns. Following development of a serum peptide pattern platform, we analyzed 500 serum samples obtained from healthy individuals. Samples from breast (n = 84), lung (n = 70), and rectal (n = 30) cancer patients were also examined. Extensive data analysis revealed negligible contributions of age to serum peptide patterns except in healthy individuals between 20–30 and 60+ years of age. Gender-related variations in the serum patterns of healthy individuals were only observed in 20–30 year-old individuals. Our results revealed substantial variation in individual peptide profiles, but 65 peptides were detected at a 20% higher frequency in the healthy population. A peptide profile was developed for each type of cancer, containing 10 discriminating peptides not prevalent in healthy individuals. Sequence identification of 111 signature peptides revealed that they fell into several tight clusters and most were exopeptidase products of serum resident proteins. We have obtained a MS-based serum peptide profile for healthy individuals, providing a reference for observing the occurrence of cancer-specific peptides.

ECHS1 interacts with STAT3 and negatively regulates STAT3 signaling

STAT3 physically interacts with ECHS1 by anti bait co immunoprecipitation (View interaction)

Phosphorylation-triggered CUEDC2 degradation promotes UV-induced G1 arrest through APC/CCdh1 regulation

DNA damage triggers cell cycle arrest to provide a time window for DNA repair. Failure of arrest could lead to genomic instability and tumorigenesis. DNA damage-induced G1 arrest is generally achieved by the accumulation of Cyclin-dependent kinase inhibitor 1 (p21). However, p21 is degraded and does not play a role in UV-induced G1 arrest. The mechanism of UV-induced G1 arrest thus remains elusive. Here, we have identified a critical role for CUE domain-containing protein 2 (CUEDC2) in this process. CUEDC2 binds to and inhibits anaphase-promoting complex/cyclosome-Cdh1 (APC/CCdh1), a critical ubiquitin ligase in G1 phase, thereby stabilizing Cyclin A and promoting G1–S transition. In response to UV irradiation, CUEDC2 undergoes ERK1/2-dependent phosphorylation and ubiquitin-dependent degradation, leading to APC/CCdh1-mediated Cyclin A destruction, Cyclin-dependent kinase 2 inactivation, and G1 arrest. A nonphosphorylatable CUEDC2 mutant is resistant to UV-induced degradation. Expression of this stable mutant effectively overrides UV-induced G1–S block. These results establish CUEDC2 as an APC/CCdh1 inhibitor and indicate that regulated CUEDC2 degradation is critical for UV-induced G1 arrest.

Ubiquitin-associated domain-containing ubiquitin regulatory X (UBX) protein UBXN1 is a negative regulator of nuclear factor κB (NF-κB) signaling.

Background: Excessive NF-κB hyperactivation should be tightly controlled in cells. Results: UBXN1 inhibits TNFα-triggered NF-κB signaling by sequestering cIAPs from being recruited to TNFR1. Conclusion: UBXN1 is a novel negative regulator of TNFα-triggered NF-κB signaling. Significance: Our study identified a novel ubiquitin-linked protein UBXN1 as a negative regulator of NF-κB signaling pathway independent of VCP/p97 and provided important insight into the new regulatory mechanism of the UBX protein family. Excessive nuclear factor κB (NF-κB) activation should be precisely controlled as it contributes to multiple immune and inflammatory diseases. However, the negative regulatory mechanisms of NF-κB activation still need to be elucidated. Various types of polyubiquitin chains have proved to be involved in the process of NF-κB activation. Many negative regulators linked to ubiquitination, such as A20 and CYLD, inhibit IκB kinase activation in the NF-κB signaling pathway. To find new NF-κB signaling regulators linked to ubiquitination, we used a small scale siRNA library against 51 ubiquitin-associated domain-containing proteins and screened out UBXN1, which contained both ubiquitin-associated and ubiquitin regulatory X (UBX) domains as a negative regulator of TNFα-triggered NF-κB activation. Overexpression of UBXN1 inhibited TNFα-triggered NF-κB activation, although knockdown of UBXN1 had the opposite effect. UBX domain-containing proteins usually act as valosin-containing protein (VCP)/p97 cofactors. However, knockdown of VCP/p97 barely affected UBXN1-mediated NF-κB inhibition. At the same time, we found that UBXN1 interacted with cellular inhibitors of apoptosis proteins (cIAPs), E3 ubiquitin ligases of RIP1 in the TNFα receptor complex. UBXN1 competitively bound to cIAP1, blocked cIAP1 recruitment to TNFR1, and sequentially inhibited RIP1 polyubiquitination in response to TNFα. Therefore, our findings demonstrate that UBXN1 is an important negative regulator of the TNFα-triggered NF-κB signaling pathway by mediating cIAP recruitment independent of VCP/p97.

