Barbara A. Niemeyer

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Verified

Barbara verified their affiliation via an institutional email.

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• PhD
• Professor (Full) at Saarland University

Membrane contact sites (MCSs) are hubs allowing various cell organelles to coordinate their activities. The dynamic nature of these sites and their small size hinder analysis by current imaging techniques. To overcome these limitations, we here design a series of reversible chemogenetic reporters incorporating improved, low-affinity variants of spl...

Membrane contact sites (MCSs) are hubs allowing various cell organelles to coordinate their activities. The dynamic nature of these sites and their small size hinder analysis by current imaging techniques. To overcome these limitations, we here designed a series of reversible chemogenetic reporters incorporating improved, low-affinity variants of s...

By virtue of mitochondrial control of energy production, reactive oxygen species (ROS) generation, and maintenance of Ca²⁺ homeostasis, mitochondria play an essential role in modulating T cell function. The mitochondrial Ca²⁺ uniporter (MCU) is the pore-forming unit in the main protein complex mediating mitochondrial Ca²⁺ uptake. Recently, MCU has...

Balanced Ca2+ homeostasis is essential for cellular functions. STIM2 mediated Store-Operated Ca2+ Entry (SOCE) regulates cytosolic and ER Ca2+ concentrations, stabilizes dendritic spine formation and drives presynaptic spontaneous transmission and ER stress in neurons. Recently identified alternative spliced variants expand the STIM protein reperto...

Pathogenic variants in SPAST, the gene coding for spastin, are the single most common cause of hereditary spastic paraplegia, a progressive motor neuron disease. Spastin regulates key cellular functions, including microtubule-severing and endoplasmic reticulum-morphogenesis. However, it remains unclear how alterations in these cellular functions du...

Radiation therapy efficiently eliminates cancer cells and reduces tumor growth. To understand collateral agonistic and antagonistic effects of this treatment on the immune system, we examined the impact of x-ray irradiation on human T cells. We find that, in a major population of leukemic Jurkat T cells and peripheral blood mononuclear cells, clini...

Plasma membrane protein channels provide a passageway for ions to access the intracellular milieu. Rapid entry of calcium ions into cells is controlled mostly by ion channels, while Ca²⁺-ATPases and Ca²⁺ exchangers ensure that cytosolic Ca²⁺ levels ([Ca²⁺]cyt) are maintained at low (~100 nM) concentrations. Some channels, such as the Ca²⁺-release-a...

Alternative splicing is a potent modifier of protein function. Stromal interaction molecule 1 (Stim1) is the essential activator of store-operated Ca2+ entry (SOCE) triggering activation of transcription factors. Here, we characterize Stim1A, a splice variant with an additional 31 amino acid domain inserted in frame within its cytosolic domain. Pro...

The endoplasmic reticulum (ER) is extensively remodelled during the development of professional secretory cells to cope with high protein production. Since ER is the principal Ca2+ store in the cell, we characterised the Ca2+ homeostasis in NALM-6 and RPMI 8226 cells, which are commonly used as human pre-B and antibody secreting plasma cell models,...

Background: Cardiovascular diseases (CVD) and chronic kidney disease (CKD) are highly prevalent, aggravate each other, and account for substantial mortality. Both conditions are characterized by activation of the innate immune system. The alarmin IL-1α is expressed in a variety of cell types promoting (sterile) systemic inflammation. The aim of the...

Immune responses involve mobilization of T cells within naïve and memory compartments. Tightly regulated Ca²⁺ levels are essential for balanced immune outcomes. How Ca²⁺ contributes to regulating compartment stoichiometry is unknown. Here, we show that plasma membrane Ca²⁺ ATPase 4 (PMCA4) is differentially expressed in human CD4⁺ T compartments yi...

Aims Inflammation plays an important role in cardiovascular disease (CVD) development. The NOD-like receptor protein-3 (NLRP3) inflammasome contributes to the development of atherosclerosis in animal models. Components of the NLRP3 inflammasome pathway such as interleukin-1β can therapeutically be targeted. Associations of genetically determined in...

Immune responses involve mobilization of T cells within naïve and memory compartments. Tightly regulated Ca2+ levels are essential for balanced immune outcomes. How Ca2+ contributes to regulating compartment stoichiometry is unknown. Here, we show that plasma membrane Ca2+ ATPase 4 (PMCA4) is differentially expressed in human CD4+ T compartments yi...

