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Importance: Approximately 125 million people worldwide have psoriasis. Patients with psoriasis experience substantial morbidity and increased rates of inflammatory arthritis, cardiometabolic diseases, and mental health disorders.Observations: Plaque psoriasis is the most common variant of psoriasis. The most rapid advancements addressing plaque psoriasis have been in its pathogenesis, genetics, comorbidities, and biologic treatments. Plaque psoriasis is associated with a number of comorbidities including psoriatic arthritis, cardiometabolic diseases, and depression. For patients with mild psoriasis, topical agents remain the mainstay of treatment, and they include topical corticosteroids, vitamin D analogues, calcineurin inhibitors, and keratolytics. The American Academy of Dermatology-National Psoriasis Foundation guidelines recommend biologics as an option for first-line treatment of moderate to severe plaque psoriasis because of their efficacy in treating it and acceptable safety profiles. Specifically, inhibitors to tumor necrosis factor α (TNF-α) include etanercept, adalimumab, certolizumab, and infliximab. Other biologics inhibit cytokines such as the p40 subunit of the cytokines IL-12 and IL-13 (ustekinumab), IL-17 (secukinumab, ixekizumab, bimekizumab, and brodalumab), and the p19 subunit of IL-23 (guselkumab, tildrakizumab, risankizumab, and mirikizumab). Biologics that inhibit TNF-α, p40IL-12/23, and IL-17 are also approved for the treatment of psoriatic arthritis. Oral treatments include traditional agents such as methotrexate, acitretin, cyclosporine, and the advanced small molecule apremilast, which is a phosphodiesterase 4 inhibitor. The most commonly prescribed light therapy used to treat plaque psoriasis is narrowband UV-B phototherapy.Conclusions and Relevance: Psoriasis is an inflammatory skin disease that is associated with multiple comorbidities and substantially diminishes patients' quality of life. Topical therapies remain the cornerstone for treating mild psoriasis. Therapeutic advancements for moderate to severe plaque psoriasis include biologics that inhibit TNF-α, p40IL-12/23, IL-17, and p19IL-23, as well as an oral phosphodiesterase 4 inhibitor. [ABSTRACT FROM AUTHOR]

Background: Recombinant human calcineurin B subunit (rhCNB) has been shown to be an immune‐stimulatory protein promoting cytokine production and inducing phenotypic maturation of Dendritic cells (DCs). In vivo, it has good antitumor efficacy, and has potential as an antitumor drug. Exogenous rhCNB was found to be internalized into tumor cells via the Toll‐like receptor 4 (TLR4) complex, but it was not known whether its immuno‐modulatory and antitumor functions involved entry by this same route. Methods: The production and secretion of the cytokines and chemokines in innate immune cells induced by rhCNB were determined by ELISA, and the expression of CD40, CD80, CD86, and MHCII was analyzed by FACs. Experimental Lewis lung cancer (LLC) model was prepared in C57 BL/6 wild‐type (WT) mice, TLR4−/− mice or their littermates by the inoculation of LLCs in their right armpit, and then administrated daily intraperitoneal injections (0.2 mL) of normal saline, rhCNB 20 mg/kg, and rhCNB 40 mg/kg, respectively. Results: Recombinant human calcineurin B subunit promoted the production of antitumor cytokines by innate immune cells, and culture supernatants of rhCNB‐stimulated immune cells induced apoptosis of LLCs. In addition, rhCNB up‐regulated CD40, CD80, CD86, and MHCII expression in macrophages and DCs in TLR4+ cells but failed to do so in TLR4 deficient cells. rhCNB also induced the formation of CD4+ and CD8+T cells in splenocytes from WT mice, but not from TLR4‐deficient littermates. Intraperitoneal administration of WT C57BL/6 mice with rhCNB resulted in a 50% reduction in LLC tumor growth, but failed to inhibit tumor growth in TLR4−/− littermates. Conclusions: The in vivo antitumor and immunomodulatory effects of rhCNB are mediated by the TLR4. This conclusion is important for the further understanding and development of rhCNB as an antitumor drug. [ABSTRACT FROM AUTHOR]

Aim: Serum uromodulin (sURO) was recently found as a sensitive tubular marker in early chronic kidney disease stages. Thus far, mainly early uromodulin urinary excretion was tested in kidney recipients. The aim of our study was to conduct a long-lastinlong-term assessment of sURO in kidney graft function monitoring. Material/Methods: Forty-one stable kidney recipients (aged 47 (13.7)) were studied around the 3rd month (3m) and the 2nd year (2y) after kidney transplantation. Sera were tested for sURO, creatinine and tacrolimus levels. Kidney biopsy was scored according to revised Banff 97 classification. Results: sURO level (mean 66.06ng/ml at 3m; 77.81 at 2y) increased borderline significantly (P = 0.051) in time and significantly correlated with eGFR (3m RS = 0.46; 2y RS = 0.58), creatinine levels (RS respectively -0.55 and -0.56) and donor age (3m Rs = -0.33; 2y RS = -0.41). We observed borderline correlations between sURO and Banff biopsy scoring: 3m-sURO with arteriolar hyalinosis-ah (RS = -0.3, P = 0.06) and 2y-sURO with peritubular capillaritis-ptc (RS = 0.45, P = 0.07). Correlations of sURO with 3m tacrolimus levels (Rs = 0.3, P = 0.08) were borderline, however patients with CNI toxicity lesions in biopsy had sURO significantly lower (mean 3m-sURO 52.7 vs 83.1 ng/ml; 2y-sURO 61.9 vs 98.1 ng/ml). Conclusions: sURO can reflect kidney graft quality and function. sURO correlated with ptc, which is considered to be an early marker of a chronic antibody-mediated graft injury. Tacrolimus doesn't influence sURO levels directly, but sURO is lower in patients with toxic kidney injury in biopsy. sURO can reflect kidney graft quality and function. sURO correlated with ptc, which is considered to be an early marker of a chronic antibody-mediated graft injury. Tacrolimus doesn't influence sURO levels directly, but sURO is lower in patients with toxic kidney injury in biopsy. [ABSTRACT FROM AUTHOR]

Abstract Clear cell renal cell carcinoma (ccRCC) is one of the most common malignant tumors in the urinary system, and its incidence continues to increase. Regulator of calcineurin 1 (RCAN1), one of the genes on chromosome 21, is a crucial mediator of tumor inhibition. RCAN1.4 is best characterized as an endogenous inhibitor of the phosphatase calcineurin, and it has been observed to be downregulated in numerous types of cancer. However, its essential function remains unclear in ccRCC. In the present study, we found that RCAN1.4 expression was frequently downregulated in renal cell carcinoma tissues and cells and was inversely correlated with various clinicopathological parameters. Low RCAN1.4 expression was associated with poor overall survival and disease-free survival and could act as a diagnostic indicator in ccRCC patients. Furthermore, the overexpression of RCAN1.4 inhibited cell proliferation, migration and invasion, whereas RCAN1.4 knockdown promoted these functions in ccRCC cell lines. In addition, RCAN1.4 expression was downregulated in sunitinib-resistant renal cancer cell lines, and inhibition of RCAN1.4 promoted sunitinib resistance. We also found that RCAN1.4 could regulate epithelial-mesenchymal transition (EMT) and the expression of HIF2α in sunitinib-resistant cell lines. Taken together, these findings indicate that downregulation of RCAN1.4 may be crucial for the metastasis of ccRCC and may induce sunitinib resistance. RCAN1.4 may act as a prognostic indicator and potential therapeutic target for ccRCC. [ABSTRACT FROM AUTHOR]

