Multilevel Changes in Protein Dynamics upon Complex Formation of the Calcium‐Loaded S100A4 with a Nonmuscle Myosin IIA Tail Fragment

Dysregulation of Ca²⁺‐binding S100 proteins plays important role in various diseases. The asymmetric complex of Ca²⁺‐bound S100A4 with nonmuscle myosin IIA has high stability and highly increased Ca²⁺ affinity. Here we investigated the possible causes of this allosteric effect by NMR spectroscopy. Chemical shift‐based secondary‐structure analysis did not show substantial changes for the complex. Backbone dynamics revealed slow‐timescale local motions in the H1 helices of homodimeric S100A4; these were less pronounced in the complex form and might be accompanied by an increase in dimer stability. Different mobilities in the Ca²⁺‐coordinating EF‐hand sites indicate that they communicate by an allosteric mechanism operating through changes in protein dynamics; this must be responsible for the elevated Ca²⁺ affinity. These multilevel changes in protein dynamics as conformational adaptation allow S100A4 fine‐tuning of its protein–protein interactions inside the cell during Ca²⁺ signaling.     

Mutations Q93H and E97K in TPM2 Disrupt Ca-Dependent Regulation of Actin Filaments

Tropomyosin is a two-chain coiled coil protein, which together with the troponin complex controls interactions of actin with myosin in a Ca²⁺-dependent manner. In fast skeletal muscle, the contractile actin filaments are regulated by tropomyosin isoforms Tpm1.1 and Tpm2.2, which form homo- and heterodimers. Mutations in the TPM2 gene encoding isoform Tpm2.2 are linked to distal arthrogryposis and congenital myopathy—skeletal muscle diseases characterized by hyper- and hypocontractile phenotypes, respectively. In this work, in vitro functional assays were used to elucidate the molecular mechanisms of mutations Q93H and E97K in TPM2. Both mutations tended to decrease actin affinity of homo-and heterodimers in the absence and presence of troponin and Ca²⁺, although the effect of Q93H was stronger. Changes in susceptibility of tropomyosin to trypsin digestion suggested that the mutations diversified dynamics of tropomyosin homo- and heterodimers on the filament. The presence of Q93H in homo- and heterodimers strongly decreased activation of the actomyosin ATPase and reduced sensitivity of the thin filament to [Ca²⁺]. In contrast, the presence of E97K caused hyperactivation of the ATPase and increased sensitivity to [Ca²⁺]. In conclusion, the hypo- and hypercontractile phenotypes associated with mutations Q93H and E97K in Tpm2.2 are caused by defects in Ca²⁺-dependent regulation of actin–myosin interactions.     

Natural Mutations Affect Structure and Function of gC1q Domain of Otolin-1

Otolin-1 is a scaffold protein of otoliths and otoconia, calcium carbonate biominerals from the inner ear. It contains a gC1q domain responsible for trimerization and binding of Ca²⁺. Knowledge of a structure–function relationship of gC1q domain of otolin-1 is crucial for understanding the biology of balance sensing. Here, we show how natural variants alter the structure of gC1q otolin-1 and how Ca²⁺ are able to revert some effects of the mutations. We discovered that natural substitutions: R339S, R342W and R402P negatively affect the stability of apo-gC1q otolin-1, and that Q426R has a stabilizing effect. In the presence of Ca²⁺, R342W and Q426R were stabilized at higher Ca²⁺ concentrations than the wild-type form, and R402P was completely insensitive to Ca²⁺. The mutations affected the self-association of gC1q otolin-1 by inducing detrimental aggregation (R342W) or disabling the trimerization (R402P) of the protein. Our results indicate that the natural variants of gC1q otolin-1 may have a potential to cause pathological changes in otoconia and otoconial membrane, which could affect sensing of balance and increase the probability of occurrence of benign paroxysmal positional vertigo (BPPV).     

Multifaceted Roles of ALG-2 in Ca2+-Regulated Membrane Trafficking

ALG-2 (gene name: PDCD6) is a penta-EF-hand Ca²⁺-binding protein and interacts with a variety of proteins in a Ca²⁺-dependent fashion. ALG-2 recognizes different types of identified motifs in Pro-rich regions by using different hydrophobic pockets, but other unknown modes of binding are also used for non-Pro-rich proteins. Most ALG-2-interacting proteins associate directly or indirectly with the plasma membrane or organelle membranes involving the endosomal sorting complex required for transport (ESCRT) system, coat protein complex II (COPII)-dependent ER-to-Golgi vesicular transport, and signal transduction from membrane receptors to downstream players. Binding of ALG-2 to targets may induce conformational change of the proteins. The ALG-2 dimer may also function as a Ca²⁺-dependent adaptor to bridge different partners and connect the subnetwork of interacting proteins.     