Store-operated Ca2+-entry (SOCE) regulates basal and receptor-triggered Ca2+ signaling with STIM proteins sensing the endoplasmic reticulum (ER) Ca2+ content and triggering Ca2+ entry by gating Orai channels. Although crucial for immune cells, STIM1’s role in neuronal Ca2+ homeostasis is controversial. Here, we characterize a splice variant, STIM1B...

The Ca²⁺ selective channel ORAI1 and endoplasmic reticulum (ER)-resident STIM proteins form the core of the channel complex mediating store operated Ca²⁺ entry (SOCE). Using liquid phase electron microscopy (LPEM), the distribution of ORAI1 proteins was examined at rest and after SOCE-activation at nanoscale resolution. The analysis of over seven h...

Defective ER/SR-cytosol Ca2+ cycling is associated with increased ER stress, pathological heart conditions and muscular defects. Within the SR, ryanodine receptor 2 (RyR2) is required for excitation/contraction coupling. Ca2+ release from the SR is counterbalanced by K+ influx through trimeric intracellular cation (TRIC) channels to maintain ER/SR...

In CTLs: High glucose‐culture enhances thapsigargin‐induced SOCE but decreases target recognition‐induced Ca²⁺ influx. High glucose‐culture regulates expression of ORAIs and STIMs without affecting glucose uptake. More high glucose‐cultured CTLs are prone to necrosis after execution of killing.

The endoplasmic reticulum (ER) is a vast intracellular organelle important to maintain proteome, lipid and calcium homeostasis to ensure cellular functionality. Stromal Interaction Molecule (STIM) proteins are sensors that can respond to changes in ER Ca²⁺ concentration with conformational change, oligomerization and relocalization to ER–plasma mem...

The vanilloid capsaicin is a widely consumed spice, known for its burning and “hot” sensation through activation of TRPV1 ion-channels, but also known to decrease oxidative stress, inflammation and influence tau-pathology. Beside these positive effects, little is known about its effects on amyloid-precursor-protein (APP) processing leading to amylo...

The killing efficiency of cytotoxic T lymphocytes (CTLs) is tightly regulated by intracellular Ca2+ concentration. Glucose is the key energy source for CTLs, lack of which significantly impairs CTL activation, proliferation and effector functions. The impact of high glucose on Ca2+ influx in CTLs remains largely elusive. In this work, we stimulated...

Alternative splicing is a potent modifier of protein function. Stromal interaction molecule 1 (Stim1) is the essential activator molecule of store-operated Ca ²⁺ entry (SOCE) and a sorting regulator of certain ER proteins such as Stimulator of interferon genes (STING). Here, we characterize a conserved new variant, Stim1A, where splice-insertion tr...

The Ca ²⁺ selective channel ORAI1 and endoplasmic reticulum (ER)-resident STIM proteins form the core of the channel complex mediating store operated Ca ²⁺ entry (SOCE). Using liquid phase electron microscopy (LPEM) the distribution of ORAI1 proteins was examined at rest and after SOCE-activation at nanoscale resolution. The analysis of over seven...

Store-operated Ca2+-entry (SOCE) regulates basal and receptor-triggered Ca2+ signaling with STIM proteins sensing the endoplasmic reticulum (ER) Ca2+ content and triggering Ca2+ entry by gating Orai channels. Although crucial for immune cells, STIM1’s role in neuronal Ca2+ homeostasis is controversial. Here, we characterize a splice variant, STIM1B...

ORAI1 proteins form highly selective Ca2+ channels in the plasma membrane. Crystallographic data point towards a hexameric stoichiometry of ORAI1 channels, whereas optical methods postulated ORAI1 channels to reside as dimers at rest, and other data suggests a tetrameric configuration. Here, liquid-phase scanning transmission electron microscopy (S...

NLRP3-inflammasome-driven inflammation is involved in the pathogenesis of a variety of diseases. Identification of endogenous inflammasome activators is essential for the development of new anti-inflammatory treatment strategies. Here, we identified that apolipoprotein C3 (ApoC3) activates the NLRP3 inflammasome in human monocytes by inducing an al...