Endothelial cell activation by proinflammatory stimuli drives leukocyte recruitment through enhanced expression of counter-receptors such as vascular cell adhesion molecule-1 (VCAM-1). We previously demonstrated that activation of the receptor tyrosine kinase EphA2 with its ligand ephrin-A1 induces VCAM-1 expression. Here, we sought to characterize the proinflammatory signaling pathways involved. Analysis of over-represented transcription factors in ephrin-A1-induced genes identified multiple potential transcriptional regulators, including the Rel family members nuclear factor-κB (NF-κB/p65) and nuclear factor of activated T-cells (NFAT). While ephrin-A1 failed to induce endothelial NF-κB activation, NF-κB inhibitors prevented ephrin-A1-induced VCAM-1 expression, suggesting basal NF-κB activity is required. In contrast, ephrin-A1 induced a robust EphA2-dependent increase in NFAT activation, and mutation of the NF-κB/NFAT-binding sites in the VCAM-1 promoter blunted ephrin-A1-induced promoter activity. NFAT activation classically occurs through calcium-dependent calcineurin activation, and inhibiting NFAT signaling with calcineurin inhibitors (cyclosporine A, FK506) or direct NFAT inhibitors (A-285222) was sufficient to block ephrin-A1-induced VCAM-1 expression. Consistent with robust NFAT activation, ephrin-A1-induced an EphA2-dependent calcium influx in endothelial cells that was required for ephrin—A1-induced NFAT activation and VCAM-1 expression. This work provides the first data showing EphA2-dependent calcium influx and NFAT activation and identifies NFAT as a novel EphA2-dependent proinflammatory pathway in endothelial activation. [ABSTRACT FROM AUTHOR]

Cyclooxygenase-2 (COX-2) derived-prostanoids participate in the altered vascular function and mechanical properties in cardiovascular diseases. We investigated whether regulator of calcineurin 1 (Rcan1) participates in vascular contractility and stiffness through the regulation of COX-2. For this, wild type ( Rcan1 +/+ ) and Rcan1 -deficient ( Rcan1 −/− ) mice untreated or treated with the COX-2 inhibitor rofecoxib were used. Vascular function and structure were analysed by myography. COX-2 and phospo-p65 expression were studied by western blotting and immunohistochemistry and TXA 2 production by ELISA. We found that Rcan1 deficiency increases COX-2 and IL-6 expression and NF-κB activation in arteries and vascular smooth muscle cells (VSMC). Adenoviral-mediated re-expression of Rcan1.4 in Rcan1 −/− VSMC normalized COX-2 expression. Phenylephrine-induced vasoconstrictor responses were greater in aorta from Rcan1 −/− compared to Rcan1 +/+ mice. This increased response were diminished by etoricoxib, furegrelate, SQ 29548, cyclosporine A and parthenolide, inhibitors of COX-2, TXA 2 synthase, TP receptors, calcineurin and NF-κB, respectively. Endothelial removal and NOS inhibition increased phenylephrine responses only in Rcan1 +/+ mice. TXA 2 levels were greater in Rcan1 −/− mice. In small mesenteric arteries, vascular function and structure were similar in both groups of mice; however, vessels from Rcan1 −/− mice displayed an increase in vascular stiffness that was diminished by rofecoxib. In conclusion, our results suggest that Rcan1 might act as endogenous negative modulator of COX-2 expression and activity by inhibiting calcineurin and NF-kB pathways to maintain normal contractility and vascular stiffness in aorta and small mesenteric arteries, respectively. Our results uncover a new role for Rcan1 in vascular contractility and mechanical properties. [ABSTRACT FROM AUTHOR]

ABSTRACT: Aim: Arteriolar hyalinosis (AH) is a common lesion in allograft biopsies taken following kidney transplantation. Recent studies have shown that severe AH may predict transplant outcomes and provide information about previous exposure to certain drugs, such as calcineurin inhibitors (CNI). However, the incidence of AH as a direct result of diabetic nephropathy (DN) after kidney transplantation has not been fully evaluated. This study aimed to assess the impact of primary DN on the development of AH lesions in patients who underwent kidney transplantation. Methods: Eighty‐three patients who underwent living‐donor kidney transplantation between April 2005 and June 2015 were enrolled in this study. A total of 33 patients had DN prior to transplantation. Allograft biopsies were scored according to the Banff classification, and the relationship between the individual histological lesions and clinical baseline data was assessed. Results: At early biopsy (3–12 months), there were no differences in the rates of AH lesions between the DN group and the non‐DN group (ah ≥ 1: 37% vs. 41.3%, P = 0.719; aah ≥ 1: 14.8% vs. 6.5%; P = 0.453). However, there were significant differences between the groups in biopsies taken more than 3 years after the transplant (ah ≥ 2: 83.3% vs. 36.8%, P = 0.013; aah ≥ 2: 66.7% vs. 21.1%, P = 0.011). Multivariable analysis showed that both the length of time after transplantation and the presence of DN were independent risk factors for ah ≥ 2 (odds ratio [OR]: 2.55, 95% confidence interval [CI]: 1.47–19.54, P = 0.011) and aah ≥ 2 (OR: 7.55, 95% CI: 1.49–38.33, P = 0.015). Conclusion: This is the first report showing that the presence of primary DN disease contributes to the development of severe AH late in the course after kidney allografts. [ABSTRACT FROM AUTHOR]

Almost every cell in the human body extends a primary cilium. Defective cilia function leads to a set of disorders known as ciliopathies, which are characterised by debilitating developmental defects that affect many tissues. Here, we report a new role for regulator of calcineurin 2 (RCAN2) in primary cilia function. It localises to centrioles and the basal body and is required to maintain normal cilia length. RCAN2 was identified as the most strongly upregulated gene from a comparative RNAseq analysis of cells in which expression of the Golgi matrix protein giantin had been abolished by gene editing. In contrast to previous work where we showed that depletion of giantin by RNAi results in defects in ciliogenesis and in cilia length control, giantin knockout cells generate normal cilia after serum withdrawal. Furthermore, giantin knockout zebrafish show increased expression of RCAN2. Importantly, suppression of RCAN2 expression in giantin knockout cells results in the same defects in the control of cilia length that are seen upon RNAi of giantin itself. Together, these data defineRCAN2 as a regulator of cilia function that can compensate for the loss of giantin function. [ABSTRACT FROM AUTHOR]

