Increased 14-3-3ζ Expression In The Multidrug-Resistant Leukemia Cell Line HL-60/Vcr As Compared To HL-60 Cell Line Mediates Cell Growth and Apoptosis

Abstract

Objectives Acute myeloid leukemia (AML) recurrence is largely a result of multidrug resistance (MDR). Because vincristine-resistant HL-60 cells (HL-60/VCR) express greater levels of 14-3-3¦Æ, which is associated with apoptosis and chemosensitivity in other cancers, this study sought to examine its role in AML chemosensitivity using HL-60 and HL-60/VCR cells. Methods The mRNA and protein expression of 14-3-3¦Æ, mdr1, Pgp, BCL-2 and MCL-1 were examined using semi-quantitative RT-PCR and Western blot analyses, respectively. The subcellular location of 14-3-3¦Æ protein in HL-60 and HL-60/VCR cells was determined using immunofluorescence and confocal microscopy. After siRNA-mediated silencing of 14-3-3¦Æ in HL-60 AND HL-60/VCR cells, cell growth and cell cycle progression were determined by direct counting and flow cytometry, respectively. The effect of 14-3-3¦Æ siRNA on topotecan (TPT)-induced apoptosis was evaluated using acridine orange/ethidium bromide and Annexin V staining as well as TUNEL analysis. NF-¦ÊB-DNA biding was also assessed using electrophoretic mobility shift assay. Results As compared to HL-60 cells, increased 14-3-3¦Æ mRNA and protein expression was observed in HL-60/VCR cells. In addition, increased mdr-1 mRNA as well as Pgp, Bcl-2, and Mcl-1 protein expression were observed in HL-60/VCR cells. In both HL-60 and HL-60/VCR cells, 14-3-3¦Æ was observed in the cytoplasm and nuclear compartments. 14-3-3¦Æ siRNA significantly reduced HL-60 and HL-60/VCR cell growth after 48 h and increased the proportion of cells in the G0/G1 phase. Moreover, 14-3-3¦Æ siRNA significantly increased the sensitivity of both HL-60 and HL-60/VCR cells to TPT possibly through inhibition of Bcl-2, Mcl-1 and Pgp protein expression. Conversely, increased Bad and Noxa protein expression was observed with 14-3-3¦Æ siRNA. NF-ĸB DNA binding was reduced with 14-3-3¦Æ siRNA. Conclusions Silencing of 14-3-3¦Æ increased the sensitivity of both sensitive and resistant HL-60 cells to TPT-induced apoptosis possibly through altering the expression of apoptosis-associated proteins, suggesting that it may be a potential target for MDR AML. Disclosures: No relevant conflicts of interest to declare.

Cetacean morbillivirus in Humpback whales’ exhaled breath

“…This gene encodes proteins involved in a wide range of biological processes. Specifically, YWHAZ mediates signal transduction pathways through binding to phosphoserine‐ and phosphothreonine‐containing proteins and interacts with many apoptotic proteins, playing a role in regulating cell survival (Liang et al., 2014).…”

“…This gene encodes proteins involved in a wide range of biological processes. Specifically, YWHAZ mediates signal transduction pathways through binding to phosphoserine-and phosphothreonine-containing proteins and interacts with many apoptotic proteins, playing a role in regulating cell survival (Liang et al, 2014). The PCR product of positive samples was purified using the QIAquick PCR Purification Kit (Qiagen) and directly sequenced from both ends by the SANGER method, using an ABI 3730 DNA analyser and an ABI BigDye Terminator cycle sequencing kit (Applied Biosystems).…”

14-3-3ζ negatively regulates mitochondrial biogenesis in GBM residual cells

“…The 14-3-3 family proteins are involved in cellular processes like intracellular signalling, differentiation, proliferation, transcriptional regulation, cytoskeletal organization, stress signalling, apoptosis, and tumorigenesis [5,6,7,8]. Of the seven isoforms of 14-3-3 protein, 14-3-3ζ plays a pivotal role in regulating multiple signalling pathways in cancer development, progression, and therapy resistance [9,10,11]. It is known to be overexpressed and associated with poor prognosis in breast, lung, head and neck, and glioblastoma [12].…”