Search

Glycogen synthase kinase-3 (GSK3) are ubiquitously expressed serine-threonine kinases involved in a plethora of functions ranging from the control of glycogen metabolism to transcriptional regulation. We recently demonstrated that GSK3 inhibition triggers JNK-cJUN-dependent apoptosis in human pancreatic cancer cells. However, the comprehensive picture of downstream GSK3-regulated pathways/functions remains elusive. Herein, counterbalancing the death signals, we show that GSK3 inhibition induces prosurvival signals through increased activity of the autophagy/lysosomal network. Our data also reveal a contribution of GSK3 in the regulation of the master transcriptional regulator of autophagy and lysosomal biogenesis, transcription factor EB (TFEB) in pancreatic cancer cells. Similarly to mammalian target of rapamycin (mTOR) inhibition, GSK3 inhibitors promote TFEB nuclear localization and leads to TFEB dephosphorylation through endogenous serine/ threonine phosphatase action. However, GSK3 and mTOR inhibition impinge differently and independently on TFEB phosphorylation suggesting that TFEB is regulated by a panel of kinases and/or phosphatases. Despite their differential impact on TFEB phosphorylation, both GSK3 and mTOR inhibitors promote 14-3-3 dissociation and TFEB nuclear localization. Quantitative mass spectrometry analyses further reveal an increased association of TFEB with nuclear proteins upon GSK3 and mTOR inhibition suggesting a positive impact on TFEB transcriptional function. Finally, a predominant nuclear localization of TFEB is unveiled in fully fed pancreatic cancer cells, whereas a reduction in TFEB expression significantly impairs their capacity for growth in an anchorage-independent manner. In addition, TFEB-restricted cells are more sensitive to apoptosis upon GSK3 inhibition. Altogether, our data uncover new functions under the control ofGSK3in pancreatic cancer cells in addition to providing key insight into TFEB regulation. [ABSTRACT FROM AUTHOR]

Tumor hypoxia is strongly associated with malignant progression such as increased cell invasion and metastasis. Although the invasion-related genes affected by hypoxia have been well described, the contribution of post-transcriptional mechanisms such as protein trafficking and proprotein processing associated with the hypoxic response remains poorly understood. The proprotein convertase furin, the major processing enzyme of the secretory pathway, resides in the trans-Golgi network and most studies support a model where endogenous substrates are processed by furin within this compartment. Here, we report that hypoxia triggered an unexpected relocalization of furin from the trans-Golgi network to endosomomal compartments and the cell surface in cancer cells. Exposing these cells back to normoxic conditions reversed furin redistribution, suggesting that the tumor microenvironment modulates furin trafficking in a highly regulated manner. Assessment of the mechanisms involved revealed that both Rab4GTPase-dependent recycling and interaction of furin with the cytoskeletal anchoring protein, filamin-A, are essential for the cell surface relocalization of furin. Interference with the association of furin with filamin-A, prevented cell surface relocalization of furin and abolished the ability of cancer cells to migrate in response to hypoxia. Our observations support the notion that hypoxia promotes the formation of a peripheral processing compartment where furin translocates for enhanced processing of proproteins involved in tumorigenesis. J. Cell. Physiol. 227: 789-800, 2012. © 2011 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

The hypoxic and acidic microenvironments in tumors are strongly associated with malignant progression and metastasis, and have thus become a central issue in tumor physiology and cancer treatment. Despite this, the molecular links between acidic pH- and hypoxia-mediated cell invasion/metastasis remain mostly unresolved. One of the mechanisms that tumor cells use for tissue invasion is the generation of invadopodia, which are actin-rich invasive plasma membrane protrusions that degrade the extracellular matrix. Here, we show that hypoxia stimulates the formation of invadopodia as well as the invasive ability of cancer cells. Inhibition or shRNA-based depletion of the Na+/H+ exchanger NHE-1, along with intracellular pH monitoring by live-cell imaging, revealed that invadopodia formation is associated with alterations in cellular pH homeostasis, an event that involves activation of the Na+/H+ exchange rate by NHE-1. Further characterization indicates that hypoxia triggered the activation of the p90 ribosomal S6 kinase (p90 RSK), which resulted in invadopodia formation and site-specific phosphorylation and activation of NHE-1. This study reveals an unsuspected role of p90RSK in tumor cell invasion and establishes p90RS kinase as a link between hypoxia and the acidic microenvironment of tumors. [ABSTRACT FROM AUTHOR]