Wear particles serve a central role in periprosthetic osteolysis, which leads to the aseptic loosening of prostheses. In the present study a lentiviral vector was constructed to silence macrophage colony stimulating factor (M-CSF) and receptor activator of nuclear factor κ-B ligand (RANKL) genes, which synergistically inhibit osteoclast formation and differentiation. To confirm the role of the calcineurin/nuclear factor of activated T cells (NFAT) pathway in osteolysis, we transduced murine macrophage/monocyte RAW264.7 cells with M-CSF-short hairpin (sh)RNA-RANKL-shRNA. Tumor necrosis factor-α (TNF-α) protein levels were evaluated using enzyme-linked immunosorbent assay. Transduced RAW264.7 cells were cultured in Transwell chambers in the presence of 0.1 mg/ml titanium particles to investigate the capacity of TNF-α inhibition to reduce wear debris-induced inflammation. RANKL, M-CSF, TNF-α, interleukin (IL)-1β, IL-6 and NFATc1 mRNA levels were also assessed by reverse transcription-quantitative polymerase chain reaction. Osteoclastogenesis was measured by tartrate-resistant acid phosphatase (TRAP) mRNA quantification. Lentiviral-mediated double gene inhibition is known to be able to completely inhibit inflammatory osteolysis, simultaneously decreasing the number of NFATc1- and TRAP-positive cells. The present study confirmed that the combined silencing of M-CSF and RANKL genes can inhibit the osteolysis induced by the wear particles around the prosthesis. The calcineurin/NFAT pathway serves a role in the prevention of prosthesis loosening. [ABSTRACT FROM AUTHOR]

The expression of transient receptor potential canonical 6 (TRPC6) in central nervous system (CNS) is important for neuronal functions and certain neural disorders. However, the regulatory mechanism of TRPC6 expression in neurons is still obscure. In this study, we show that TRPC6 expression in the primary cultured cortical neurons is bidirectionally regulated by glutamate. Activation of NR2A-containing NMDARs induces TRPC6 transcription through a calcineurin-dependent pathway. In contrast, activation of NR2B-containing NMDARs causes TRPC6 degradation through calpain. Thus, TRPC6 expression in neurons is regulated by glutamate in a bidirectional manner that is dependent on NR2A and NR2B. [ABSTRACT FROM AUTHOR]

FK506 (Tacrolimus), isolated from Streptomyces tsukubaenis is a powerful immunosuppressant shown to inhibit T cell activation. FK506 mediated immunosuppression requires the formation of a complex between FK506, a FK506 binding protein (FKBP) and calcineurin. Numerous FKBPs have been identified in a wide range of species, from single celled organisms to humans. FKBPs show peptidylprolyl cis/trans isomerase (PPIase) activity and have been shown to affect a wide range of cellular processes including protein folding, receptor signaling and apoptosis. FKBPs also affect numerous biological functions in addition to immunosuppression including regulation of cardiac function, neuronal function and development and have been implicated in several diseases including cardiac disease, cancer and neurodegenerative diseases such as Alzheimer's disease. More recently, FKBPs have proven useful as molecular tools for studying protein interactions, localization and functions. This review provides an overview of the current state of knowledge of FKBPs and their numerous biological functions and uses. [ABSTRACT FROM AUTHOR]

Cardiovascular incidences are markedly higher in men than in pre-menstrual women. However, this advantage in women declines with aging and therefore can be correlated with the sex hormone 17β-Estradiol (E2) which is reported to protect heart cells by acting though estrogen receptors (ERs). In this study we have determined the effect of E2/ERβ against ISO induced cellular hypertrophy in H9c2 cardiomyoblast cells. The results confirm that ISO induced cardiac-hypertrophy by elevating the levels of hypertrophy associated proteins, ANP and BNP and further by upregulating p-CaMKII, calcineurin, p-GATA4 and NFATc3 which was correlated with a significant enlargement of the H9c2 cardiomyoblast. However, overexpression of ERβ and/or administration of E2 inhibited ISO-induced hypertrophy in H9c2 cells. In addition, E2/ERβ also inhibited ISO-induced NFATc3 translocation, and reduced the protein level of downstream marker, BNP. Furthermore, by testing with the calcineurin inhibitor (CsA), it was confirmed that calcineurin acted as a key mediator for the anti-hypertrophic effect of E2/ERβ. In cells treated with calcium blocker (BATPA), the inhibitory effect of E2/ERβ on ISO-induced Ca2+ influx and hypertrophic effects were totally blocked suggesting that E2/ERβ inhibited calcineurin activity to activate I-1 protein and suppress PP1, then induce PLB protein phosphorylation and activation, resulting in Ca2+ reuptake into sarcoplasmic reticulum through SR Ca2+ cycling modification. In conclusion, E2/ERβ suppresses the Ca2+ influx and calcineurin activity induced by ISO to enhance the PLB protein activity and SR Ca2+ cycling. [ABSTRACT FROM AUTHOR]

Dysfunction of the fast-inactivating Kv3.4 potassium current in dorsal root ganglion (DRG) neurons contributes to the hyperexcitability associated with persistent pain induced by spinal cord injury (SCI). However, the underlying mechanism is not known. In light of our previous work demonstrating modulation of the Kv3.4 channel by phosphorylation, we investigated the role of the phosphatase calcineurin (CaN) using electrophysiological, molecular, and imaging approaches in adult female Sprague Dawley rats. Pharmacological inhibition of CaN in small-diameter DRG neurons slowed repolarization of the somatic action potential (AP) and attenuated the Kv3.4 current. Attenuated Kv3.4 currents also exhibited slowed inactivation. We observed similar effects on the recombinant Kv3.4 channel heterologously expressed in Chinese hamster ovary cells, supporting our findings inDRGneurons. Elucidating the molecular basis of these effects, mutation of four previously characterized serines within the Kv3.4 N-terminal inactivation domain eliminated the effects of CaN inhibition on the Kv3.4 current. SCI similarly induced concurrent Kv3.4 current attenuation and slowing of inactivation. Although there was little change in CaN expression and localization after injury, SCI induced upregulation of the native regulator of CaN 1 (RCAN1) in the DRG at the transcript and protein levels. Consistent with CaN inhibition resulting from RCAN1 upregulation, overexpression of RCAN1 in naive DRG neurons recapitulated the effects of pharmacological CaN inhibition on the Kv3.4 current and the AP. Overall, these results demonstrate a novel regulatory pathway that links CaN, RCAN1, and Kv3.4 inDRGneurons. Dysregulation of this pathway might underlie a peripheral mechanism of pain sensitization induced by SCI. [ABSTRACT FROM AUTHOR]