Polo-like kinase 1 coordinates biosynthesis during cell cycle progression by directly activating pentose phosphate pathway

Two hallmarks for cancer cells are the accelerated cell cycle progression as well as the altered metabolism, however, how these changes are coordinated to optimize the growth advantage for cancer cells are still poorly understood. Here we identify that Polo-like kinase 1 (Plk1), a key regulator for cell mitosis, plays a critical role for biosynthesis in cancer cells through activating pentose phosphate pathway (PPP). We find that Plk1 interacts with and directly phosphorylates glucose-6-phosphate dehydrogenase (G6PD). By activating G6PD through promoting the formation of its active dimer, Plk1 increases PPP flux and directs glucose to the synthesis of macromolecules. Importantly, we further demonstrate that Plk1-mediated activation of G6PD is critical for its role to promote cell cycle progression and cancer cell growth. Collectively, these findings establish a critical role for Plk1 in regulating biosynthesis in cancer cells, exemplifying how cell cycle progression and metabolic reprogramming are coordinated for cancer progression.Polo-like kinase 1 (Plk1) is a key regulator of mitosis. Here, the authors show that Plk1 activates the pentose phosphate pathway in cancer cells by directly phosphorylating glucose-6-phosphate dehydrogenase (G6PD) and that such activation is critical for cell cycle progression and cancer cell growth.

RNF4 negatively regulates NF‐κB signaling by down‐regulating TAB2

Most of NF‐κB (nuclear factor kappa B) signaling molecules have various types of post‐translational modifications. In this study, we focused on ubiquitination and designed a siRNA library including most ubiquitin‐binding domains. With this library, we identified several candidate regulators of canonical NF‐κB pathway, including RNF4. Overexpression of RNF4 impaired NF‐κB activation in a dose‐dependent manner, whereas RNF4 knockdown potentiated NF‐κB activation. We showed that RNF4 interacts with the TAK1–TAB2–TAB3 complex, but not TAB1. Further, we found that RNF4 specifically down‐regulated TAB2 through a lysosomal pathway, and knockdown of RNF4 impaired endogenous TAB2 degradation. Therefore, our findings will provide new insights into the negative regulation of NF‐κB signaling.

CUEDC2 modulates cardiomyocyte oxidative capacity by regulating GPX1 stability

The irreversible loss of cardiomyocytes due to oxidative stress is the main cause of heart dysfunction following ischemia/reperfusion (I/R) injury and ageing‐induced cardiomyopathy. Here, we report that CUEDC2, a CUE domain‐containing protein, plays a critical role in oxidative stress‐induced cardiac injury. Cuedc2−/− cardiomyocytes exhibited a greater resistance to oxidative stress‐induced cell death. Loss of CUEDC2 enhanced the antioxidant capacity of cardiomyocytes, promoted reactive oxygen species (ROS) scavenging, and subsequently inhibited the redox‐dependent activation of signaling pathways. Notably, CUEDC2 promoted E3 ubiquitin ligases tripartite motif‐containing 33 (TRIM33)‐mediated the antioxidant enzyme, glutathione peroxidase 1 (GPX1) ubiquitination, and proteasome‐dependent degradation. Ablation of CUEDC2 upregulated the protein level of GPX1 in the heart significantly. Strikingly, in vivo, the infarct size of Cuedc2−/− heart was significantly decreased after I/R injury, and aged Cuedc2−/− mice preserved better heart function as the overall ROS levels in their hearts were significantly lower. Our results demonstrated a novel role of CUEDC2 in cardiomyocyte death regulation. Manipulating CUEDC2 level might be an attractive therapeutic strategy for promoting cardiomyocyte survival following oxidative stress‐induced cardiac injury.