Abstract
Solid tumor development is frequently accompanied by energy-deficient conditions such as glucose deprivation and hypoxia. Follistatin (FST), a secretory protein originally identified from ovarian follicular fluid, has been suggested to be involved in tumor development. However, whether it plays a role in cancer cell survival under energy-deprived conditions remains elusive. In this study, we demonstrated that glucose deprivation markedly enhanced the expression and nucleolar localization of FST in HeLa cells. The nucleolar localization of FST relied on its nuclear localization signal (NLS) comprising the residues 64-87. Localization of FST to the nucleolus attenuated rRNA synthesis, a key process for cellular energy homeostasis and cell survival. Overexpression of FST delayed glucose deprivation-induced apoptosis, whereas down-regulation of FST exerted the opposite effect. These functions depended on the presence of an intact NLS because the NLS-deleted mutant of FST lost the rRNA inhibition effect and the cell protective effect. Altogether, we identified a novel nucleolar function of FST, which is of importance in the modulation of cancer cell survival in response to glucose deprivation.
Highlights
Transcription of rRNA is catalyzed by polymerase I
Versus 42.5%), whereas knockdown of FST promoted glucose deprivation-induced apoptosis (61.1% versus 41.1%), further confirming the protective effect of FST (data not FST was traditionally recognized as a secretory protein and has been reported to exert its biological functions through extracellularly inactivating activin [36], myostatin [18], and bone morphogenic proteins
Nuclear localization apoptosis [34, 35], and down-regulating rRNA transcription is of FST has been observed in epithelial cells of breast tissue [37]
Summary
DMEM containing 25 mM glucose (Invitrogen) supplemented with 10% fetal bovine serum (Thermo Fisher) and penicillinstreptomycin mixed solution (Invitrogen). HeLa cells cultured with or without glucose were stained with FST monoclonal antibodies using the indirect immunofluorescence method and observed under a confocal microscope. The data showed that FST was detected in both the cytoplasm and the or FST/⌬NLS-expressing clones were obtained for further nucleus, whereas the nuclear FST increased when glucose experiments. Assays for Apoptosis Detection—Both floating (dead) and that FST is a nucleolar protein and that glucose deprivation attached cells were harvested and applied to different assays promotes its expression and nucleolar localization. Total proteins were extracted, and PARP tial NLS, a series of FST truncated fragments were fused to the cleavage was visualized by immunoblotting
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