Mammalian inner hair cells (IHCs) transduce sound into depolarization and transmitter release. Big conductance and voltage‐ and Ca²⁺‐activated K⁺ (BK) channels are responsible for fast membrane repolarization and small time constants of mature IHCs. For unknown reasons, they activate at around ‐75 mV with a voltage of half‐maximum activation (Vhalf...

Reactive oxygen species (ROS) are emerging as important regulators of cancer growth and metastatic spread. However, how cells integrate redox signals to affect cancer progression is not fully understood. Mitochondria are cellular redox hubs, which are highly regulated by interactions with neighboring organelles. Here, we investigated how ROS at the...

Basophils are a small population of innate immune cells, but their release of the cytokine interleukin-4 (IL-4) is important for mounting an efficient immune response against distinct parasites. Yoshikawa et al . (in the 9 April 2019 issue) showed that whereas STIM1 is essential for IL-4 release after stimulation of FcεRI, STIM2 mediates a delayed...

Release of Ca ²⁺ from endoplasmatic retriculum (ER) Ca ²⁺ stores causes stromal interaction molecules (STIM) in the ER membrane and ORAI proteins in the plasma membrane (PM) to interact and form the Ca ²⁺ release activated Ca ²⁺ (CRAC) channels, which represent a major Ca ²⁺ entry route in non-excitable cells and thus control various cell functions...

Quantitative Studies of Membrane Proteins in Whole Cells with Different Methods of Liquid Phase Scanning Transmission Electron Microscopy - Volume 25 Issue S1 - Diana B. Peckys, Dalia Alansary, Barbara Niemeyer, Niels de Jonge

In recent years, channels that mediate Store-Operated Calcium Entry (SOCE, i.e. the ability of cells to sense a decrease in endoplasmic reticulum luminal calcium and induce calcium entry across the plasma membrane) have been associated to a number of disorders, spanning from immune disorders to acute pancreatitis and have been suggested to be drugg...

Adjusting intracellular calcium signaling is an important feature in the regulation of immune cell function and survival. Here we show that miR-34a-5p, a small non-coding RNA that is deregulated in many common diseases, is a regulator of store-operated Ca2+ entry (SOCE) and calcineurin signaling. Upon miR-34a-5p overexpression, we observed both a d...

The oncometabolite (R)-2-hydroxyglutarate (R-2-HG) produced by isocitrate dehydrogenase (IDH) mutations promotes gliomagenesis via DNA and histone methylation. Here, we identify an additional activity of R-2-HG: tumor cell-derived R-2-HG is taken up by T cells where it induces a perturbation of nuclear factor of activated T cells transcriptional ac...

Application of Liquid Phase Scanning Transmission Electron Microscopy Methods for the Quantitative Study of Membrane Proteins in Whole Cells - Volume 24 Supplement - Diana B. Peckys, Indra N. Dahmke, Dalia Alansary, Barbara Niemeyer, Niels de Jonge

Release of $Ca^{2+}$ from endoplasmatic retriculum (ER) $Ca^{2+}$ stores causes stromal interaction molecules (STIM) in the ER membrane and ORAI proteins in the plasma membrane (PM) to interact and form the $Ca^{2+}$ release activated $Ca^{2+}$ (CRAC) channels, which represent a major $Ca^{2+}$ entry route in non-excitable cells and thus control va...

Release of Ca ²⁺ from endoplasmatic retriculum (ER) Ca ²⁺ stores causes stromal interaction molecules (STIM) in the ER membrane and ORAI proteins in the plasma membrane (PM) to interact and form the Ca ²⁺ release activated Ca ²⁺ (CRAC) channels, which represent a major Ca ²⁺ entry route in non-excitable cells and thus control various cell functions...

Differentiation of naïve CD4+T cells into effector subtypes with distinct cytokine profiles and physiological roles is a tightly regulated process, the imbalance of which can lead to an inadequate immune response or autoimmune disease. The crucial role of Ca2+signals, mainly mediated by the store operated Ca2+entry (SOCE) in shaping the immune resp...

Significance The work presents a unique understanding of the organization and function of two ubiquitously expressed proteins, central in generating calcium signals in all cell types. These are the intracellular calcium sensing “STIM” proteins, and the highly selective cell surface “Orai” calcium channels. We reveal that STIM proteins can cross-lin...