Regulator of calcineurin 1 (RCAN1) is an endogenous inhibitor of the calcineurin pathway in cells. It is expressed as two isoforms in vertebrates: RCAN1.1 is constitutively expressed in most tissues, whereas transcription of RCAN1.4 is induced by several stimuli that activate the calcineurin-NFAT pathway. RCAN1.4 is highly upregulated in response to VEGF in human endothelial cells in contrast to RCAN1.1 and is essential for efficient endothelial cell migration and tubular morphogenesis. Here, we show that RCAN1.4 has a role in the regulation of agonist-stimulated VEGFR-2 internalisation and establishment of endothelial cell polarity. siRNA-mediated gene silencing revealed that RCAN1 plays a vital role in regulating VEGF-mediated cytoskeletal reorganisation and directed cell migration and sprouting angiogenesis. Adenoviral-mediated overexpression of RCAN1.4 resulted in increased endothelial cell migration. Antisense-mediated morpholino silencing of the zebrafish RCAN1.4 orthologue revealed a disrupted vascular development further confirming a role for the RCAN1.4 isoform in regulating vascular endothelial cell physiology. Our data suggest that RCAN1.4 plays a novel role in regulating endothelial cell migration by establishing endothelial cell polarity in response to VEGF. [ABSTRACT FROM AUTHOR]

Ca2+ signals are transient, hence, upon a stimulus-induced increase in cytosolic Ca2+ concentration, cells have to reestablish resting Ca2+ levels. Ca2+ extrusion is operated by a wealth of transporters, such as Ca2+ pumps and Ca2+/H+ antiporters, which often require a rise in Ca2+ concentration to be activated. Here, we report a regulatory fine-tuning mechanism of the Arabidopsis thaliana plasma membrane-localized Ca2+-ATPase isoform ACA8 that is mediated by calcineurin B-like protein (CBL) and CBL-interacting protein kinase (CIPK) complexes. We show that two CIPKs (CIPK9 and CIPK14) are able to interact with ACA8 in vivo and phosphorylate it in vitro. Transient co-overexpression of ACA8 with CIPK9 and the plasma membrane Ca2+ sensor CBL1 in tobacco leaf cells influences nuclear Ca2+ dynamics, specifically reducing the height of the second peak of the wound-induced Ca2+ transient. Stimulus-induced Ca2+ transients in mature leaves and seedlings of an aca8 T-DNA insertion line exhibit altered dynamics when compared with the wild type. Altogether our results identify ACA8 as a prominent in vivo regulator of cellular Ca2+ dynamics and reveal the existence of a Ca2+-dependent CBL-CIPK-mediated regulatory feedback mechanism, which crucially functions in the termination of Ca2+ signals. [ABSTRACT FROM AUTHOR]

Background: Immunosuppressive therapies are important parts of renal transplantation. Objective: To assess the present literature on the effectiveness of early introduction of mTOR inhibitors with or without calcineurin inhibitors (CNI) in renal transplant recipients in terms of renal functioning and graft survival. Methods: The current literature was reviewed following PROSPERO approval, assessing the role of immunosuppressive agent, mTOR inhibitors as an alternative to CNI within 6 months of renal transplantation by searching PubMed, EMBASE, Cochrane, Crossref, and Scopus. Results: 6 articles of early withdrawal of CNI and introduction of mTOR inhibitors within 6 months of renal transplantation were sought. Glomerular filtration rate (GFR) and serum creatinine were better in mTOR inhibitor group at 12 months. Biopsy-proven acute rejection (BPAR) was significantly higher in mTOR inhibitor group, though survival was comparable. Conclusion: On the basis of present literature, the early introduction of mTOR inhibitors causes substantial CNI minimization. The mTOR inhibitors are more favorable due to their complementary mechanism of action and favorable nephrotoxicity profile, better glomerular filtration, and lower serum creatinine with equivalent survival. However, the higher rejection rate may influence the use of these regimens in patients with moderate to high immunological risk. [ABSTRACT FROM AUTHOR]

Accumulation of misfolded/unfolded aggregated proteins in the brain is a hallmark of many neurodegenerative diseases affecting humans and animals. Dysregulation of calcium (Ca) and disruption of fast axonal transport (FAT) are early pathological events that lead to loss of synaptic integrity and axonal degeneration in early stages of neurodegenerative diseases. Dysregulated Ca in the brain is triggered by accumulation of misfolded/unfolded aggregated proteins in the endoplasmic reticulum (ER), a major Ca storing organelle, ultimately leading to neuronal dysfunction and apoptosis. Calcineurin (CaN), a Ca/calmodulin-dependent serine/threonine phosphatase, has been implicated in T cells activation through the induction of nuclear factor of activated T cells (NFAT). In addition to the involvement of several other signaling cascades, CaN has been shown to play a role in early synaptic dysfunction and neuronal death. Therefore, inhibiting hyperactivated CaN in early stages of disease might be a promising therapeutic strategy for treating patients with protein misfolding diseases. In this review, we briefly summarize the structure of CaN, inhibition mechanisms by which immunosuppressants inhibit CaN, role of CaN in maintaining neuronal and synaptic integrity and homeostasis and the role played by CaN in protein unfolding/misfolding neurodegenerative diseases. [ABSTRACT FROM AUTHOR]

Introduction Tuberculosis ( TB) is an important cause of morbidity and mortality in renal transplant recipients. Immunosuppressive drugs are one of the most important risk factor for post-transplant tuberculosis ( PTTB). A paucity of data exists about the impact of the type of calcineurin inhibitor on PTTB. Methods In this retrospective study, all adult patients on calcineurin inhibitor-based immunosuppression were included. Patients receiving TB chemoprophylaxis were excluded. Diabetes, duration of dialysis, hepatitis B and C, past treated TB, induction therapy, type of antimetabolite, acute rejection, new onset of diabetes after renal transplantation ( RT) ( NODAT) and cytomegalovirus ( CMV) were analyzed in tacrolimus (Tac) and cyclosporine (CsA) groups. Primary outcome was incidence of TB and secondary outcomes were timeline of development of TB after RT and pattern of TB in the two groups. Results Of the 1664 patients included, 582 patients received CsA-based immunosuppression while 1082 received Tac-based immunosuppression. Duration of dialysis, positive tuberculin skin test, use of induction, mycophenolate mofetil use, CMV infection, and NODAT were significantly more, and hepatitis B infection, past treated TB, and acute rejection episodes were significantly less in the Tac group. At the end of follow-up, incidence of TB in the Tac group was significantly less than in the CsA group (6.1% vs 19.9%, P<.001). Mean time for development of TB after RT was similar in both the groups and nodal and disseminated TB were more common in the Tac group. Conclusion In conclusion, our study shows that use of Tac as compared to CsA significantly decreases incidence of PTTB. Time of infection since transplant was similar in both the groups. However, nodal and disseminated TB were more common in the Tac group. [ABSTRACT FROM AUTHOR]