Diethylstilbestrol,a Novel ANO1 Inhibitor,Exerts an Anticancer Effect on Non-Small Cell Lung Cancer via Inhibition of ANO1

Non-small cell lung cancer (NSCLC) is one of the leading causes of cancer-related mortality; thus, therapeutic targets continue to be developed. Anoctamin1 (ANO1), a novel drug target considered for the treatment of NSCLC, is a Ca²⁺-activated chloride channel (CaCC) overexpressed in various carcinomas. It plays an important role in the development of cancer; however, the role of ANO1 in NSCLC is unclear. In this study, diethylstilbestrol (DES) was identified as a selective ANO1 inhibitor using high-throughput screening. We found that DES inhibited yellow fluorescent protein (YFP) fluorescence reduction caused by ANO1 activation but did not inhibit cystic fibrosis transmembrane conductance regulator channel activity or P2Y activation-related cytosolic Ca²⁺ levels. Additionally, electrophysiological analyses showed that DES significantly reduced ANO1 channel activity, but it more potently reduced ANO1 protein levels. DES also inhibited the viability and migration of PC9 cells via the reduction in ANO1, phospho-ERK1/2, and phospho-EGFR levels. Moreover, DES induced apoptosis by increasing caspase-3 activity and PARP-1 cleavage in PC9 cells, but it did not affect the viability of hepatocytes. These results suggest that ANO1 is a crucial target in the treatment of NSCLC, and DES may be developed as a potential anti-NSCLC therapeutic agent.     

Impact of A134 and E218 Amino Acid Residues of Tropomyosin on Its Flexibility and Function

Tropomyosin (Tpm) is one of the major actin-binding proteins that play a crucial role in the regulation of muscle contraction. The flexibility of the Tpm molecule is believed to be vital for its functioning, although its role and significance are under discussion. We choose two sites of the Tpm molecule that presumably have high flexibility and stabilized them with the A134L or E218L substitutions. Applying differential scanning calorimetry (DSC), molecular dynamics (MD), co-sedimentation, trypsin digestion, and in vitro motility assay, we characterized the properties of Tpm molecules with these substitutions. The A134L mutation prevented proteolysis of Tpm molecule by trypsin, and both substitutions increased the thermal stability of Tpm and its bending stiffness estimated from MD simulation. None of these mutations affected the primary binding of Tpm to F-actin; still, both of them increased the thermal stability of the actin-Tpm complex and maximal sliding velocity of regulated thin filaments in vitro at a saturating Ca²⁺ concentration. However, the mutations differently affected the Ca²⁺ sensitivity of the sliding velocity and pulling force produced by myosin heads. The data suggest that both regions of instability are essential for correct regulation and fine-tuning of Ca²⁺-dependent interaction of myosin heads with F-actin.     

Cinobufagin Exerts Anticancer Activity in Oral Squamous Cell Carcinoma Cells through Downregulation of ANO1

Anoctamin1 (ANO1), a calcium-activated chloride channel, is frequently overexpressed in several cancers, including oral squamous cell carcinoma (OSCC). OSCC is a highly aggressive cancer and the most common oral malignancy. ANO1 has been proposed as a potential candidate for targeted anticancer therapy. In this study, we performed a cell-based screening to identify novel regulators leading to the downregulation of ANO1, and discovered cinobufagin, which downregulated ANO1 expression in oral squamous cell carcinoma CAL-27 cells. ANO1 protein levels were significantly reduced by cinobufagin in a dose-dependent manner with an IC₅₀ value of ~26 nM. Unlike previous ANO1 inhibitors, short-term (≤10 min) exposure to cinobufagin did not alter ANO1 chloride channel activity and ANO1-dependent intestinal smooth muscle contraction, whereas long-term (24 h) exposure to cinobufagin significantly reduced phosphorylation of STAT3 and mRNA expression of ANO1 in CAL-27 cells. Notably, cinobufagin inhibited cell proliferation of CAL-27 cells expressing high levels of ANO1 more potently than that of ANO1 knockout CAL-27 cells. In addition, cinobufagin significantly reduced cell migration and induced caspase-3 activation and PARP cleavage in CAL-27 cells. These results suggest that downregulation of ANO1 by cinobufagin is a potential mechanism for the anticancer effect of cinobufagin in OSCC.     