Key points: Cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells are required to eliminate cancer cells. We analysed the Ca2+dependence of CTL and NK cell cytotoxicity and found that in particular CTLs have a very low optimum of [Ca2+]i(between 122 and 334 nm) and [Ca2+]o(between 23 and 625 μm) for efficient cancer cell elimination, well b...

Cysteines are among the least abundant amino acids found in proteins. Due to their unique nucleophilic thiol group, they are able to undergo a broad range of chemical modifications besides their known role in disulfide formation, such as S-sulfenylation (-SOH), S-sulfinylation (-SO(2)H), S-sufonylation (-SO(3)H), S-glutathionylation (-SSG), and S-s...

Background: IDH1-mutant tumors form the majority of grade II and grade III gliomas, qualifying as a new entity in the 2016 WHO classification of CNS tumors. We have previously shown that a peptide vaccine encoding the IDH1(R132H) epitope evokes an MHC II-restricted CD4+ T Helper-1 cell (Th1) immune response and is therapeutic in an MHC-humanized A2...

Peptidergic dorsal root ganglion (DRG) neurons transmit sensory and nociceptive information from the periphery to the central nervous system. Their synaptic activity is profoundly affected by neuromodulatory peptides stored and released from large dense-core vesicles (LDCVs). However, the mechanism of peptide secretion from DRG neurons is poorly un...

Background The point mutation IDH1R132H is the most frequent one in grade II and grade III gliomas and secondary glioblastomas. We have previously shown that a peptide vaccine encoding the IDH1R132H epitope evokes an MHC II-restricted CD4+ T Helper-1 cell (Th1) immune response and is therapeutic in an MHC-humanized A2.DR1 mouse tumor model. Ongoing...

Store-operated Ca²⁺ entry mediated by STIM1-gated Orai1 channels is essential to activate immune cells and its inhibition or gain-of-function can lead to immune dysfunction and other pathologies. Reactive oxygen species interacting with cysteine residues can alter protein function. Pretreatment of the Ca²⁺ selective Orai1 with the oxidant H2O2 redu...

ORAI1 proteins are ion channel subunits and the essential pore-forming units of the calcium release-activated calcium channel complex essential for T-cell activation and many other cellular processes. In this study, we used environmental scanning electron microscopy (ESEM) with scanning transmission electron microscopy (STEM) detection to image pla...

In phagocytes, pathogen recognition is followed by Ca2+ mobilization and NADPH oxidase 2 (NOX2)-mediated "oxidative burst," which involves the rapid production of large amounts of reactive oxygen species (ROS). We showed that ORAI Ca2+ channels control store-operated Ca2+ entry, ROS production, and bacterial killing in primary human monocytes. ROS...

N-glycosylation of cell surface proteins affects protein function, stability, and interaction with other proteins. Orai channels, which mediate store-operated Ca2+ entry (SOCE), are composed of N-glycosylated subunits. Upon activation by Ca2+ sensor proteins (stromal interaction molecules STIM1 or STIM2) in the endoplasmic reticulum, Orai Ca2+ chan...

A wide variety of cellular function depends on the dynamics of intracellular Ca(2+) signals. Especially for relatively slow and lasting processes such as gene expression, cell proliferation and often migration, cells rely on the store-operated Ca(2+) entry (SOCE) pathway, which is particularly prominent in immune cells. Here, sensor proteins (STIM1...

Introduction: Antigen mediated stimulation of naïve CD4+ T cells triggers differentiation into effector regulatory or helper subtypes in a cytokine and antigen dependent manner. Imbalances between resulting subtypes lead to an inadequate immune response or to an autoimmune disease. The crucial role of Ca2+ signals in shaping the resulting immune re...

Understanding the molecular machinery underlying cellular function demands innovative microscopy techniques. Scanning transmission electron microscopy (STEM) of liquid specimens, so-called Liquid STEM, resolved the constituents of individual protein complexes within the context of intact cells in liquid state. Contrast was obtained on specific prot...

Cellular homeostasis relies upon precise regulation of Ca2+ concentration. Stromal interaction molecule (STIM) proteins regulate store-operated calcium entry (SOCE) by sensing Ca2+ concentration in the ER and forming oligomers to trigger Ca2+ entry through plasma membrane-localized Orai1 channels. Here we characterize a STIM2 splice variant, STIM2....