The management of connective tissue disease-associated interstitial lung disease (CTD-ILD) is complex and this arena offers many challenges to the practicing clinician. Unfortunately, treatment strategies and recommendations are often based on experience rather than evidence, and there are few effective therapeutic options. Pharmacologic intervention with immunosuppression is usually the mainstay of therapy and is reserved for those with clinically significant and/or progressive ILD. There is a desperate need for controlled trials across the spectrum of CTD-ILD and a number of potentially promising novel therapies warrant further study. It is important to address co-morbid conditions or aggravating factors (e.g., gastroesophageal reflux, aspiration, bone health, pulmonary hypertension,Pneumocystis jiroveciprophylaxis) and to institute non-pharmacologic management strategies (e.g., supplemental oxygen and cardiopulmonary rehabilitation) as part of a comprehensive treatment plan in CTD-ILD. [ABSTRACT FROM AUTHOR]

Objectives Cardiomyocyte hypertrophy is an important structural feature of diabetic cardiomyopathy. Calcineurin/nuclear factor of activated T-cell ( NFAT) pathway plays a central role in the pathogenesis of cardiac hypertrophy. The purpose of this study was to investigate the effects of tropisetron, a novel calcineurin inhibitor, on high glucose ( HG)-induced cardiomyocyte hypertrophy and its underlying mechanism. Methods H9c2 myocardial cells were treated with tropisetron or cyclosporine A 1 h before exposure to HG for 48 h. Key findings Exposure to HG resulted in enhanced cell size, protein content and atrial natriuretic peptide ( ANP) protein expression. HG significantly increased Ca2+ level, calcineurin expression and nuclear translocation of NFATc4. Both tropisetron and cyclosporine A markedly prevented the hypertrophic characteristic features, calcineurin overexpression and nuclear localization of NFATc4 while intracellular Ca2+ was not affected. Conclusion Our results showed that tropisetron may have protective effects against HG-induced cardiomyocyte hypertrophy. The mechanism responsible for this beneficial effect seems to be, at least in part, blockade of calcineurin/ NFAT signalling pathway. [ABSTRACT FROM AUTHOR]

Saccharomyces cerevisiae Rch1 is structurally similar to both the vertebrate solute carrier SLC10A7 and Candida albicans Rch1. We show here that ScRCH1 is a functional homolog of CaRCH1 . In S. cerevisiae , overexpression of ScRCH1 suppresses, but deletion of ScRCH1 does not affect, the lithium and rapamycin tolerance of pmr1 cells. Overexpression of ScRCH1 reduces expression of ENA1 , prevents sustained accumulation of cytosolic calcium and reduces the activation level of calcium/calcineurin signaling in pmr1 cells. Therefore, similar to the situation in the pathogen C. albicans , ScRch1 negatively regulates the cytosolic homeostasis in response to high levels of extracellular calcium. ScRch1 proteins distribute as multiple foci in the plasma membrane prior to cell division, move toward and concentrate at the bud neck as the bud grows in size, and disperse again along the plasma membrane immediately prior to cytokinesis. Furthermore, our genetic and biochemical data also demonstrate that transcriptional expression of RCH1 is positively regulated by calcium/calcineurin signaling through the sole CDRE element in its promoter. [ABSTRACT FROM AUTHOR]

Regulator of calcineurin (RCAN) is a calcineurin-interacting protein that inhibits calcineurin phosphatase when overexpressed, often upregulated under neuropathological conditions with impaired learning and memory processes, such as Down syndrome or Alzheimer's disease. Thermotactic behavior in the nematode Caenorhabditis elegans is a form of memory in which calcineurin signaling plays a pivotal role in the thermosensation of AFD neurons. In this study, we found that rcan-1 deletion mutants exhibited cryophilic behavior dependent on tax-6 , which was rescued by expressing rcan-1 in AFD neurons. Interaction between RCAN-1 and TAX-6 requires the conserved PxIxIT motif of RCAN-1, without which thermotactic behavior could not be fully rescued. In addition, the loss of crh-1 /CREB suppressed the thermotaxis phenotypes of rcan-1 and tax-6 mutants, indicating that crh-1 is crucial in thermotaxis memory in these mutants. Taken together, our results suggest that rcan-1 is an inhibitory regulator of tax-6 and that it acts in the formation of thermosensory behavioral memory in C . elegans . [ABSTRACT FROM AUTHOR]

Calcineurin inhibitors, such as cyclosporine A and FK506, are used as immunosuppressant drugs, but their adverse effects on male reproductive function remain unclear. The testis expresses somatic calcineurin and a sperm-specific isoform that contains a catalytic subunit (PPP3CC) and a regulatory subunit (PPP3R2). We demonstrate herein that male mice lacking Ppp3cc or Ppp3r2 genes (knockout mice) are infertile, with reduced sperm motility owing to an inflexible midpiece. Treatment of mice with cyclosporine A or FK506 creates phenocopies of the sperm motility and morphological defects. These defects appear within 4 to 5 days of treatment, which indicates that sperm-specific calcineurin confers midpiece flexibility during epididymal transit. Male mouse fertility recovered a week after we discontinued treatment. Because human spermatozoa contain PPP3CC and PPP3R2 as a form of calcineurin, inhibition of this sperm-specific calcineurin may lead to the development of a reversible male contraceptive that would target spermatozoa in the epididymis. [ABSTRACT FROM AUTHOR]

Fluorescence complementation ( FC) techniques are expedient for analyzing bimolecular protein-protein interactions. Here we aimed to develop a method for visualization of ternary protein complexes using dual-color trimolecular fluorescence complementation (Tri FC)., Dual-color TriFC combines protein fragments of mCherry and mVenus, in which a scaffold protein is bilaterally fused to C-terminal fragments of both fluorescent proteins and combined with potential interacting proteins fused to an N-terminal fluorescent protein fragment. For efficient visual verification of ternary complex formation, TriFC was combined with a cytoplasm to plasma membrane translocation assay., Modular vector sets were designed which are fully compatible with previously reported bimolecular fluorescence complementation (Bi FC) vectors. As a proof-of-principle, the ternary complex formation of the PP2B protein phosphatase Calcineurin-A/Calcineurin-B with Calmodulin-2 was investigated in transiently transformed Nicotiana benthamiana leaf epidermal cells. The results indicate a Calcineurin-B-induced interaction of Calmodulin-2 with Calcineurin-A., Tri FC and the translocation of Tri FC complexes provide a novel tool to investigate ternary complex formations with the simplicity of a Bi FC approach. The robustness of FC applications and the opportunity to quantify fluorescence complementation render this assay suitable for a broad range of interaction analyses. [ABSTRACT FROM AUTHOR]

The view of the lysosome as the terminal end of cellular catabolic pathways has been challenged by recent studies showing a central role of this organelle in the control of cell function. Here we show that a lysosomal Ca2+ signalling mechanism controls the activities of the phosphatase calcineurin and of its substrate TFEB, a master transcriptional regulator of lysosomal biogenesis and autophagy. Lysosomal Ca2+ release through mucolipin 1 (MCOLN1) activates calcineurin, which binds and dephosphorylates TFEB, thus promoting its nuclear translocation. Genetic and pharmacological inhibition of calcineurin suppressed TFEB activity during starvation and physical exercise, while calcineurin overexpression and constitutive activation had the opposite effect. Induction of autophagy and lysosomal biogenesis through TFEB required MCOLN1-mediated calcineurin activation. These data link lysosomal calcium signalling to both calcineurin regulation and autophagy induction and identify the lysosome as a hub for the signalling pathways that regulate cellular homeostasis. [ABSTRACT FROM AUTHOR]