Palmitoylethanolamide Inhibits Glutamate Release in Rat Cerebrocortical Nerve Terminals

The effect of palmitoylethanolamide (PEA), an endogenous fatty acid amide displaying neuroprotective actions, on glutamate release from rat cerebrocortical nerve terminals (synaptosomes) was investigated. PEA inhibited the Ca²⁺-dependent release of glutamate, which was triggered by exposing synaptosomes to the potassium channel blocker 4-aminopyridine. This release inhibition was concentration dependent, associated with a reduction in cytosolic Ca²⁺ concentration, and not due to a change in synaptosomal membrane potential. The glutamate release-inhibiting effect of PEA was prevented by the Cav2.1 (P/Q-type) channel blocker ω-agatoxin IVA or the protein kinase A inhibitor H89, not affected by the intracellular Ca²⁺ release inhibitors dantrolene and {"type":"entrez-protein","attrs":{"text":"CGP37157","term_id":"875406365","term_text":"CGP37157"}}CGP37157, and partially antagonized by the cannabinoid CB1 receptor antagonist AM281. Based on these results, we suggest that PEA exerts its presynaptic inhibition, likely through a reduction in the Ca²⁺ influx mediated by Cav2.1 (P/Q-type) channels, thereby inhibiting the release of glutamate from rat cortical nerve terminals. This release inhibition might be linked to the activation of presynaptic cannabinoid CB1 receptors and the suppression of the protein kinase A pathway.     

P2X4 Receptors Mediate Ca2+ Release from Lysosomes in Response to Stimulation of P2X7 and H1 Histamine Receptors

The P2X4 purinergic receptor is targeted to endolysosomes, where it mediates an inward current dependent on luminal ATP and pH. Activation of P2X4 receptors was previously shown to trigger lysosome fusion, but the regulation of P2X4 receptors and their role in lysosomal Ca²⁺ signaling are poorly understood. We show that lysosomal P2X4 receptors are activated downstream of plasma membrane P2X7 and H₁ histamine receptor stimulation. When P2X4 receptors are expressed, the increase in near-lysosome cytosolic [Ca²⁺] is exaggerated, as detected with a low-affinity targeted Ca²⁺ sensor. P2X4-dependent changes in lysosome properties were triggered downstream of P2X7 receptor activation, including an enlargement of lysosomes indicative of homotypic fusion and a redistribution of lysosomes towards the periphery of the cell. Lysosomal P2X4 receptors, therefore, have a role in regulating lysosomal Ca²⁺ release and the regulation of lysosomal membrane trafficking.     

Structural Insights into Retinal Guanylate Cyclase Activator Proteins (GCAPs)

Retinal guanylate cyclases (RetGCs) promote the Ca²⁺-dependent synthesis of cGMP that coordinates the recovery phase of visual phototransduction in retinal rods and cones. The Ca²⁺-sensitive activation of RetGCs is controlled by a family of photoreceptor Ca²⁺ binding proteins known as guanylate cyclase activator proteins (GCAPs). The Mg²⁺-bound/Ca²⁺-free GCAPs bind to RetGCs and activate cGMP synthesis (cyclase activity) at low cytosolic Ca²⁺ levels in light-activated photoreceptors. By contrast, Ca²⁺-bound GCAPs bind to RetGCs and inactivate cyclase activity at high cytosolic Ca²⁺ levels found in dark-adapted photoreceptors. Mutations in both RetGCs and GCAPs that disrupt the Ca²⁺-dependent cyclase activity are genetically linked to various retinal diseases known as cone-rod dystrophies. In this review, I will provide an overview of the known atomic-level structures of various GCAP proteins to understand how protein dimerization and Ca²⁺-dependent conformational changes in GCAPs control the cyclase activity of RetGCs. This review will also summarize recent structural studies on a GCAP homolog from zebrafish (GCAP5) that binds to Fe²⁺ and may serve as a Fe²⁺ sensor in photoreceptors. The GCAP structures reveal an exposed hydrophobic surface that controls both GCAP1 dimerization and RetGC binding. This exposed site could be targeted by therapeutics designed to inhibit the GCAP1 disease mutants, which may serve to mitigate the onset of retinal cone-rod dystrophies.     

