Development of an Anti-canine PD-L1 Antibody and Caninized PD-L1 Mouse Model as Translational Research Tools for the Study of Immunotherapy in Humans.

Unlabelled: Immune checkpoint blockade therapy, one of the most promising cancer immunotherapies, has shown remarkable clinical impact in multiple cancer types. Despite the recent success of immune checkpoint blockade therapy, however, the response rates in patients with cancer are limited (∼20%-40%). To improve the success of immune checkpoint blockade therapy, relevant preclinical animal models are essential for the development and testing of multiple combination approaches and strategies.

CtIP Regulates Mitotic Spindle Assembly by Modulating the TPX2-Aurora A Signaling Axis.

CtBP-interacting protein (CtIP) plays a critical role in controlling the homologous recombination-mediated DNA double-stranded break (DSB) repair pathway through DNA end resection, and recent studies suggest that it also plays a role in mitosis. However, the mechanism by which CtIP contributes to mitosis regulation remains elusive. Here, we show that depletion of CtIP leads to a delay in anaphase progression resulting in misaligned chromosomes, an aberrant number of centrosomes, and defects in chromosome segregation.

Cdk1 phosphorylation negatively regulates the activity of Net1 towards RhoA during mitosis.

The Neuroepithelial transforming gene 1 (Net1) is a RhoA subfamily guanine nucleotide exchange factor that is overexpressed in a number of cancers and contributes to cancer cell motility and proliferation. Net1 also plays a Rho GTPase independent role in mitotic progression, where it promotes centrosomal activation of Aurora A and Pak2, and aids in chromosome alignment during prometaphase. To understand regulatory mechanisms controlling the mitotic function of Net1, we examined whether it was phosphorylated by the mitotic kinase Cdk1.

Saccharide analog, 2-deoxy-d-glucose enhances 4-1BB-mediated antitumor immunity via PD-L1 deglycosylation.

Triple-negative breast cancer (TNBC) lacks a well-defined molecular target and is associated with poorer outcomes compared to other breast cancer subtypes. Programmed cell death protein 1 (PD-1)/programmed death-ligand 1 (PD-L1) blockade therapy shows a 10% to 20% response rate in TNBC patients. Our previous studies show that PD-L1 proteins are heavily glycosylated in TNBC, and the glycosylation plays an important role in the PD-L1 protein's stability and immunosuppressive function.

Stress-activated MAPKs and CRM1 regulate the subcellular localization of Net1A to control cell motility and invasion.

The neuroepithelial cell transforming gene 1A (Net1A, an isoform of Net1) is a RhoA subfamily guanine nucleotide exchange factor (GEF) that localizes to the nucleus in the absence of stimulation, preventing it from activating RhoA. Once relocalized in the cytosol, Net1A stimulates cell motility and extracellular matrix invasion. In the present work, we investigated mechanisms responsible for the cytosolic relocalization of Net1A.

Phosphorylation of p53 at threonine 155 is required for Jab1-mediated nuclear export of p53.

The Jun activation-domain binding protein 1 (Jab1) induces p53 nuclear export and cytoplasmic degradation, but the underlying mechanism is poorly understood. Here, we show that phosphorylation at the threonine 155 residue is essential for Jab1-mediated p53 nuclear export. Jab1 stimulated phosphorylation of p53 at T155 was inhibited by curcumin, an inhibitor of COP9 signalosome (CSN)-associated kinases.

An essential role for UTX in resolution and activation of bivalent promoters.

Trimethylated histone H3 lysine 27 (H3K27me3) is linked to gene silencing, whereas H3K4me3 is associated with gene activation. These two marks frequently co-occupy gene promoters, forming bivalent domains. Bivalency signifies repressed but activatable states of gene expression and can be resolved to active, H3K4me3-prevalent states during multiple cellular processes, including differentiation, development and epithelial mesenchymal transition.

Minireview: Mouse Models of Rho GTPase Function in Mammary Gland Development, Tumorigenesis, and Metastasis.

Ras homolog (Rho) family small GTPases are critical regulators of actin cytoskeletal organization, cell motility, proliferation, and survival. Surprisingly, the large majority of the studies underlying our knowledge of Rho protein function have been carried out in cultured cells, and it is only recently that researchers have begun to assess Rho GTPase regulation and function in vivo. The purpose of this review is to evaluate our current knowledge of Rho GTPase function in mouse mammary gland development, tumorigenesis and metastasis.

Acetylation of the RhoA GEF Net1A controls its subcellular localization and activity.

Net1 isoform A (Net1A) is a RhoA GEF that is required for cell motility and invasion in multiple cancers. Nuclear localization of Net1A negatively regulates its activity, and we have recently shown that Rac1 stimulates Net1A relocalization to the plasma membrane to promote RhoA activation and cytoskeletal reorganization. However, mechanisms controlling the subcellular localization of Net1A are not well understood.

Rho GTPase independent regulation of ATM activation and cell survival by the RhoGEF Net1A.

ATM activation following DNA damage is a critical event which is required for efficient DNA repair and cell survival, yet signaling mechanisms controlling its activation are incompletely understood. The RhoGEF Net1 has previously been reported to control Rho GTPase activation and downstream cell survival outcomes following double strand DNA damage. However the role of Net1 isoforms in controlling ATM-dependent cell signaling has not been assessed.

Controlling the switches: Rho GTPase regulation during animal cell mitosis.