Mycobacteriophage D29 encodes a protein Gp66 which has been predicted to be a calcineurin family phosphoesterase. Phylogenetically Gp66 and related proteins mostly derived from mycobacteriophages form a distinct clade within this family. Interestingly, the presence of gene 66 orthologs can be traced to bacteria of diverse phylogenetic lineages such as Aquifex aeolicus, a deep branching eubacteria and Methanococcus jannaschii, an archaebacteria. The promiscuous nature of gene 66 suggests that it may have been transferred across genus barriers by horizontal gene transfer mechanisms. The biological function of members of this novel clade comprising mostly the mycobacteriophage phosphoesterases have not been elucidated so far. In this investigation, it has been demonstrated for the first time that Gp66, a member of this novel family, is a 2′, 3′ cyclic phosphodiesterase. The gene is expressed during phage infection and the net result is negative regulation of bacteriophage as well as bacterial growth. [ABSTRACT FROM AUTHOR]

Background: Conversion from calcineurin inhibitor (CNI) to mTOR inhibitors may reduce and even halt the progression of chronic allograft dysfunction (CAD) which is the most important cause of renal allograft loss. We aimed to investigate the effects of conversion from CNI to everolimus on parameters of fibrosis, inflammation, glomerulotubular damage and vascular functions in renal transplant recipients. Methods: Fifteen stable renal transplant recipients who were under CNI treatment (male/female 13/2, mean age 41 ± 10 years) were enrolled and switched to everolimus. Serum and urinary transforming growth factor-β (TGF-β), urinary neutrophil gelatinase-associated lipocalin (NGAL) and monocyte chemoattractant protein-1 (MCP-1) were measured as markers of fibrosis, tubular damage and inflammation. As parameters of vascular functions, pulse wave velocity (PWV), augmentation index (AIx), serum asymmetric dimethyl-arginine and fibroblast growth factor-23 (FGF-23) were measured. All these measurements were repeated at the 3rd month of conversion. Results: Estimated GFR (52 ± 7-57 ± 11 ml/min/l.73 m, p = 0.02) (was increased after conversion to everolimus. However, serum uric acid levels were significantly decreased (6.21 ± 1.21-5.50 ± 1.39 mg/dL, p = 0.01). Serum TGF-β levels (8727 ± 2897-1943 ± 365 pg/mL, p = 0.03) and urinary NGAL levels (26 ± 10-12 ± 2 ng/mg creatinine, p = 0.05) were significantly decreased. However, urinary MCP-1, FGF-23, PWV and AIx did not change. Urinary TGF-β was associated with urinary NGAL ( r = 0.62, p = 0.01), urinary MCP-1 ( r = 0.68, p = 0.005) and proteinuria ( r = 0.50, p = 0.05). Conclusion: Conversion from CNI to everolimus resulted in significant decreases of serum TGF-β and urinary NGAL which may represent less fibrosis and tubular damage. Association of urinary TGF-β with NGAL and MCP-1 suggests that tubular damage, fibrosis and inflammation may act together for progression of CAD. [ABSTRACT FROM AUTHOR]

Chaperone-mediated autophagy (CMA) targets soluble proteins for lysosomal degradation. Here we found that CMA was activated in T cells in response to engagement of the T cell antigen receptor (TCR), which induced expression of the CMA-related lysosomal receptor LAMP-2A. In activated T cells, CMA targeted the ubiquitin ligase Itch and the calcineurin inhibitor RCAN1 for degradation to maintain activation-induced responses. Consequently, deletion of the gene encoding LAMP-2A in T cells caused deficient in vivo responses to immunization or infection with Listeria monocytogenes. Impaired CMA activity also occurred in T cells with age, which negatively affected their function. Restoration of LAMP-2A in T cells from old mice resulted in enhancement of activation-induced responses. Our findings define a role for CMA in regulating T cell activation through the targeted degradation of negative regulators of T cell activation. [ABSTRACT FROM AUTHOR]

This viewpoint presents a unifying concept for the treatment of atopic dermatitis ( AD) that is based on the improvement of deficient Notch signalling, which appears to represent the fundamental epithelial defect of AD resulting in epidermal and immunological barrier dysfunction. One study of AD patients demonstrated a marked epidermal deficiency of Notch receptors and several mouse models with genetically suppressed Notch signalling exhibit dry skin, signs of scratching, skin barrier abnormalities, increased transepidermal water loss and Th2 cell-mediated immunological changes closely resembling human AD. Notch signalling is critically involved in the differentiation of regulatory T cells, in the feedback inhibition of activated innate immunity, in the repression of activating protein-1 ( AP-1), the regulation of late epidermal differentiation associated with filaggrin- and stratum corneum barrier lipid processing, in aquaporin 3- and claudin-1 expression and in keratinocyte-mediated release of thymic stromal lymphopoietin ( TSLP), which promotes Th2-driven immune responses with TSLP- and IL-31-mediated stimulation of cutaneous sensory neurons involved in the induction of itch. Translational evidence will be provided that all major therapeutic regimens employed for the treatment of AD such as glucocorticoids, calcineurin inhibitors and UV radiation may converge in the upregulation of impaired Notch signalling, the proposed pathogenic defect of AD. [ABSTRACT FROM AUTHOR]

Calcineurin is an essential serine/threonine phosphatase that plays vital roles in neuronal development and function, heart growth, and immune system activation. Calcineurin is unique in that it is the only phosphatase known to be activated by calmodulin in response to increasing intracellular calcium concentrations. Calcium-loaded calmodulin binds to the regulatory domain of calcineurin, resulting in a conformational change that removes an autoinhibitory domain from the active site of the phosphatase. We have determined a 1.95 Å crystal structure of calmodulin bound to a peptide corresponding to its binding region from calcineurin. In contrast to previous structures of this complex, our structure has a stoichiometry of 1:1 and has the canonical collapsed, wraparound conformation observed for many calmodulin–substrate complexes. In addition, we have used size-exclusion chromatography and time-resolved fluorescence to probe the stoichiometry of binding of calmodulin to a construct corresponding to almost the entire regulatory domain from calcineurin, again finding a 1:1 complex. Taken in sum, our data strongly suggest that a single calmodulin protein is necessary and sufficient to bind to and activate each calcineurin enzyme. [ABSTRACT FROM AUTHOR]