NAD(H) Regulates the Permeability Transition Pore in Mitochondria through an External Site

The opening of the permeability transition pore (mPTP) in mitochondria initiates cell death in numerous diseases. The regulation of mPTP by NAD(H) in the mitochondrial matrix is well established; however, the role of extramitochondrial (cytosolic) NAD(H) is still unclear. We studied the effect of added NADH and NAD⁺ on: (1) the Ca²⁺-retention capacity (CRC) of isolated rat liver, heart, and brain mitochondria; (2) the Ca²⁺-dependent mitochondrial swelling in media whose particles can (KCl) or cannot (sucrose) be extruded from the matrix by mitochondrial carriers; (3) the Ca²⁺-dependent mitochondrial depolarization and the release of entrapped calcein from mitochondria of permeabilized hepatocytes; and (4) the Ca²⁺-dependent mitochondrial depolarization and subsequent repolarization. NADH and NAD⁺ increased the CRC of liver, heart, and brain mitochondria 1.5–2.5 times, insignificantly affecting the rate of Ca²⁺-uptake and the free Ca²⁺ concentration in the medium. NAD(H) suppressed the Ca²⁺-dependent mitochondrial swelling both in KCl- and sucrose-based media but did not induce the contraction and repolarization of swollen mitochondria. By contrast, EGTA caused mitochondrial repolarization in both media and the contraction in KCl-based medium only. NAD(H) delayed the Ca²⁺-dependent depolarization and the release of calcein from individual mitochondria in hepatocytes. These data unambiguously demonstrate the existence of an external NAD(H)-dependent site of mPTP regulation.     

Homology Modeling Study of Bovine μ-Calpain Inhibitor-Binding Domains

The activated mammalian CAPN-structures, the CAPN/CAST complex in particular, have become an invaluable target model using the structure-based virtual screening of drug candidates from the discovery phase to development for over-activated CAPN linked to several diseases, such as post-ischemic injury and cataract formation. The effect of Ca²⁺-binding to the enzyme is thought to include activation, as well as the dissociation, aggregation, and autolysis of small regular subunits. Unfortunately, the Ca²⁺-activated enzyme tends to aggregate when provided as a divalent ion at the high-concentration required for the protease crystallization. This is also makes it very difficult to crystallize the whole-length enzyme itself, as well as the enzyme-inhibitor complex. Several parameters that influence CAPN activity have been investigated to determine its roles in Ca²⁺-modulation, autoproteolysis, phosphorylation, and intracellular distribution and inhibition by its endogenous inhibitor CAST. CAST binds and inhibits CAPN via its CAPN-inhibitor domains (four repeating domains 1–4; CAST1–4) when CAPN is activated by Ca²⁺-binding. An important key to understanding CAPN1 inhibition by CAST is to determine how CAST interacts at the molecular level with CAPN1 to inhibit its protease activity. In this study, a 3D structure model of a CAPN1 bound bovine CAST4 complex was built by comparative modeling based on the only known template structure of a rat CAPN2/CAST4 complex. The complex model suggests certain residues of bovine CAST4, notably, the TIPPKYQ motif sequence, and the structural elements of these residues, which are important for CAPN1 inhibition. In particular, as CAST4 docks near the flexible active site of CAPN1, conformational changes at the interaction site after binding could be directly related to CAST4 inhibitory activity. These functional interfaces can serve as a guide to the site-mutagenesis in research on bovine CAPN1 structure-function relationships for the design of small molecules inhibitors to prevent uncontrolled and unspecific degradation in the proteolysis of key protease substrates.     

