The mitochondrial fractionation/western blotting rihanna puma slides ( Fig. 5a ) showed PUMA to be primarily localized in the non-mitochondrial fractions of U87MG-EGFRvIII cells under unstressed condition and to undergo a modest mitochondrial translocalization following exposure to the apoptotic inducers, staurosporin (ST) and anisomycin (AN). As indicated by the low mtPUMA index (0-0.02), the majority of PUMA is sequestered in the non-mitochondrial fractions of the EGFRvIII-expressing U87MGEGFRvIII cells ( Fig. 5a ).
The extent of PUMA mitochondrial translocalization, mtPUMA Index, is computed as described earlier in Materials and Methods. (A) PUMA is primarily localized in the cytoplasm of U87MG-EGFRvIII cells. U87MG-EGFRvIII cells puma fenty slides treated with vehicle control, staurosporin (ST, 1 uM) or anisomycin (AN, 100 ng/ml) were harvested and fractionated into mitochondrial and non-mitochondrial fractions. Protein extracts from both fractions were subjected to western blotting to puma shoes for women detect PUMA.
PUMA is primarily localized in the cytoplasm of these cells, under unstressed and stressed conditions, as indicated by the low mtPUMA indices.Similar observations were further found in T98G GBM cells that naturally express EGFR ( Fig. 5c ). The modest detection of mitochondrial PUMA in these cells may potentially be the result of insufficient cytoplasmic EGFR to interact with and sequester all the PUMA molecules rihanna puma shoes in the cytoplasm.
Importantly, siRNA-mediated EGFR expression knockdown led to a significant increase (16-fold) of mitochondrial PUMA ( Fig. 5c-left ), further suggesting the ability of EGFR to modulate PUMA mitochondrial translocalization. EGFR-specific siRNA was effective in reducing EGFR expression as shown by the western blots ( Fig. 5c-right ). In addition to GBM cells, we found PUMA to be localized in the cytoplasm of MDA-MB-468 human breast cancer cells .
That are known to express significant levels of endogenous EGFR [ 34 ]. Collectively, these results demonstrate that PUMA is sequestered in the cytoplasm of EGFR- and EGFRvIII-expressing puma blaze of glory GBM and breast cancer cells, constitutively and under apoptotic stress.To gain insight into the factors that modulate the interaction of EGFR with PUMA, we examined the requirement of EGFR activation for the EGFR-PUMA interaction.
Fig. 6a (left panel) shows that the ability of EGFR to bind to PUMA was similar in U87MG-EGFR cells with and without EGF stimultion following serum starvation. As indicated by the absence of auto-phosphorylated EGFR (p-EGFR, Y1068), serum-starved U87MG-EGFR cells express inactive EGFR ( Fig. 6a-right ). In contrast, p-EGFR is readily detected in EGF-treated cells, [img]http://www.czcrush.com/images/detail/puma blaze of glory-230wmy.jpg[/img] indicating that EGF efficiently activated EGFR in these cells.
 |
Strona 1 z 1 |
| |
Strona 1 z 1 |
|
|||||
 To forum działa w systemie phorum.pl Masz pomysł na forum? Załóż forum za darmo! Forum narusza regulamin? Powiadom nas o tym! Powered by Active24, phpBB © phpBB Group Design by phpBBStyles.com for LearnPHP.org |
|||||