Bone remodeling is a continuous process of osteoblastic bone formation and osteoclastic bone resorption to maintain normal bone mass. NFATc1 is the master regulator of osteoclastogensis and transcriptionally activated by c-Fos and NF-кB in response to receptor activator of NF-кB ligand (RANKL) treatment. Defective entry into mitosis 1 (Dim1) is a nuclear protein that is implicated in pre-mRNA splicing and cell cycle progression, but the possible role of Dim1 in regulating other cellular processes remains unknown. Here, we demonstrate that Dim1 attenuates RANKL-induced osteoclastogenesis by targeting NFATc1 signaling pathway. Expression levels of Dim1 and NFATc1 are significantly increased during the formation of multinucleated osteoclasts. RNAi-mediated knockdown of Dim1 markedly enhances the expression of NFATc1 and its target genes, leading to the increase of RANKL-induced osteoclastogenesis in bone marrow-derived macrophages. Conversely, ectopic expression of Dim1 decreases RANKL-induced osteoclast differentiation by silencing NFATc1 and its target genes, further linking Dim1 to the dynamic regulation of osteoclastogenesis. Consistent with this notion, ChIP and interaction studies show that Dim1 directly associates with c-Fos and prevents c-Fos from binding to the NFATc1 promoter, resulting in targeted inactivation of the NFATc1 gene. Therefore, our studies reveal an unrecognized role for Dim1 as a master modulator of osteoclast differentiation, as well as the molecular mechanism underlying its repressive action toward osteoclastogensis. [ABSTRACT FROM AUTHOR]

Abstract: Cisapride has been shown to have electrophysiological effects on the heart. The aim of this study was to investigate whether cisapride has effects on cardiac hypertrophy. Rat and cellular models of cardiac hypertrophy were used in this study. Cell surface area (CSA), mRNA and protein expression were used to evaluate cardiac hypertrophy. Cardiac function was measured by echocardiography. Cisapride attenuated ISO-induced increase in CSA in a dose-dependent manner in cultured neonatal rat cardiomyocytes. A significant anti-hypertrophic effect was achieved by cisapride 0.01μM (P<0.05). Cisapride repressed the increased mRNA levels of ANP, BNP, β-MHC in ISO-treated cells (P<0.05). However, mallotoxin or GR113808 did not influence anti-hypertrophic effects of cisapride. In addition, cisapride inhibited the increase of intracellular Ca2+ ([Ca2+]i) and the upregulation of protein levels of calcineurin and NFATc-3 (P<0.05) as well as prevented the downregulation of p-NFATc-3 (P<0.01) induced by ISO. Consistently, cisapride (0.5mg/kg/day) produced inhibitory effects on cardiac hypertrophy, including the suppression of ANP, BNP, β-MHC, calcineurin, and NFATc-3; elevation of p-NFATc-3; reduction of cross-sectional area of cardiomyocytes in rat heart; and restoration of cardiac dysfunction by improving left ventricular diastolic and systolic performance. Importantly, cisapride 0.5 and 5.0mg/kg/day did not cause prolongation of QT and QTc intervals in rats. In conclusion, cisapride possesses a prominent anti-hypertrophic property which is likely to be conferred by its ability to downregulate Ca2+/calcineurin/NFAT and the present data provide new insight into this drug action. [Copyright &y& Elsevier]

Aims Calcineurin inhibitors ( CNIs) taken after heart transplantation lead to excellent short-term outcomes, but long-term use may cause chronic nephrotoxicity. Our aim was to identify, appraise, select and analyse all high-quality research evidence relevant to the question of the clinical impact of CNI-sparing strategies in heart transplant patients. Methods We carried out a systematic review and meta-analysis of randomized controlled trials on CNI reduction in heart transplant recipients. Primary outcomes were kidney function and acute rejection after 1 year. Secondary outcomes included graft loss, all-cause mortality and adverse events. Results Eight open-label studies were included, with 723 patients (four tested de novo CNI reduction and four maintenance CNI reduction). Calcineurin inhibitor reduction did not improve creatinine clearance at 12 months 5.46 [−1.17, 12.03] P = 0.32 I2 = 65.4%. Acute rejection at 12 months (55/360 vs. 52/332), mortality (18/301 vs. 15/270) and adverse event rates (55/294 vs. 52/281) did not differ between the low- CNI and standard- CNI groups. There was significant benefit on creatinine clearance in patients with impaired renal function at 6 months [+12.23 (+5.26, +18.82) ml min−1, P = 0.0003] and at 12 months 4.63 [−4.55, 13.82] P = 0.32 I2 = 75%. Conclusions This meta-analysis did not demonstrate a favourable effect of CNI reduction on kidney function, but there was no increase in acute rejection. To provide a better analysis of the influence of CNI reduction patterns and associated treatments, a meta-analysis of individual patient data should be performed. [ABSTRACT FROM AUTHOR]

Background: The calcium-dependent phosphatase calcineurin is highly expressed in the amygdala, a brain area important for behaviors related to mood disorders and anxiety. Organ transplant patients are administered the calcineurin inhibitor cyclosporine A (CsA) chronically and demonstrate an increased incidence of anxiety and mood disorders. It is therefore important to determine whether chronic blockade of calcineurin may contribute to symptoms of anxiety and depression in these patients. Methods: Pharmacological (CSA) and viral-mediated gene transfer (adeno-associated viral expression of short hairpin RNA [shRNA]) approaches were used to inhibit calcineurin activity systemically or selectively in the amygdala of the mouse brain to determine the role of calcineurin in behaviors related to anxiety and depression. Results: Systemic inhibition of calcineurin activity with CsA or local downregulation of calcineurin levels in the amygdala using adeno-associated viral-delivered shRNAs targeting calcineurin B increased measures of anxiety-like behavior in the elevated plus maze, the light/dark box, and the open field test. A decrease in locomotor activity was also observed in mice treated systemically with CsA. In the forced swim model of depression-like behavior, both systemic CsA treatment and shRNA-mediated calcineurin blockade in the amygdala significantly increased immobility. Conclusions: Taken together, these data demonstrate that decreasing calcineurin activity in the amygdala increases anxiety-like behaviors and to some extent depression-like behaviors. These studies suggest that chronic administration of CsA to organ transplant patients could have significant effects on anxiety and mood and this should be recognized as a potential clinical consequence of treatment to prevent transplant rejection. [ABSTRACT FROM AUTHOR]

Abstract: Cord blood transplantation (CBT) is a known risk factor for human herpesvirus-6 (HHV-6) infection. We analyzed the nature of HHV-6 infections in 125 double-unit CBT recipients (median age, 42 years) transplanted for hematologic malignancies with calcineurin inhibitor/mycophenolate mofetil prophylaxis and no antithymocyte globulin. One hundred seventeen patients (94%) reactivated HHV-6 by quantitative plasma PCR (median peak, 7600 copies/mL; range, 100 to 160,000) at a median of 20 days (range, 10 to 59) after transplantation. HHV-6 encephalitis occurred in 2 patients (1.6%), of whom 1 died and 1 recovered with therapy. No association was found between high-level HHV-6 viremia (≥10,000 or ≥25,000 copies/mL) and age, diagnosis, conditioning intensity, or dominant unit characteristics or between high-level viremia and transplant outcomes (engraftment, cytomegalovirus reactivation, day 100 grades II to IV acute graft-versus-host disease, day 100 transplant-related mortality, or 1-year disease-free survival). HHV-6 therapy delayed the onset of cytomegalovirus reactivation. Interestingly, HHV-6 resolution was observed in untreated patients, and resolution of viremia correlated with absolute lymphocyte count recovery. We observed a low incidence of encephalitis and no association with CBT outcomes. Our data suggest therapy in uncomplicated viremia may not be warranted. However, further investigation of the risk-to-benefit of HHV-6 viremia treatment and standardization of PCR testing is required. [Copyright &y& Elsevier]

