Abstract
BackgroundMultiple myeloma (MM) is at present an incurable malignancy, characterized by apoptosis-resistant tumor cells. Interferon (IFN) treatment sensitizes MM cells to Fas-induced apoptosis and is associated with an increased activation of Signal transducer and activator of transcription (Stat)1. The role of Stat1 in MM has not been elucidated, but Stat1 has in several studies been ascribed a pro-apoptotic role. Conversely, IL-6 induction of Stat3 is known to confer resistance to apoptosis in MM.MethodsTo delineate the role of Stat1 in IFN mediated sensitization to apoptosis, sub-lines of the U-266-1970 MM cell line with a stable expression of the active mutant Stat1C were utilized. The influence of Stat1C constitutive transcriptional activation on endogenous Stat3 expression and activation, and the expression of apoptosis-related genes were analyzed. To determine whether Stat1 alone would be an important determinant in sensitizing MM cells to apoptosis, the U-266-1970-Stat1C cell line and control cells were exposed to high throughput compound screening (HTS).ResultsTo explore the role of Stat1 in IFN mediated apoptosis sensitization of MM, we established sublines of the MM cell line U-266-1970 constitutively expressing the active mutant Stat1C. We found that constitutive nuclear localization and transcriptional activity of Stat1 was associated with an attenuation of IL-6-induced Stat3 activation and up-regulation of mRNA for the pro-apoptotic Bcl-2 protein family genes Harakiri, the short form of Mcl-1 and Noxa. However, Stat1 activation alone was not sufficient to sensitize cells to Fas-induced apoptosis. In a screening of > 3000 compounds including bortezomib, dexamethasone, etoposide, suberoylanilide hydroxamic acid (SAHA), geldanamycin (17-AAG), doxorubicin and thalidomide, we found that the drug response and IC50 in cells constitutively expressing active Stat1 was mainly unaltered.ConclusionWe conclude that Stat1 alters IL-6 induced Stat3 activity and the expression of pro-apoptotic genes. However, this shift alone is not sufficient to alter apoptosis sensitivity in MM cells, suggesting that Stat1 independent pathways are operative in IFN mediated apoptosis sensitization.
Highlights
Multiple myeloma (MM) is at present an incurable malignancy, characterized by apoptosis-resistant tumor cells
We report here that constitutive transcriptional activation of Stat1 in the U-266-1970 MM cell was associated with an attenuation of Stat3 activation and a differential expression of several genes involved in apoptosis
In the present study we show that Stat3 protein was decreased and phosphorylation of Stat3 was attenuated in an IL-6 dependent MM cell line, the U-266-1970, expressing a constitutively active mutant of Stat1, the Stat1C
Summary
Multiple myeloma (MM) is at present an incurable malignancy, characterized by apoptosis-resistant tumor cells. Common complications of MM include anemia, renal dysfunction, and bone destruction Conventional drugs such as alkylating agents and. The aim of this study was to evaluate the importance of signal transducer and activator of transcription (Stat) 1 in influencing apoptosis and drug resistance in MM. Stat belongs to a family of transcription factors that are associated with regulation of growth and survival in hematopoietic cells. Upon cytokine stimulation, these proteins become phosphorylated by kinases, such as janus activated kinases (JAKs), associate with the cytoplasmic part of the cytokine receptor, form homo- or heterodimers and, translocate to the nucleus where they bind to specific DNA elements and directly regulate transcription [3,4]
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