Xue-Song Liu

Rig-I−/− mice develop colitis associated with downregulation of Gαi2

RIG-I (retinoid acid-inducible gene-I), a putative RNA helicase with a cytoplasmic caspase-recruitment domain (CARD), was identified as a pattern-recognition receptor (PRR) that mediates antiviral immunity by inducing type I interferon production. To further study the biological function of RIG-I, we generated Rig-I−/− mice through homologous recombination, taking a different strategy to the previously reported strategy. Our Rig-I−/− mice are viable and fertile. Histological analysis shows that Rig-I−/− mice develop a colitis-like phenotype and increased susceptibility to dextran sulfate sodium-induced colitis. Accordingly, the size and number of Peyers patches dramatically decreased in mutant mice. The peripheral T-cell subsets in mutant mice are characterized by an increase in effector T cells and a decrease in naïve T cells, indicating an important role for Rig-I in the regulation of T-cell activation. It was further found that Rig-I deficiency leads to the downregulation of G protein αi2 subunit (Gαi2) in various tissues, including T and B lymphocytes. By contrast, upregulation of Rig-I in NB4 cells that are treated with ATRA is accompanied by elevated Gαi2 expression. Moreover, Gαi2 promoter activity is increased in co-transfected NIH3T3 cells in a Rig-I dose-dependent manner. All these findings suggest that Rig-I has crucial roles in the regulation of Gαi2 expression and T-cell activation. The development of colitis may be, at least in part, associated with downregulation of Gαi2 and disturbed T-cell homeostasis.

Disruption of palladin results in neural tube closure defects in mice.

Palladin is a newly identified actin-associated protein which was proposed to be involved in actin cytoskeleton organization and nervous system development. Here, we show that inactivation of palladin leads to embryonic lethality due to severe defects of cranial neural tube closure and herniation of liver and intestine. It was found that palladin(-/-) embryos died around E15.5 and developed cranial neural tube closure defects (NTDs) with 100% penetrance. Whole mount in situ hybridization revealed that expression of palladin in early wild type embryos (E8.5) was specifically restricted in the elevating cranial neural folds where the neural tube closure is initiated. Palladin expression closely mirrors the phenotypic defects observed in palladin(-/-) mutants. While in E 9.5 and E10.5 embryos palladin was ubiquitously expressed. In vitro study revealed that formation of stress fibers in cytoplasm, cell adherent ability to extra-cellular matrix protein fibronectin and cell migration were dramatically disturbed in palladin(-/-) murine embryonic fibroblast cells (MEFs). Our findings suggest that palladin plays important roles in actin stress fiber formation, cell adhesion and migration. We propose that palladin is required for the initiation of neural tube closure and provides an important new candidate that may be implicated in the etiology of human NTDs.

Rig-I-/- mice develop colitis associated with downregulation of G alpha i2.

RIG-I (retinoid acid-inducible gene-I), a putative RNA helicase with a cytoplasmic caspase-recruitment domain (CARD), was identified as a pattern-recognition receptor (PRR) that mediates antiviral immunity by inducing type I interferon production. To further study the biological function of RIG-I, we generated Rig-I−/− mice through homologous recombination, taking a different strategy to the previously reported strategy. Our Rig-I−/− mice are viable and fertile. Histological analysis shows that Rig-I−/− mice develop a colitis-like phenotype and increased susceptibility to dextran sulfate sodium-induced colitis. Accordingly, the size and number of Peyers patches dramatically decreased in mutant mice. The peripheral T-cell subsets in mutant mice are characterized by an increase in effector T cells and a decrease in naïve T cells, indicating an important role for Rig-I in the regulation of T-cell activation. It was further found that Rig-I deficiency leads to the downregulation of G protein αi2 subunit (Gαi2) in various tissues, including T and B lymphocytes. By contrast, upregulation of Rig-I in NB4 cells that are treated with ATRA is accompanied by elevated Gαi2 expression. Moreover, Gαi2 promoter activity is increased in co-transfected NIH3T3 cells in a Rig-I dose-dependent manner. All these findings suggest that Rig-I has crucial roles in the regulation of Gαi2 expression and T-cell activation. The development of colitis may be, at least in part, associated with downregulation of Gαi2 and disturbed T-cell homeostasis.