Functional and Molecular Properties of DYT-SGCE Myoclonus-Dystonia Patient-Derived Striatal Medium Spiny Neurons

Myoclonus-dystonia (DYT-SGCE, formerly DYT11) is characterized by alcohol-sensitive, myoclonic-like appearance of fast dystonic movements. It is caused by mutations in the SGCE gene encoding ε-sarcoglycan leading to a dysfunction of this transmembrane protein, alterations in the cerebello-thalamic pathway and impaired striatal plasticity. To elucidate underlying pathogenic mechanisms, we investigated induced pluripotent stem cell (iPSC)-derived striatal medium spiny neurons (MSNs) from two myoclonus-dystonia patients carrying a heterozygous mutation in the SGCE gene (c.298T>G and c.304C>T with protein changes W100G and R102X) in comparison to two matched healthy control lines. Calcium imaging showed significantly elevated basal intracellular Ca²⁺ content and lower frequency of spontaneous Ca²⁺ signals in SGCE MSNs. Blocking of voltage-gated Ca²⁺ channels by verapamil was less efficient in suppressing KCl-induced Ca²⁺ peaks of SGCE MSNs. Ca²⁺ amplitudes upon glycine and acetylcholine applications were increased in SGCE MSNs, but not after GABA or glutamate applications. Expression of voltage-gated Ca²⁺ channels and most ionotropic receptor subunits was not altered. SGCE MSNs showed significantly reduced GABAergic synaptic density. Whole-cell patch-clamp recordings displayed elevated amplitudes of miniature postsynaptic currents and action potentials in SGCE MSNs. Our data contribute to a better understanding of the pathophysiology and the development of novel therapeutic strategies for myoclonus-dystonia.     

Molecular Mechanisms of the Deregulation of Muscle Contraction Induced by the R90P Mutation in Tpm3.12 and the Weakening of This Effect by BDM and W7

Point mutations in the genes encoding the skeletal muscle isoforms of tropomyosin can cause a range of muscle diseases. The amino acid substitution of Arg for Pro residue in the 90th position (R90P) in γ-tropomyosin (Tpm3.12) is associated with congenital fiber type disproportion and muscle weakness. The molecular mechanisms underlying muscle dysfunction in this disease remain unclear. Here, we observed that this mutation causes an abnormally high Ca²⁺-sensitivity of myofilaments in vitro and in muscle fibers. To determine the critical conformational changes that myosin, actin, and tropomyosin undergo during the ATPase cycle and the alterations in these changes caused by R90P replacement in Tpm3.12, we used polarized fluorimetry. It was shown that the R90P mutation inhibits the ability of tropomyosin to shift towards the outer domains of actin, which is accompanied by the almost complete depression of troponin’s ability to switch actin monomers off and to reduce the amount of the myosin heads weakly bound to F-actin at a low Ca²⁺. These changes in the behavior of tropomyosin and the troponin–tropomyosin complex, as well as in the balance of strongly and weakly bound myosin heads in the ATPase cycle may underlie the occurrence of both abnormally high Ca²⁺-sensitivity and muscle weakness. BDM, an inhibitor of myosin ATPase activity, and W7, a troponin C antagonist, restore the ability of tropomyosin for Ca²⁺-dependent movement and the ability of the troponin–tropomyosin complex to switch actin monomers off, demonstrating a weakening of the damaging effect of the R90P mutation on muscle contractility.     

Synthesis and luminescence properties of single‐component Ca5(PO4)3F:Dy3+, Eu3+ white‐emitting phosphors

A series of Ca₅(PO₄)₃F:Dy³⁺, Eu³⁺ phosphors was synthesized by a solid‐state reaction method. The XRD results show that all as‐prepared Ca₅(PO₄)₃F:Dy³⁺, Eu³⁺ samples match well with the standard Ca₅(PO₄)₃F structure and the doped Dy³⁺ and Eu³⁺ ions have no effect on the crystal structure. Under near‐ultraviolet excitation, Dy³⁺ doped Ca₅(PO₄)₃F phosphor shows blue (486 nm) and yellow (579 nm) emissions, which correspond to ⁴F_9/2→⁶H_15/2 and ⁴F_9/2→⁶H_13/2 transitions respectively. Eu³⁺ co‐doped Ca₅(PO₄)₃F:Dy³⁺ phosphor shows the additional red emission of Eu³⁺ at 631 nm, and an improved color rendering index. The chromaticity coordinates of Ca₅(PO₄)₃F:Dy³⁺, Eu³⁺ phosphors also indicate the excellent warm white emission characteristics and low correlated color temperature. Overall, these results suggest that the Ca₅(PO₄)₃F:Dy³⁺, Eu³⁺ phosphors have potential applications in warm white light‐emitting diodes as single‐component phosphor.     