Orai1 subunits interacting with STIM1 molecules comprise the major components responsible for calcium release-activated calcium (CRAC) channels. The homologs Orai2 and Orai3 yield smaller store-operated currents when overexpressed and are mostly unable to substitute Orai1. Orai3 subunits are also essential components of store independent channel co...

Ca2+ signaling depends on a tight regulation of the intracellular Ca2+ concentration. Alterations in basal Ca2+ can lead to various diseases and likely contribute to development of abnormal growth. Different regulators such as calmodulin and Ca2+ pumps limit cytosolic [Ca2+] and their down-regulation by siRNA lead to an increased basal [Ca2+]. Anot...

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Cell homeostasis and Ca2+ signaling depend on a tight regulation of the intracellular Ca2+ concentration. Alterations in basal Ca2+ can lead to various diseases and likely contribute to development of abnormal growth. Different regulators such as calmodulin and Ca2+ pumps limit cytosolic [Ca2+] and their down-regulation by siRNA lead to an increase...

Reactive oxygen and nitrogen species (ROS/RNS) are often byproducts of biochemical reactions but are increasingly recognized as important second messengers involved in regulation of distinct cellular functions. Mild and reversible oxidation of certain amino acids within protein polypeptide chains is known to precisely control function of transcript...

We have tested the action of three agents microinjected into the ventral nerve photoreceptor of Limulus on the electrical response to dim light. 1. A monoclonal antibody (mAb 4A) against the G alpha subunit of frog transducin reduces the size of the receptor current to 60%, suggesting an interaction with G alpha in the Limulus photoreceptor. 2. Inj...

Spontaneous melanoma phenotype switching is controlled by unknown environmental factors and may determine melanoma outcome and responsiveness to anti-cancer therapy. We show that Orai1 and STIM2 are highly expressed and control store operated Ca(2+) entry in human melanoma. Lower extracellular Ca(2+) or silencing of Orai1/STIM2 caused a decrease in...

Plasma-membrane-localized Orai1 ion channel subunits interacting with ER-localized STIM1 molecules comprise the major subunit composition responsible for calcium release-activated calcium channels. STIM1 "translates" the Ca(2+) store content into Orai1 activity, making it a store-operated channel. Surprisingly, in addition to being the physical act...

To analyze the influence of Ca2+ microdomains on the global cytosolic Ca2+ concentration, we consider the polarization and activation of T-cells after the formation of an immunological synapse as a model system. For T-cell proliferation and activation, a high and robust Ca2+ signal lasting from minutes up to hours is needed. This raises the intrigu...

T cell receptor mediated activation requires the influx of calcium from the extracellular medium. Here, the endoplasmic reticulum (ER) Ca2+ sensor STIM1 reports the decrease in luminal [Ca2+] caused by IP3 receptor activity, clusters and directly binds and activates Orai1 channels, providing Ca2+ influx (store-operated, SOCE) essential for immune c...

A drop of endoplasmic reticulum Ca2+ concentration triggers its Ca2+ ssensor protein stromal interaction molecule 1 (STIM1) to oligomerize and accumulate within endoplasmic reticulum-plasma membrane junctions where it activates Orai1 channels, providing store-operated Ca2+ entry. To elucidate the functional significance of N-glycosylation sites of...

T-lymphocytes rely on a Ca(2+) signal known as store operated calcium entry (SOCE) for their activation. This Ca(2+) signal is generated by activation of T-cell receptor, depletion of endoplasmic reticulum (ER) Ca(2+) stores and activation of Ca(2+) release-activated Ca(2+) currents (I(CRAC)). Here we report that the ER protein ORMDL3, the product...

Abstract  Store-operated Ca(2+) entry (SOCE) is a widespread mechanism in cells to raise cytosolic Ca(2+) and to refill Ca(2+) stores. T cells critically rely on SOCE mediated by stromal interaction molecules (STIM) and Orai molecules for their activation and regulation of gene transcription; cells such as muscle cells, neurons or melanocytes proba...

A genetically encoded sensor for parallel measurements of phosphatidylinositol 3-kinase activity and hydrogen peroxide (H(2)O(2)) levels (termed PIP-SHOW) was developed. Upon elevation of local phosphatidylinositol 3,4,5-trisphosphate (PIP(3)) concentration, the sensor translocates from the cytosol to the plasma membrane, while a ratiometric excita...