Zbtb7a suppresses prostate cancer through repression of a Sox9-dependent pathway for cellular senescence bypass and tumor invasion

Zbtb7a has previously been described as a powerful proto-oncogene. Here we unexpectedly demonstrate that Zbtb7a has a critical oncosuppressive role in the prostate. Prostate-specific inactivation of Zbtb7a leads to a marked acceleration of Pten loss–driven prostate tumorigenesis through bypass of Pten loss–induced cellular senescence (PICS). We show that ZBTB7A physically interacts with SOX9 and functionally antagonizes its transcriptional activity on key target genes such as MIA, which is involved in tumor cell invasion, and H19, a long noncoding RNA precursor for an RB-targeting microRNA. Inactivation of Zbtb7a in vivo leads to Rb downregulation, PICS bypass and invasive prostate cancer. Notably, we found that ZBTB7A is genetically lost, as well as downregulated at both the mRNA and protein levels, in a subset of human advanced prostate cancers. Thus, we identify ZBTB7A as a context-dependent cancer gene that can act as an oncogene in some contexts but also has oncosuppressive-like activity in PTEN-null tumors.

Multiple synostoses syndrome is due to a missense mutation in exon 2 of FGF9 gene.

Fibroblast growth factors (FGFs) play diverse roles in several developmental processes. Mutations leading to deregulated FGF signaling can cause human skeletal dysplasias and cancer.(1,2) Here we report a missense mutation (Ser99Asp) in exon 2 of FGF9 in 12 patients with multiple synostoses syndrome (SYNS) in a large Chinese family. In vitro studies demonstrate that FGF9(S99N) is expressed and secreted as efficiently as wild-type FGF9 in transfected cells. However, FGF9(S99N) induces compromised chondrocyte proliferation and differentiation, which is accompanied by enhanced osteogenic differentiation and matrix mineralization of bone marrow-derived mesenchymal stem cells (BMSCs). Biochemical analysis reveals that S99N mutation in FGF9 leads to significantly impaired FGF signaling, as evidenced by diminished activity of Erk1/2 pathway and decreased beta-catenin and c-Myc expression when compared with wild-type FGF9. Importantly, the binding of FGF9(S99N) to its receptor is severely impaired although the dimerization ability of mutant FGF9 itself or with wild-type FGF9 is not detectably affected, providing a basis for the defective FGFR signaling. Collectively, our data demonstrate a previously uncharacterized mutation in FGF9 as one of the causes of SYNS, implicating an important role of FGF9 in normal joint development.

Palladin regulates cell and extracellular matrix interaction through maintaining normal actin cytoskeleton architecture and stabilizing Beta1-integrin

Cell and extracellular matrix (ECM) interaction plays an important role in development and normal cellular function. Cell adhesion and cell spreading on ECM are two basic cellular behaviors related to cell‐ECM interaction. Here we show that palladin, a novel actin cytoskeleton‐associated protein, is actively involved in the regulation of cell‐ECM interaction. It was found that palladin‐deficient mouse embryonic fibroblasts (MEFs) display decreased cell adhesion and compromised cell spreading on various ECMs. Disorganized actin cytoskeleton architecture characterized by faint stress fibers, less lamellipodia and focal adhesions can account for the weakened cell‐ECM interaction in palladin−/− MEFs. Furthermore, decreased polymerized filament actin and increased globular actin can be observed in palladin−/− MEFs, strongly suggesting that palladin is essential for the formation or stabilization of polymerized filament actin. Elevated phospho‐cofilin level and proper responses in cofilin phosphorylation to either Rho signal agonist or antagonist in palladin−/− MEFs indicate that disrupted stress fibers in palladin−/− MEFs is not associated with cofilin phosphorylation. More interestingly, the protein level of ECM receptor β1‐integrin is dramatically decreased in MEFs lacking palladin. Down‐regulation of β1‐integrin protein can be restored by proteasome inhibitor MG‐132 treatment. All these data implicate that palladin is essential for cell‐ECM interaction through maintaining normal actin cytoskeleton architecture and stabilizing β1‐integrin protein. J. Cell. Biochem. 100: 1288–1300, 2007.

Vulnerabilities of PTEN–TP53-Deficient Prostate Cancers to Compound PARP–PI3K Inhibition