Protease and Amylase Stability in the Presence of Chelators Used in Laundry Detergent Applications: Correlation Between Chelator Properties and Enzyme Stability in Liquid Detergents

Chelators are a common ingredient in most laundry detergents. They have a number of different functions such as reducing water hardness, assisting in keeping particulate soil in suspension and the removal of certain stains, thus complementing the action of the anionic surfactants. Another important group of components in a modern liquid detergent is enzymes, mainly proteases and amylases. As the most commonly used enzymes within the detergent industry are dependent on bound calcium ions to maintain conformational stability and function, the presence of both chelators and enzymes in a liquid detergent presents a challenge. The three commonly used Ca²⁺ chelators: citrate, DTPA (diethylene triamine pentaacetic acid) and HEDP (1-hydroxyethane-1,1-diyl)bis(phosphonic acid), were studied with regard to their impact on protease and amylase stability in buffer and in a model liquid detergent. Enzyme stability was characterized by differential scanning calorimetry (DSC) and activity studies, and correlated to the chelator-Ca²⁺ interaction properties. The results show that a chelator’s ability to reduce water hardness and its Ca²⁺ affinity are in reality two separate aspects in the context of their use in liquid detergents. In the presence of DTPA, stoichiometric surplus of free Ca²⁺ is required to maintain sufficient amylase and protease stability. In the presence of the weaker chelators, HEDP and citrate, the total Ca²⁺ concentration is more important to protein stability than stoichiometric balancing between chelator and Ca²⁺. Thus, for these chelators their total concentration only has a minor impact on the Ca²⁺ concentration required to maintain or improve enzyme storage stability. The results underline the importance of Ca²⁺ in liquid detergent formulations, and suggest how proper balancing of chelators and Ca²⁺ can be used to improve overall enzyme stability.     

Genetic Dissection of Vps13 Regulation in Yeast Using Disease Mutations from Human Orthologs

The VPS13 family of proteins have emerged as key players in intracellular lipid transport and human health. Humans have four different VPS13 orthologs, the dysfunction of which leads to different diseases. Yeast has a single VPS13 gene, which encodes a protein that localizes to multiple different membrane contact sites. The yeast vps13Δ mutant is pleiotropic, exhibiting defects in sporulation, protein trafficking, endoplasmic reticulum (ER)-phagy and mitochondrial function. Non-null alleles resulting from missense mutations can be useful reagents for understanding the multiple functions of a gene. The exceptionally large size of Vps13 makes the identification of key residues challenging. As a means to identify critical residues in yeast Vps13, amino acid substitution mutations from VPS13A, B, C and D, associated with human disease, were introduced at the cognate positions of yeast VPS13, some of which created separation-of-function alleles. Phenotypic analyses of these mutants have revealed that the promotion of ER-phagy is a fourth, genetically separable role of VPS13 and provide evidence that co-adaptors at the endosome mediate the activity of VPS13 in vacuolar sorting.     

Interferon Beta Activity Is Modulated via Binding of Specific S100 Proteins

Interferon-β (IFN-β) is a pleiotropic cytokine used for therapy of multiple sclerosis, which is also effective in suppression of viral and bacterial infections and cancer. Recently, we reported a highly specific interaction between IFN-β and S100P lowering IFN-β cytotoxicity to cancer cells (Int J Biol Macromol. 2020; 143: 633–639). S100P is a member of large family of multifunctional Ca²⁺-binding proteins with cytokine-like activities. To probe selectivity of IFN-β—S100 interaction with respect to S100 proteins, we used surface plasmon resonance spectroscopy, chemical crosslinking, and crystal violet assay. Among the thirteen S100 proteins studied S100A1, S100A4, and S100A6 proteins exhibit strictly Ca²⁺-dependent binding to IFN-β with equilibrium dissociation constants, K_d, of 0.04–1.5 µM for their Ca²⁺-bound homodimeric forms. Calcium depletion abolishes the S100—IFN-β interactions. Monomerization of S100A1/A4/A6 decreases K_d values down to 0.11–1.0 nM. Interferon-α is unable of binding to the S100 proteins studied. S100A1/A4 proteins inhibit IFN-β-induced suppression of MCF-7 cells viability. The revealed direct influence of specific S100 proteins on IFN-β activity uncovers a novel regulatory role of particular S100 proteins, and opens up novel approaches to enhancement of therapeutic efficacy of IFN-β.