Targeting a ribonucleoprotein complex containing the caprin-1 protein and the c-Myc mRNA suppresses tumor growth in mice: an identification of a novel oncotarget

Tylophorine compounds have been the focus of drug development for decades. Tylophorine derivatives exhibit anti-cancer activities but their cellular targets remain unknown. We used a biotinylated tylophorine derivative to probe for the interacting cellular target(s) of tylophorine. Tylophorine directly binds to caprin-1 and consequently enhances the recruitment of G3BP1, c-Myc mRNA, and cyclin D2 mRNA to form a ribonucleoprotein complex. Subsequently, this tylophorine targeted ribonucleoprotein complex is sequestered to the polysomal fractions and the protein expressions of the associated mRNA-transcripts are repressed. Caprin-1 depleted carcinoma cells become more resistant to tylophorine, associated with decreased formation of the ribonucleoprotein complex targeted by tylophorine. Consequently, tylophorine downregulates c-Myc and cyclins D1/D2, causing hypophosphorylation of Rb and suppression of both processing-body formation and the Warburg effect. Gene expression profiling and gain-of-c-Myc-function experiments also revealed that the downregulated c-Myc contributes to the anti-oncogenic effects of tylophorine compounds. Furthermore, the potent tylophorine derivative dibenzoquinoline-33b elicited a similar effect, as c-Myc protein levels were also decreased in xenograft tumors treated with dibenzoquinoline-33b. Thus, tylophorine compounds exert anti-cancer activity predominantly by targeting and sequestering the caprin-1 protein and c-Myc mRNA associated ribonucleoprotein complex

Chemo- and Thermosensory Responsiveness of Grueneberg Ganglion Neurons Relies on Cyclic Guanosine Monophosphate Signaling Elements

Neurons of the Grueneberg ganglion (GG) in the anterior nasal region of mouse pups respond to cool temperatures and to a small set of odorants. While the thermosensory reactivity appears to be mediated by elements of a cyclic guanosine monophosphate (cGMP) cascade, the molecular mechanisms underlying the odor-induced responses are unclear. Since odor-responsive GG cells are endowed with elements of a cGMP pathway, specifically the transmembrane guanylyl cyclase subtype GC-G and the cyclic nucleotide-gated ion channel CNGA3, the possibility was explored whether these cGMP signaling elements may also be involved in chemosensory GG responses. Experiments with transgenic mice deficient for GC-G or CNGA3 revealed that GG responsiveness to given odorants was significantly diminished in these knockout animals. These findings suggest that a cGMP cascade may be important for both olfactory and thermosensory signaling in the GG. However, in contrast to the thermosensory reactivity, which did not decline over time, the chemosensory response underwent adaptation upon extended stimulation, suggesting that the two transduction processes only partially overlap. Copyright (C) 2011 S. Karger AG, Base

Guanylyl cyclase-G modulates jejunal apoptosis and inflammation in mice with intestinal ischemia and reperfusion.

Membrane bound guanylyl cyclase-G (mGC-G), a novel form of GC mediates ischemia and reperfusion (IR)-induced renal injury. We investigated the roles of mGC-G in intestinal IR-induced jejunal damage, inflammation, and apoptosis.Male C57BL/6 wild-type (WT) and mGC-G gene knockout (KO) mice were treated with a sham operation or 45 min of superior mesenteric arterial obstruction followed by 3, 6, 12, or 24 h of reperfusion.Sham-operated KO mice had significantly lower plasma nitrate and nitrite (NOx) levels and jejunal villus height, crypt depth, and protein expression of phosphorylated-nuclear factor-kappa-B (NF-κB), phosphorylated-c-Jun N-terminal kinases (JNK) 2/3, phosphorylated-p38, and B-cell lymphoma-2 (Bcl-2). They had significantly greater jejunal interleukin-6 mRNA, cytochrome c protein, and apoptotic index compared with sham-operated WT mice. Intestinal IR significantly decreased plasma NOx in WT mice and increased plasma NOx in KO mice. The jejunal apoptotic index and caspase 3 activities were significantly increased, and nuclear phosphorylated-NF-κB and phosphorylated-p38 protein were significantly decreased in WT, but not KO mice with intestinal IR. After reperfusion, KO mice had an earlier decrease in jejunal cyclic GMP, and WT mice had an earlier increase in jejunal proliferation and a later increase in cytosol inhibitor of kappa-B-alpha. Intestinal IR induced greater increases in plasma and jejunal interleukin-6 protein in WT mice and a greater increase in jejunal interleukin-6 mRNA in KO mice.mGC-G is involved in the maintenance of jejunal integrity and intestinal IR-induced inflammation and apoptosis. These results suggest that targeting cGMP pathway might be a potential strategy to alleviate IR-induced jejunal damages