UNLABELLED Prostate cancer is the most prevalent cancer in males, and treatment options are limited for advanced forms of the disease. Loss of the PTEN and TP53 tumor suppressor genes is commonly observed in prostate cancer, whereas their compound loss is often observed in advanced prostate cancer. Here, we show that PARP inhibition triggers a p53-dependent cellular senescence in a PTEN-deficient setting in the prostate. Surprisingly, we also find that PARP-induced cellular senescence is morphed into an apoptotic response upon compound loss of PTEN and p53. We further show that superactivation of the prosurvival PI3K-AKT signaling pathway limits the efficacy of a PARP single-agent treatment, and that PARP and PI3K inhibitors effectively synergize to suppress tumorigenesis in human prostate cancer cell lines and in a Pten/Trp53-deficient mouse model of advanced prostate cancer. Our findings, therefore, identify a combinatorial treatment with PARP and PI3K inhibitors as an effective option for PTEN-deficient prostate cancer. SIGNIFICANCE The paucity of therapeutic options in advanced prostate cancer displays an urgent need for the preclinical assessment of novel therapeutic strategies. We identified differential therapeutic vulnerabilities that emerge upon the loss of both PTEN and p53, and observed that combined inhibition of PARP and PI3K provides increased efficacy in hormone-insensitive advanced prostate cancer.

ZBTB7A acts as a tumor suppressor through the transcriptional repression of glycolysis

Elevated glycolysis is a common metabolic trait of cancer, but what drives such metabolic reprogramming remains incompletely clear. We report here a novel transcriptional repressor-mediated negative regulation of glycolysis. ZBTB7A, a member of the POK (POZ/BTB and Krüppel) transcription repressor family, directly binds to the promoter and represses the transcription of critical glycolytic genes, including GLUT3, PFKP, and PKM. Analysis of The Cancer Genome Atlas (TCGA) data sets reveals that the ZBTB7A locus is frequently deleted in many human tumors. Significantly, reduced ZBTB7A expression correlates with up-regulation of the glycolytic genes and poor survival in colon cancer patients. Remarkably, while ZBTB7A-deficient tumors progress exceedingly fast, they exhibit an unusually heightened sensitivity to glycolysis inhibition. Our study uncovers a novel tumor suppressor role of ZBTB7A in directly suppressing glycolysis.

Rig-I regulates NF-κB activity through binding to Nf-κb1 3′-UTR mRNA

Retinoic acid inducible gene I (RIG-I) senses viral RNAs and triggers innate antiviral responses through induction of type I IFNs and inflammatory cytokines. However, whether RIG-I interacts with host cellular RNA remains undetermined. Here we report that Rig-I interacts with multiple cellular mRNAs, especially Nf-κb1. Rig-I is required for NF-κB activity via regulating Nf-κb1 expression at posttranscriptional levels. It interacts with the multiple binding sites within 3′-UTR of Nf-κb1 mRNA. Further analyses reveal that three distinct tandem motifs enriched in the 3′-UTR fragments can be recognized by Rig-I. The 3′-UTR binding with Rig-I plays a critical role in normal translation of Nf-κb1 by recruiting the ribosomal proteins [ribosomal protein L13 (Rpl13) and Rpl8] and rRNAs (18S and 28S). Down-regulation of Rig-I or Rpl13 significantly reduces Nf-κb1 and 3′-UTR–mediated luciferase expression levels. These findings indicate that Rig-I functions as a positive regulator for NF-κB signaling and is involved in multiple biological processes in addition to host antivirus immunity.

Germinal Cell Aplasia in Kif18a Mutant Male Mice Due to Impaired Chromosome Congression and Dysregulated BubR1 and CENP-E

Chromosomal instability during cell division frequently causes cell death or malignant transformation. Orderly chromosome congression at the metaphase plate, a paramount process to vertebrate mitosis and meiosis, is controlled by a number of molecular regulators, including kinesins. Kinesin-8 (Kif18A) functions to control mitotic chromosome alignment at the mid-zone by negative regulation of kinetochore oscillation. Here the authors report that disrupting Kif18a function results in complete sterility in male but not in female mice. Histological examination reveals that Kif18a(-/-) testes exhibit severe developmental impairment of seminiferous tubules. Testis atrophy in Kif18a(-/-) mice is caused by perturbation of microtubule dynamics and spindle pole integrity, leading to chromosome congression defects during mitosis and meiosis. Depletion of KIF18A via RNAi causes mitotic arrest accompanied by unaligned chromosomes and increased microtubule nucleating centers in both GC-1 and HeLa cells. Prolonged depletion of KIF18A causes apoptosis due to perturbed microtubule dynamics. Further studies reveal that KIF18A silencing results in degradation of CENP-E and BubR1, which is accompanied by premature sister chromatid separation. KIF18A physically interacts with BubR1 and CENP-E, and this interaction is modulated during mitosis. Combined, the studies indicate that KIF18A is essential for normal chromosome congression during cell division and that the absence of KIF18A function causes severe defects in microtubule dynamics, spindle integrity, and checkpoint activation, leading to germinal cell aplasia in mice.