In Situ Hybridization in Cancer and Normal Tissue

Kadkol, Shrihari S.; Juang, Jyuhn‐Huarng; Wu, T. C.

doi:10.1385/1-59259-329-1:51

Cited by 2 publications

(2 citation statements)

References 68 publications

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“…23,2003 SILENCING OF Snk/Plk2 AND MITOTIC CATASTROPHE 5557 with a Dig RNA labeling kit (SP6/T7) (Roche). In situ hybridization was performed as described previously (13,18). Flow cytometry.…”

Section: Methodsmentioning

confidence: 99%

Silencing of the Novel p53 Target Gene Snk/Plk2 Leads to Mitotic Catastrophe in Paclitaxel (Taxol)-Exposed Cells

Burns

Fei

Scata

et al. 2003

Molecular and Cellular Biology

216

207

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Loss of p53 sensitizes to antimicrotubule agents in human tumor cells, but little is known about its role during mitosis. We have identified the Polo-like kinase family member serum inducible kinase (Snk/Plk2) as a novel p53 target gene. Snk/Plk2 mutagenesis demonstrated that its kinase activity is negatively regulated by its C terminus. Small interfering RNA (siRNA)-mediated Snk/Plk2 silencing in the presence of the mitotic poisons paclitaxel (Taxol) or nocodazole significantly increased apoptosis, similar to p53 mutations, which confer paclitaxel sensitivity. Furthermore, we have demonstrated that the apoptosis due to silencing of Snk/Plk2 in the face of spindle damage occurs in mitotic cells and not in cells that have progressed to a G 1 -like state without dividing. Since siRNA directed against Snk/Plk2 promoted death of paclitaxel-treated cells in mitosis, we envision a mitotic checkpoint wherein p53-dependent activation of Snk/Plk2 prevents mitotic catastrophe following spindle damage. Finally, these studies suggest that disruption of Snk/Plk2 may be of therapeutic value in sensitizing paclitaxel-resistant tumors.

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Section: Methodsmentioning

confidence: 99%

Silencing of the Novel p53 Target Gene Snk/Plk2 Leads to Mitotic Catastrophe in Paclitaxel (Taxol)-Exposed Cells

Burns

Fei

Scata

et al. 2003

Molecular and Cellular Biology

216

207

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“…The following ISH protocol (Kadkol et al, 2003) was used with minor modifications. After deparaffinization and rehydration in graded alcohols, esophageal sections were heated in citrate buffer (0.01 M, pH 6.0) in a microwave oven (90-941C, 3 Â 5 min) and digested with proteinase K (371C, 15 min).…”

Section: Ish Analysismentioning

confidence: 99%

Differentially expressed genes execute zinc-induced apoptosis in precancerous esophageal epithelium of zinc-deficient rats

et al. 2004

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Zinc deficiency (ZD) in rats increases esophageal cell proliferation and the incidence of N-nitrosomethylbenzylamine-induced esophageal tumors. Conversely, zinc replenishment (ZR) rapidly induces apoptosis in esophageal epithelia and reverses cancer development. We investigated gene expression changes in ZR versus ZD esophageal epithelia to identify differentially expressed genes associated with the antitumor effect of ZR. Weanling rats were fed a ZD diet for 6 weeks to establish esophageal cell proliferation or a zinc-sufficient (ZS) diet. Then, 10 ZD rats were treated with zinc gluconate intragastrically and switched to ZS diet; the remaining 10 ZD and ZS animals were treated with saline. All animals were killed 26-28 h later. Using cDNA microarrays, realtime polymerase chain reaction amplification and RNA hybridization techniques, we identified novel differentially expressed genes, including a RNA-binding protein with two RNA recognition motifs and a zinc knuckle (ZD7), and a DNA/RNA helicase with a DEAD box (ZD10) with two splice variants, ZD10a and ZD10b. In situ hybridization detected increased mRNA expression of ZD7, ZD10a and ZD10b in ZR esophageal epithelia, which displayed markedly increased occurrence of apoptotic cells, relative to ZD epithelia. Overexpression of ZD7 in human esophageal cancer cells resulted in induction of apoptosis and activation of caspase-3 and -7, activities that were inhibited by caspase-specific inhibitors. In addition, ZD7 mRNA levels and zinc-induced apoptosis in rat squamous carcinoma cells were reduced by specific small interfering ribonucleic acids. Thus, ZR rapidly induces ZD7 and ZD10 expression, which in turn stimulates apoptosis. These results provide the beginnings of a molecular pathway for zinc-induced apoptosis under conditions that reverse esophageal tumor initiation.

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In Situ Hybridization in Cancer and Normal Tissue

Cited by 2 publications

References 68 publications

Silencing of the Novel p53 Target Gene Snk/Plk2 Leads to Mitotic Catastrophe in Paclitaxel (Taxol)-Exposed Cells

Silencing of the Novel p53 Target Gene Snk/Plk2 Leads to Mitotic Catastrophe in Paclitaxel (Taxol)-Exposed Cells

Differentially expressed genes execute zinc-induced apoptosis in precancerous esophageal epithelium of zinc-deficient rats

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