Result
Since STAT5 promotes leukemia cell survival, cell viability following STAT5 depletion was also assessed. Reduced STAT5 and BCL10 expression decreased Kit225 cell viability in a dose dependent manner, regardless of the absence or presence of IL-2 in the culture medium. This data further suggest that non-cytokine activated STAT5 dependent gene regulation may be important in tumor cells such as Kit225. IL-2 starved Kit225 cells were greater than 90% viable after 72 hours, although tyrosine phosphorylated STAT5 were destroyed within 24 hours. These results support the hypothesis that the leukemia cell survival promoting activities of STAT5 are, at least partially, cytokine independent; while targets such as BCL10 may be responsible for this phenotype. To support this notice, the effect of STAT5 depletion on NFκB function was explored. Nuclear proteins were isolated from STAT5 antisense and treated Kit225 cells for 24 hours and then incubated with [32P]-labeled NFκB probe. The results of this isolation showed reduced DNA binding of NFκB following STAT5 depletion as compared to control ODN treated samples. This data suggest that STAT5 regulates NFκB signaling in an IL-2-independent manner in Kit225 cells.
Conclusion
In conclusion, the results demonstrate that STAT5 mediated BCL10 expression occurs in the absence or presence of cytokine stimulation and STAT5 tyrosine phosphorylation. Also, the data indicates that STAT5 and that the NFκB pathways are all connected and critical for regulating leukemic cancer cell genes. The applicable relevance of these findings is that the therapeutic strategies that seek to stop the cancers progression by blocking STAT tyrosine phosphorylation status might not be effective and could possibly be tumor dependent. Therefore, both targeted disruption of tyrosine and non-tyrosine phosphorylated forms of STAT5 may be required to disrupt the growth and possibly kill the cancer cells.