Nuclear translocation of the Hsp70/Hsp90 organizing protein mSTI1 is regulated by cell cycle kinases

Longshaw, Victoria M.; Chapple, J. Paul; Balda, María S.; Cheetham, Michael E.; Blatch, Gregory L.

doi:10.1242/jcs.00905

Cited by 111 publications

(89 citation statements)

References 44 publications

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“…There is evidence to suggest that localisation of Hop may be due to phosphorylation. Hop can be phosphorylated by cell cycle kinases, phosphorylation by cdc2 (cell division cycle 2) kinase promotes cytoplasmic retention, whereas phosphorylation by casein kinase II (CKII) promotes nuclear Hop, suggesting that Hop might move between the cytoplasm and the nucleus under certain cell cycle conditions [40,41]. The nuclear accumulation of Hop is primarily stress regulated and CKII phosphorylation has been shown not to disrupt Hop-Hsp90 binding [36].…”

Section: Discussionmentioning

confidence: 99%

RNAi knockdown of Hop (Hsp70/Hsp90 organising protein) decreases invasion via MMP-2 down regulation

et al. 2011

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Keywords:Hop/STIP1/STI1 (Hsp70/Hsp90-organising protein) Pancreatic cancer Immunohistochemistry In vitro invasion MMP-2 a b s t r a c tWe previously identified Hop as over expressed in invasive pancreatic cancer cell lines and malignant tissues of pancreatic cancer patients, suggesting an important role for Hop in the biology of invasive pancreatic cancer. Hop is a co-chaperone protein that binds to both Hsp70/Hsp90. We hypothesised that by targeting Hop, signalling pathways modulating invasion and client protein stabilisation involving Hsp90-dependent complexes may be altered.In this study, we show that Hop knockdown by small interfering (si)RNA reduces the invasion of pancreatic cancer cells, resulting in decreased expression of the downstream target gene, matrix metalloproteinases-2 (MMP-2). Hop in conditioned media co-immunoprecipitates with MMP-2, implicating a possible extracellular function for Hop. Knockdown of Hop expression also reduced expression levels of Hsp90 client proteins, HER2, Bcr-Abl, c-MET and v-Src. Furthermore, Hop is strongly expressed in high grade PanINs compared to lower PanIN grades, displaying differential localisation in invasive ductal pancreatic cancer, indicating that the localisation of Hop is an important factor in pancreatic tumours.Our data suggests that the attenuation of Hop expression inactivates key signal transduction proteins which may decrease the invasiveness of pancreatic cancer cells possibly through the modulation of Hsp90 activity. Therefore, targeting Hop in pancreatic cancer may constitute a viable strategy for targeted cancer therapy.

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Section: Discussionmentioning

confidence: 99%

RNAi knockdown of Hop (Hsp70/Hsp90 organising protein) decreases invasion via MMP-2 down regulation

et al. 2011

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“…Stress-induced phosphoprotein-1 was shown to interact with Hsp70 and Hsp90 and facilitates transfer of substrates from Hsp70 to Hsp90 and is important for proper protein folding, maturation, and cell cycle progression (8,22,36,67). We propose that severe downregulation of stress-induced phosphoprotein-1 with high dose alone might have disturbed the key interaction between stress-induced phosphoprotein-1 and various HSPs, leading to cell death, compared with sustained expression of stress-induced phosphoprotein-1 and activation of survival pathways in autoprotected mice.…”

Section: Discussionmentioning

confidence: 99%

“…Group III consisted of five mice to ensure a minimum of three surviving mice, because mice started dying between 108 and 120 h (36 -48 h after the high dose) in the lethality experiment (Table 1). Mice were terminated under diethyl ether anesthesia at 6,12,24,36,72,78,84,96,144,240, and 336 h after the low-dose administration in groups II and IV. Mice in group III (n ϭ 3/time point) were terminated after the administration of DW at 6,12,24,36,72,78,84,96, and 108 h. Plasma was collected from mice at each time point to assess renal function.…”

Section: Reagentsmentioning

confidence: 99%

“…Mice were terminated under diethyl ether anesthesia at 6,12,24,36,72,78,84,96,144,240, and 336 h after the low-dose administration in groups II and IV. Mice in group III (n ϭ 3/time point) were terminated after the administration of DW at 6,12,24,36,72,78,84,96, and 108 h. Plasma was collected from mice at each time point to assess renal function. Kidneys were also isolated from these mice at each time point after respective treatment for further analysis of renal ATP content, histopathology, and immunohistochemical staining.…”

