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Grand Rapids, MI, United States

Zarif J.C.,Van Andel Research Institute | Zarif J.C.,Michigan State University | Lamb L.E.,Beaumont Health System Research Institute | Schulz V.V.,Van Andel Research Institute | And 3 more authors.
Oncotarget | Year: 2015

Castration-resistant prostate cancers still depend on nuclear androgen receptor (AR) function despite their lack of dependence on exogenous androgen. Second generation anti-androgen therapies are more efficient at blocking nuclear AR; however resistant tumors still develop. Recent studies indicate Src is highly active in these resistant tumors. By manipulating AR activity in several different prostate cancer cell lines through RNAi, drug treatment, and the use of a nuclear-deficient AR mutant, we demonstrate that androgen acting on cytoplasmic AR rapidly stimulates Src tyrosine kinase via a non-genomic mechanism. Cytoplasmic AR, acting through Src enhances laminin integrin-dependent invasion. Active Matriptase, which cleaves laminin, is elevated within minutes after androgen stimulation, and is subsequently shed into the medium. Matriptase activation and shedding induced by cytoplasmic AR is dependent on Src. Concomitantly, CDCP1/gp140, a Matriptase and Src substrate that controls integrin-based migration, is activated. However, only inhibition of Matriptase, but not CDCP1, suppresses the AR/Src-dependent increase in invasion. Matriptase, present in conditioned medium from AR-stimulated cells, is sufficient to enhance invasion in the absence of androgen. Thus, invasion is stimulated by a rapid but sustained increase in Src activity, mediated non-genomically by cytoplasmic AR, leading to rapid activation and shedding of the laminin protease Matriptase.


Niemi N.M.,Van Andel Research Institute | Niemi N.M.,Van Andel Institute Graduate School | Lanning N.J.,Van Andel Research Institute | Westrate L.M.,Van Andel Research Institute | MacKeigan J.P.,Van Andel Research Institute
PLoS ONE | Year: 2013

Protein Tyrosine Phosphatase localized to the Mitochondrion 1 (PTPMT1) is a dual specificity phosphatase exclusively localized to the mitochondria, and has recently been shown to be a critical component in the cardiolipin biosynthetic pathway. The downregulation of PTPMT1 in pancreatic beta cells has been shown to increase cellular ATP levels and insulin production, however, the generalized role of PTPMT1 in cancer cells has not been characterized. Here we report that downregulation of PTPMT1 activity is sufficient to induce apoptosis of cancer cells. Additionally, the silencing of PTPMT1 decreases cardiolipin levels in cancer cells, while selectively increasing ATP levels in glycolytic media. Additionally, sublethal downregulation of PTPMT1 synergizes with low doses of paclitaxel to promote cancer cell death. Our data suggest that inhibition of PTPMT1 causes a metabolic crisis in cancer cells that induces cell death, and may be a mechanism by which cancer cells can be sensitized to currently available therapies. © 2013 Niemi et al.


Niemi N.M.,Van Andel Research Institute | Niemi N.M.,Van Andel Institute Graduate School | Mackeigan J.P.,Van Andel Research Institute
Antioxidants and Redox Signaling | Year: 2013

Significance: Apoptosis is a complex cellular process subject to multiple layers of regulation. One such layer of regulation includes post-translational modifications, including acetylation and phosphorylation. In particular, phosphorylation of proteins directly implicated in the apoptotic process has been extensively documented. Importantly, these phosphorylation events often have functional consequences, affecting the onset of apoptotic cell death. Recent Advances: Large-scale proteomics studies have identified multiple novel phosphorylation sites on proteins involved in the apoptotic process. The delineation of the regulation and functional consequences of these phosphorylation events will be important in understanding the regulatory complexity of apoptosis. Critical Issues: Multiple mitochondrial-localized proteins involved in apoptosis are functionally affected by phosphorylation, which can ultimately dictate whether a cell lives or dies. The dynamic interplay between these phosphorylated proteins and their regulatory enzymes is critical for understanding the complex cellular decision to undergo apoptosis. Future Directions: Detailed analysis of the kinetic and spatial regulation of phosphorylation events on apoptotic proteins, as well as how these dynamics influence the cell death process, will illuminate the complex interplay between the network of proteins that control the decision to undergo cell death. Antioxid. Redox Signal. 19, 572-582. © 2013, Mary Ann Liebert, Inc.


White Y.,University of California at San Francisco | Bagchi A.,Helen DeVos Childrens Hospital | Bagchi A.,Van Andel Institute Graduate School | Van Ziffle J.,University of California at San Francisco | And 8 more authors.
Nature Communications | Year: 2016

Oncogenic KRAS mutations introduce discrete amino acid substitutions that reduce intrinsic Ras GTPase activity and confer resistance to GTPase-activating proteins (GAPs). Here we discover a partial duplication of the switch 2 domain of K-Ras encoding a tandem repeat of amino acids G60-A66dup in a child with an atypical myeloproliferative neoplasm. K-Ras proteins containing this tandem duplication or a similar five amino acid E62-A66dup mutation identified in lung and colon cancers transform the growth of primary myeloid progenitors and of Ba/F3 cells. Recombinant K-RasG60-A66dup and K-RasE62-A66dup proteins display reduced intrinsic GTP hydrolysis rates, accumulate in the GTP-bound conformation and are resistant to GAP-mediated GTP hydrolysis. Remarkably, K-Ras proteins with switch 2 insertions are impaired for PI3 kinase binding and Akt activation, and are hypersensitive to MEK inhibition. These studies illuminate a new class of oncogenic KRAS mutations and reveal unexpected plasticity in oncogenic Ras proteins that has diagnostic and therapeutic implications.


Martin K.R.,Van Andel Research Institute | Barua D.,Los Alamos National Laboratory | Kauffman A.L.,Van Andel Research Institute | Westrate L.M.,Van Andel Research Institute | And 5 more authors.
Autophagy | Year: 2013

Macroautophagy (autophagy) is a cellular recycling program essential for homeostasis and survival during cytotoxic stress. This process, which has an emerging role in disease etiology and treatment, is executed in four stages through the coordinated action of more than 30 proteins. An effective strategy for studying complicated cellular processes, such as autophagy, involves the construction and analysis of mathematical or computational models. When developed and refined from experimental knowledge, these models can be used to interrogate signaling pathways, formulate novel hypotheses about systems, and make predictions about cell signaling changes induced by specific interventions. Here, we present the development of a computational model describing autophagic vesicle dynamics in a mammalian system. We used time-resolved, live-cell microscopy to measure the synthesis and turnover of autophagic vesicles in single cells. The stochastically simulated model was consistent with data acquired during conditions of both basal and chemicallyinduced autophagy. The model was tested by genetic modulation of autophagic machinery and found to accurately predict vesicle dynamics observed experimentally. Furthermore, the model generated an unforeseen prediction about vesicle size that is consistent with both published findings and our experimental observations. Taken together, this model is accurate and useful and can serve as the foundation for future efforts aimed at quantitative characterization of autophagy. © 2013 Landes Bioscience.

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