The apoptotic signaling pathway activated by Toll-like receptor-2

The innate immune system uses Toll family receptors to signal for the presence of microbes and initiate host defense. Bacterial lipoproteins (BLPs), which are expressed by all bacteria, are potent activators of Toll-like receptor-2 (TLR2). Here we show that the adaptor molecule, myeloid differentiation factor 88 (MyD88), mediates both apoptosis and nuclear factor-κB (NF-κB) activation by BLP-stimulated TLR2. Inhibition of the NF-κB pathway downstream of MyD88 potentiates apoptosis, indicating that these two pathways bifurcate at the level of MyD88. TLR2 signals for apoptosis through MyD88 via a pathway involving Fas-associated death domain protein (FADD) and caspase 8. Moreover, MyD88 binds FADD and is sufficient to induce apoptosis. These data indicate that TLR2 is a novel ‘death receptor’ that engages the apoptotic machinery without a conventional cytoplasmic death domain. Through TLR2, BLP induces the synthesis of the precursor of the pro-inflammatory cytokine interleukin-1β (IL-1β). Interestingly, BLP also activates caspase 1 through TLR2, resulting in proteolysis and secretion of mature IL-1β. These results indicate that caspase activation is an innate immune response to microbial pathogens, culminating in apoptosis and cytokine production

Molecules of the mitogen-activated protein kinases (MAPK) signaling pathway in the jejunum.

<p>A western blot assay to detect the protein expression of total and phosphorylated JNK, ERK, and p38 and the internal control β-actin (A). Protein expression of phosphorylated JNK1 (B), phosphorylated JNK 2/3 (C), phosphorylated ERK1 (D), phosphorylated ERK2 (E), and phosphorylated p38 (F). Protein quantification was carried out by densitometric analysis, normalized by the internal control β-actin, and calculated as the percentages of the WT-SH group. Means ± SEM, n = 10 for the WT groups and n = 9 for the KO groups. *P<.05, KO vs. WT mice with sham operation or with intestinal IR at the same time point (Student’s t-test,). Values with different superscripts indicate significant differences within all of the time points in WT and KO mice, individually (one-way ANOVA with LSD, P<.05).</p

Immunofluorescence analysis of apoptotic cells and immunohistochemical analysis of cell proliferation in jejunal sections.

<p><i>In situ</i> detection of apoptotic cells using TUNEL assay is shown with green fluorescein label in the jejunum of WT (A) and KO (B) mice. A negative control section treated with DNase without transferase is shown in the bottom right (A). The total nuclei stained with 4′,6-diamino-2-phenylindole (DAPI) were detected by UV light (blue) as shown in the bottom right (B). <i>In situ</i> detection of proliferative cells using Ki-67 staining is shown in the brown labeled jejunal crypts in WT (C) and KO (D) mice. The apoptotic index (E) and proliferation index (F) in per 5 villi and per 10 crypts, respectively. Means ± SEM, n = 10 for the WT groups and n = 9 for the KO groups. *P<.05, KO vs. WT mice with sham operation or with intestinal IR at the same time point (Student’s t-test,). Values with different superscripts indicate significant differences within all of the time points in WT and KO mice, individually (one-way ANOVA with LSD, P<.05).</p

Jejunal morphology. H&E staining in WT (A) and KO (B) mice.

<p>Scale bars  = 25 µm. The average villus height (C), crypt depth (D), and mucosal dry weight (E) and protein content (F) per centimeter of jejunum. Means ± SEM, n = 10 for WT groups and n = 9 for KO groups. *P<.05, KO vs. WT mice with sham operation or with intestinal IR at the same time point (Student’s t-test,). Values with different superscripts indicate significant differences within all of the time points in WT and KO mice, individually (one-way ANOVA with LSD, P<.05).</p