Section: Reagentsmentioning

confidence: 99%

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Proteomics ofS-(1, 2-dichlorovinyl)-l-cysteine-induced acute renal failure and autoprotection in mice

Korrapati¹,

Chilakapati²,

Witzmann³

et al. 2007

American Journal of Physiology-Renal Physiology

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“…Our in vitro kinase assay confirmed that Y354 was indeed the direct phosphorylation site of SYK. Besides, it is known that STIP1 promotes the association between 70 kDa heat shock cognate protein (HSC70) and heat shock protein 90 (HSP90) (52). HSC70 and HSP90 can be phosphorylated by SYK both in vitro and in vivo (28).…”

Section: Validating Novel Syk Substrates In Vitro and In Vivo-mentioning

confidence: 99%

Identification of Direct Tyrosine Kinase Substrates Based on Protein Kinase Assay-Linked Phosphoproteomics

Xue¹,

Geahlen

Tao

2013

Molecular & Cellular Proteomics

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Protein kinases are implicated in multiple diseases such as cancer, diabetes, cardiovascular diseases, and central nervous system disorders. Identification of kinase substrates is critical to dissecting signaling pathways and to understanding disease pathologies. However, methods and techniques used to identify bona fide kinase substrates have remained elusive. Here we describe a proteomic strategy suitable for identifying kinase specificity and direct substrates in high throughput. This approach includes an in vitro kinase assay-based substrate screening and an endogenous kinase dependent phosphorylation profiling. In the in vitro kinase reaction route, a pool of formerly phosphorylated proteins is directly extracted from whole cell extracts, dephosphorylated by phosphatase treatment, after which the kinase of interest is added. Protein phosphorylation plays a pivotal role in regulating biological events such as protein-protein interactions, signal transduction, subcellular localization, and apoptosis (1). Deregulation of kinase-substrate interactions often leads to disease states such as human malignancies, diabetes, and immune disorders (2). Although a number of kinases are being targeted to develop new drugs, our understanding of the precise relationships between protein kinases and their direct substrates is incomplete for the majority of protein kinases (3). Thus, mapping kinase-substrate relationships is essential for the understanding of biological signaling networks and the discovery and development of drugs for targeted therapies (4). Toward this goal, various in vitro kinase assays using synthetic peptide libraries (5), phage expression libraries (6), protein arrays (7-9), or cell extracts (10, 11) have been explored for the screening of kinase substrates.Besides classical biochemical and genetic methods, mass spectrometry-based high throughput approaches have become increasingly attractive because they are capable of sequencing proteins and localizing phosphorylation sites at the same time. Mass spectrometry-based proteomic methods have been extensively applied to kinase-substrate interaction mapping (12) and global phosphorylation profiling (13-15). Although thousands of phosphorylation events can be inspected simultaneously (16,17), large-scale phosphoproteomics does not typically reveal direct relationships between protein kinases and their substrates.Recently, several mass spectrometry-based proteomic strategies have been introduced for identifying elusive kinase substrates (7,18,19). Taking advantage of recent advances of high speed and high-resolution mass spectrometry, these methods used purified, active kinases to phosphorylate cell extracts in vitro, followed by mass spectrometric analysis to identify phosphoproteins. These approaches commonly face the major challenge of distinguishing phosphorylation events triggered by the kinase reaction from background signals introduced by endogenous kinase activities (20). To dissect the phosphorylation cascade, Shokat and colleagues developed an appro...

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Nuclear translocation of the Hsp70/Hsp90 organizing protein mSTI1 is regulated by cell cycle kinases

Cited by 111 publications

References 44 publications

RNAi knockdown of Hop (Hsp70/Hsp90 organising protein) decreases invasion via MMP-2 down regulation

RNAi knockdown of Hop (Hsp70/Hsp90 organising protein) decreases invasion via MMP-2 down regulation

Proteomics ofS-(1, 2-dichlorovinyl)-l-cysteine-induced acute renal failure and autoprotection in mice

Identification of Direct Tyrosine Kinase Substrates Based on Protein Kinase Assay-Linked Phosphoproteomics

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