Kindlin-2 promotes invasiveness of prostate cancer cells via NF-κB-dependent upregulation of matrix metalloproteinases.

Invasive progression is the major lethal cause of prostate cancer. In this study, we aimed to investigate the role of kindlin-2, an integrin-binding focal adhesion protein, in the regulation of invasiveness of prostate cancer. We found that downregulation of kindlin-2 using small interfering RNA (siRNA) technology significantly inhibited the invasion of PC-3 and DU-145 prostate cancer cells in a Matrigel Transwell assay. Conversely, overexpression of kindlin-2 promoted the invasiveness of prostate cancer cells. Kindlin-2 overexpression was found to activate nuclear factor (NF)-κB-dependent signaling and upregulate the expression of matrix metalloproteinase-9 (MMP-9) and MMP-2, whereas kindlin-2 silencing led to opposing effects on the expression of NF-κB and MMPs. Most importantly, kindlin-2-induced invasiveness was almost completely abolished by pretreatment with pyrrolidine dithiocarbamate (an inhibitor of NF-κB signaling) or co-transfection with MMP-9 or MMP-2 siRNA. Taken together, our data indicate that kindlin-2 promotes the invasiveness of prostate cancer cells largely through NF-κB-dependent upregulation of MMP-9 and MMP-2. Further studies are warranted to evaluate the significance of kindlin-2 as a therapeutic target for metastatic prostate cancer.

[Effects of adenovirus-mediated PTEN on the proliferation of prostate cancer PC-3 cells and expressions of cyclin D1 and p21].

OBJECTIVE: To construct a recombinant adenovirus expression vector containing the anti-oncogene PTEN and to investigate the effects of the PTEN gene on the proliferation of prostate cancer PC-3 cells and the expressions of cyclin D1 and p21 in the PC-3 cells. METHODS: The PTEN gene was amplified from the rat hippocampus by RT-PCR and cloned into the shuttle plasmid pEN-TR2A. The plasmids were constructed and amplified in 293A cells. Prostate cancer PC-3 cells were cultured in vitro and infected with the adenoviral vector carrying the PTEN gene (Ad-PTEN). The up-regulation of the PTEN protein was measured by indirect immuno-fluorescence assay; the expressions of PTEN, cyclin D1 and p21 in the cells infected with Ad-PTEN and Ad-LacZ were determined by RESULTS: The Western blot; and the effect of PTEN on the cell proliferation was detected by MTT assay and plate colony formation. recombinant adenoviral vector Ad-PTEN was successfully constructed. Western blot showed a significantly increased expression of the PTEN protein in the PC-3 cells infected with Ad-PTIEN (0.215 +/-0.065) as compared with that in the control ([0.052 +/-0.009], t = 4. 30, P <0.05) and the Ad-LacZ group ( [0. 056 +/- 0.008 ] , t =4.21, P <0.05). The expression of cyclin D1 was significantly lower in the Ad-PTEN-infected PC-3 cells (0. 256 +/- 0. 072) than in the control ( [0. 502 +/- 0. 087 ], t = 3.77, P < 0.05) and the Ad-LacZ group ([0.498 +/-0.081] , t =3.87, P <0.05), while the expression of p21 remarkably higher in the Ad-PTEN-infected PC-3 cells (0.589 +/-0. 076) than in the control ([0. 146 +/-0.026] , t = 9.55, P<0. 01) and the Ad-LacZ group ([0. 163 +/-0. 024] , t = 9.26, P <0.01). Ad-PTEN significantly inhibited the growth of the PC-3 cells (21.98%) at 48 h (t = 6.80, P <0.01). The colony formation rate of the PC-3 cells was (37.4 +/-4. 18)% in the Ad-PTEN group, significantly lower than (54.9 +/-4.81)% in the control (t =4.76, P<0.01) and (56.5 +/- 5.42)% in the Ad-LacZ group (t=4.83, P<0.01). CONCLUSION: The expression of PTEN induced by Ad-PTEN can significantly inhibit the proliferation of PC-3 cells, down-regulate the expression of cyclin D1, and up-regulate the expression of p21.

[Effects of staurosporine on the proliferation and apoptosis of prostate cancer PC-3 cells].

OBJECTIVE: To investigate the effects of staurosporine (ST) on the proliferation and apoptosis of prostate cancer PC-3 cells. METHODS: Prostate cancer PC-3 cells were treated in vitro with ST at 10(-8) mol/L. The expressions of cyclin A and cyclin D1 proteins in the cells were detected by Western blot, the effect of ST on the proliferation of the cells determined by MTT assay and plate colony formation, the apoptosis of the cells examined by flow cytometry, and their morphological changes observed under the light microscope. RESULTS: ST treatment markedly decreased the expressions of cyclin A and cyclin D1 in the PC-3 cells, and significantly inhibited the growth of the PC-3 cells (19.35%) at 48 h. (F = 31.06, P < 0.01). The colony formation rate of the PC-3 cells was (37.10 +/- 3.43) % in the ST group, significantly lower than (64.80 +/- 4.34) % in the control (chi2 = 14.59, P < 0.05) and (62.80 +/- 4.36) % in the DMSO group (chi2 = 12.50, P < 0.05), while the apoptosis rate of the cells was remarkably higher in the ST group ([19.6 +/- 2.20] %) than in the control ([5.33 +/- 1.40] %) and the DMSO group ([5.50 +/- 0.96] %) (F = 104.36, P < 0.01). Under the light microscope, the ST-treated cells were round with indistinct margins as compared with those of the other two groups. CONCLUSION: ST could significantly inhibit the proliferation and induce the apoptosis of PC-3 cells.

Genomic characterization of a novel dsRNA virus detected in the phytopathogenic fungus Verticillium dahliae Kleb.

Four novel double-stranded RNA segments were detected in a Verticillium dahliae Kleb. strain (V. dahliae isolate 0-21), a causal fungal agent of Verticillium wilt disease of cotton. Each dsRNA genome segment contains a single large open reading frame (ORF) that encodes a distinctive protein with modest levels of sequence similarities to the corresponding putative proteins in the genus Chrysovirus. These include an RNA-dependent RNA polymerase (RdRp), a coat protein, an undefined replication-related protein and an ovarian tumor domain peptidase. Phylogenetic analysis of the four putative proteins unanimously indicated that they are evolutionarily related to viruses in Chrysovirus. The 5'- and 3'-untranslated regions of the four dsRNAs share highly similar internal sequence and contain conserved sequence stretches of UGAUAAAAAA(/U)UG(/U)AAAAA- (in the 5'-UTR) and -UUUACUACU (in the 3'-UTR), indicating that they have a common virus origin. Indeed, isometric virus-like particles (VLPs) with a diameter of approximately 34nm were extracted from the fungal mycelia, and the four dsRNA segments were also detected in the virus-like particle (VLP) fraction. These results suggest that the mycovirus with four different dsRNA genome segments from the fungal isolate 0-21 is a new member of the genus Chrysovirus. We named the virus Verticillium dahliae chrysovirus 1 (VdCV1).