UV radiation of the skin triggers keratinocytes to secrete endothelin-1 (ET-1) that binds to endothelin receptors on neighboring melanocytes. Melanocytes respond with a prolonged increase in intracellular Ca(2+) concentration ([Ca(2+)](i)), which is necessary for proliferation and melanogenesis. A major fraction of the Ca(2+) signal is caused by en...

Using biochemical and electrophysiological methods we found that pentam-idine exclusively inhibits hERG export from the endoplasmic reticulum to the cell surface in a heterologous expression system as well as in cardiomyo-cytes. hERG trafficking inhibition could be rescued in the presence of the pharmacological chaperone astemizole. We used rescue...

Store-operated CRAC channels encoded by the Orai genes mediate calcium entry in T cells. In this issue of Immunity, Omilusik et al. (2011) record Ca(V)1.4-mediated voltage-gated calcium currents in T cells and address their role for T cell development and function.

Reactive oxygen species (ROS) are increasingly recognized as second messengers in many cellular processes. While high concentrations of oxidants damage proteins, lipids and DNA, ultimately resulting in cell death, selective and reversible oxidation of key residues in proteins is a physiological mechanism that can transiently alter their activity an...

Cell polarization enables restriction of signalling into microdomains. Polarization of lymphocytes following formation of a mature immunological synapse (IS) is essential for calcium-dependent T-cell activation. Here, we analyse calcium microdomains at the IS with total internal reflection fluorescence microscopy. We find that the subplasmalemmal c...

Store-operated Ca(2+) entry is a major Ca(2+) entry mechanism that is present in most cell types. In immune cells, store-operated Ca(2+) entry is almost exclusively mediated by Ca(2+) release-activated Ca(2+) (CRAC) channels. Ca(2+) entry through these channels and the corresponding cytosolic Ca(2+) signals are required for many immune cell functio...

Reactive oxygen species (ROS) are involved in many physiological and pathophysiological cellular processes. We used lymphocytes, which are exposed to highly oxidizing environments during inflammation, to study the influence of ROS on cellular function. Calcium ion (Ca(2+)) influx through Ca(2+) release-activated Ca(2+) (CRAC) channels composed of p...

Ca(2+) homeostasis requires balanced uptake and extrusion, and dysregulation leads to disease. TRPV6 channels are homeostasis regulators, are upregulated in certain cancers, and show an unusual allele-specific evolution in humans. To understand how Ca(2+) uptake can be adapted to changes in metabolic status, we investigate regulation of Ca(2+)-infl...

The calcium selective ion channel TRPV6 shows a highly unusual evolution of one of its polymorphic alleles which might have conferred a selective advantage during migration of humans out of Africa. Because the ancestral allele contains an additional PKC consensus site, we analyzed regulation by intracellular ATP and phosphorylation. We found that A...

The ion channel TRPV6 is likely to function as an epithelial calcium channel in organs with high calcium transport requirements such as the intestine, kidney, and placenta. Transcriptional regulation of TRPV6 messenger RNA (mRNA) is controlled by 1,25-dihydroxyvitamin D, which is the active hormonal form of vitamin D3, and by additional calcium-dep...

TRPM (transient receptor potential melastatin-like) channels are distinct from many other members of the transient receptor potential family in regard to their overall size (>1000 amino acids), the lack of N-terminal ankyrin-like repeats, and hydrophobicity predictions that may allow for more than six transmembrane regions. Common to each TRPM memb...

The Ca(2+) homeostasis within cells controls a diversity of cellular processes including gene transcription, proliferation and apoptosis. Perturbance of Ca(2+) signaling may induce deregulation of cell proliferation and suppression of cell death providing the basis for cancer development. In human prostate cancer, a correlation between the mRNA exp...

In recent years many new members of the family of TRP ion channels have been identified. These channels are classified into several subgroups and participate in many sensory and physiological functions. TRPV channels are important for the perception of pain, temperature sensing, osmotic regulation, and maintenance of calcium homeostasis, and much r...

Life expectancy for patients suffering from prostate cancer is inversely correlated with the degree of extraprostatic metastasis. In order to find pharmacological tools to treat this aggressive growth it is important to define targets whose expression not only correlates with the malignancy of the cancerous cells, but that are also amenable to phar...