Animal cell division is a fundamental process that requires complex changes in cytoskeletal organization and function. Aberrant cell division often has disastrous consequences for the cell and can lead to cell senescence, neoplastic transformation or death. As important regulators of the actin cytoskeleton, Rho GTPases play major roles in regulating many aspects of mitosis and cytokinesis.

Rho GTPase-independent regulation of mitotic progression by the RhoGEF Net1.

Neuroepithelial transforming gene 1 (Net1) is a RhoA-subfamily-specific guanine nucleotide exchange factor that is overexpressed in multiple human cancers and is required for proliferation. Molecular mechanisms underlying its role in cell proliferation are unknown. Here we show that overexpression or knockdown of Net1 causes mitotic defects.

Regulation of focal adhesion kinase activation, breast cancer cell motility, and amoeboid invasion by the RhoA guanine nucleotide exchange factor Net1.

Net1 is a RhoA guanine nucleotide exchange factor (GEF) that is overexpressed in a subset of human cancers and contributes to cancer cell motility and invasion in vitro. However, the molecular mechanism accounting for its role in cell motility and invasion has not been described. In the present work, we show that expression of both Net1 isoforms in breast cancer cells is required for efficient cell motility.

MKRN1 induces degradation of West Nile virus capsid protein by functioning as an E3 ligase.

West Nile virus capsid protein (WNVCp) displays pathogenic toxicity via the apoptotic pathway. However, a cellular mechanism protective against this toxic effect has not been observed so far. Here, we identified Makorin ring finger protein 1 (MKRN1) as a novel E3 ubiquitin ligase for WNVCp.

PML-IV functions as a negative regulator of telomerase by interacting with TERT.

Maintaining proper telomere length requires the presence of the telomerase enzyme. Here we show that telomerase reverse transcriptase (TERT), a catalytic component of telomerase, is recruited to promyelocytic leukemia (PML) nuclear bodies through its interaction with PML-IV. Treatment of interferon-alpha (IFNalpha) in H1299 cells resulted in the increase of PML proteins with a concurrent decrease of telomerase activity, as previously reported.

Differential regulation of p53 and p21 by MKRN1 E3 ligase controls cell cycle arrest and apoptosis.

Makorin Ring Finger Protein 1 (MKRN1) is a transcriptional co-regulator and an E3 ligase. Here, we show that MKRN1 simultaneously functions as a differentially negative regulator of p53 and p21. In normal conditions, MKRN1 could destabilize both p53 and p21 through ubiquitination and proteasome-dependent degradation.

West Nile virus capsid protein induces p53-mediated apoptosis via the sequestration of HDM2 to the nucleolus.

The capsid protein of the West Nile virus (WNV) functions as an apoptotic agonist via the induction of mitochondrial dysfunction and the activation of caspases-9 and -3. Here, we have determined that the WNV capsid (WNVCp) is capable of binding to and sequestering HDM2 into the nucleolus. WNVCp was shown to interfere with the formation of the HDM2 and p53 complex, thereby causing the stabilization of p53 and the subsequent induction of its target apoptotic protein, Bax.

Jab1 as a mediator of nuclear export and cytoplasmic degradation of p53.

Jun activation domain-binding protein 1 (Jab1) is involved in various cellular mechanisms including development in Drosophila and mouse, cell cycle control and signal transduction pathways. Recent studies also determined that Jab1 functions as a nuclear exporter and inducer of cytoplasmic degradation for several proteins including p53, p27, capsid of West Nile virus, and Smad4/7 proteins. In particular, p53 is shown to bind to and to be exported into the cytoplasm by Jab1, which helps to maintain low levels of p53 under normal conditions.

Jab1 mediates cytoplasmic localization and degradation of West Nile virus capsid protein.

The clinical manifestations of West Nile virus (WNV), a member of the Flavivirus family, include febrile illness, sporadic encephalitis, and paralysis. The capsid (Cp) of WNV is thought to participate in these processes by inducing apoptosis through mitochondrial dysfunction and activation of caspase-9 and caspase-3. To further identify the molecular mechanism of the WNV capsid protein (WNVCp), yeast two-hybrid assays were employed using WNV-Cp as bait.

Hsp70 functions as a negative regulator of West Nile virus capsid protein through direct interaction.

West Nile virus (WNV) is a member of the Flavivirus family and induces febrile illness, sporadic encephalitis, and paralysis. The capsid (Cp) of WNV is thought to play a role in inducing these symptoms through caspase-3- and caspase-9-dependent apoptosis. Using WNVCp as bait for a yeast two-hybrid assay, we identified that Hsp70 interacted with WNVCp.

Jab1 induces the cytoplasmic localization and degradation of p53 in coordination with Hdm2.

The biological mechanisms for maintaining the basal level of p53 in normal cells require nuclear exclusion and cytoplasmic degradation. Here, we showed that Jab1 facilitates p53 nuclear exclusion and its subsequent degradation in coordination with Hdm2. p53 was excluded from the nucleus in the presence of Jab1; this exclusion was prevented by leptomycin B treatment.

Cooperative interaction of Hsp40 and TPR1 with Hsp70 reverses Hsp70-HspBp1 complex formation.

Hsp40 and TPR1 are chaperone adaptors that regulate Hsp70-dependent folding processes by interacting with the amino terminal and carboxy terminal domains of Hsp70, respectively. In this study, we report cooperative interactions involving Hsp70, Hsp40, and TPR1 that enhance Hsp70-dependent folding of chemically denatured substrates. Hsp40 and Hsp70 dependent folding of chemically denatured luciferase was enhanced by up to 80% when TPR1 was also present.