Land plants have evolved a host of anatomical and molecular adaptations for terrestrial growth. Many of these adaptations are believed to be elaborations of features that were present in their algal-like progenitors. In the model plant Arabidopsis, 10 Calcineurin B-Like proteins (CBLs) function as calcium sensors and modulate the activity of 26 CBL-Interacting Protein Kinases (CIPKs). The CBL-CIPK network coordinates environmental responses and helps maintain proper ion balances, especially during abiotic stress. We identified and analyzed CBL and CIPK homologs in green lineages, including CBLs and CIPKs from charophyte green algae, the closest living relatives of land plants. Phylogenomic evidence suggests that the network expanded from a small module, likely a single CBL-CIPK pair, present in the ancestor of modern plants and algae. Extreme conservation of the NAF motif, which mediates CBL-CIPK physical interactions, among all identified CIPKs supports the interpretation of CBL and CIPK homologs in green algae and early diverging land plants as functionally linked network components. We identified the full complement of CBL and CIPK loci in the genome of Physcomitrella, a model moss. These analyses demonstrate the strong effects of a recent moss whole genome duplication: CBL and CIPK loci appear in cognate pairs, some of which appear to be pseudogenes, with high sequence similarity. We cloned all full-length transcripts from these loci and performed yeast two-hybrid analyses to demonstrate CBL-CIPK interactions and identify specific connections within the network. Using phylogenomics, we have identified three ancient types of CBLs that are discernible by N-terminal localization motifs and a "green algal-type" clade of CIPKs with members from Physcomitrella and Arabidopsis. [ABSTRACT FROM AUTHOR]

Summary: Vertebrates develop organs and appendages in a proportionally coordinated manner, and animals that regenerate them do so to the same dimensions as the original structures. Coordinated proportional growth involves controlled regulation between allometric and isometric growth programs, but it is unclear what executes this control. We show that calcineurin inhibition results in continued allometric outgrowth of regenerating fins beyond their original dimensions. Calcineurin inhibition also maintains allometric growth of juvenile fins and induces it in adult fins. Furthermore, calcineurin activity is low when the regeneration rate is highest, and its activity increases as the rate decreases. Growth measurements and morphometric analysis of proximodistal asymmetry indicate that calcineurin inhibition shifts fin regeneration from a distal growth program to a proximal program. This shift is associated with the promotion of retinoic acid signaling. Thus, we identified a calcineurin-mediated mechanism that operates as a molecular switch between position-associated isometric and allometric growth programs. [Copyright &y& Elsevier]

Transmembrane α-amino-3-hydroxy-5-methyl-4-isoxazole propionate ( AMPA) receptor regulatory proteins ( TARPs) are auxiliary subunits that regulate AMPA receptor trafficking to the plasma membrane and localization to postsynaptic sites. The classical TARP family consists of four members: stargazin/γ-2, γ-3, γ-4 and γ-8. The TARP γ-8 isoform, which is highly expressed in the hippocampus, has a unique, long C-terminal domain with five distinct regions: two glycine-rich regions, a serine/arginine-rich region, a proline/alanine ( P/ A) rich region, and a PSD-95/ Dlg/ ZO-1 ( PDZ) binding motif. We performed mass spectrometry and immunoprecipitation assays to identify specific binding partners for the γ-8 C-terminal tail and found that γ-8, but not stargazin/γ-2, co-immunoprecipitated with calcineurin/ PP2 B, a Ca2+/calmodulin-dependent Ser/ Thr phosphatase. Co-immunoprecipitation and immunoblot analyses of lysates from COS-7 cells co-transfected with calcineurin and either wild type or chimeric γ-8 revealed that a section of the C-terminal tail (residues 356-421) can bind calcineurin. Futhermore, γ-8 lacking the P/ A-rich region (residues 383-399) did not bind to calcineurin. In addition, the GST-γ-8 C-terminal tail (residues 353-414) fusion protein containing the P/ A-rich region bound to purified calcineurin in a Ca2+/calmodulin-dependent manner, whereas GST-γ-8 with a deletion of the P/ A-rich region did not. Peptide competition assays demonstrated that γ-8 may interact with the hydrophobic pocket defined by β-sheet 14 and/or adjacent regions of the catalytic A subunit of calcineurin. These results indicate that the γ-8 P/ A-rich region is essential for binding calcineurin, suggesting that the γ-8/calcineurin complex may regulate AMPA receptor phosphorylation and trafficking. Structured digital abstract with by () to by () with by () with and by () with , and by () with and by () with and by (, , ) with by () with , , and by () with by (), and by () [ABSTRACT FROM AUTHOR]

Calcineurin, a Ca2+/calmodulin-dependent protein phosphatase, plays a critical role in controlling skeletal muscle fiber type. However, little information is available concerning the expression of calcineurin in goat. Therefore, protein phosphatase 3 catalytic subunit alpha isoform (PPP3CA) gene, also called calcineurin Aa, was cloned and its expression characterized in Tianfu goat muscle. Real time quantitative polymerase chain reaction (RT-qPCR) analyses revealed that Tianfu goat PPP3CA was detected in cardiac muscle, biceps femoris muscle, abdominal muscle, longissimus dors muscle, and soleus muscle. High expression levels were found in biceps femoris muscle, longissimus muscle and abdominal muscle (p < 0.01), and low expression levels were seen in cardiac muscle and soleus muscle (p > 0.05). In addition, the spatial-temporal mRNA expression levels showed different variation trends in different muscles with the age of the goats. Western blotting further revealed that PPP3CA protein was expressed in the above-mentioned tissues, with the highest level in biceps femoris muscle, and the lowest level in soleus muscle. In this study, we isolated the full-length coding sequence of Tianfu goat PPP3CA gene, analyzed its structure, and investigated its expression in different muscle tissues from different age stages. These results provide a foundation for understanding the function of the PPP3CA gene in goats. [ABSTRACT FROM AUTHOR]

Recently there has been much interest in the Regulators of Calcineurin (RCAN) proteins which are important endogenous modulators of the calcineurin-NFATc signalling pathway. They have been shown to have a crucial role in cellular programmes such as the immune response, muscle fibre remodelling and memory, but also in pathological processes such as cardiac hypertrophy and neurodegenerative diseases. In vertebrates, the RCAN family form a functional subfamily of three members RCAN1, RCAN2 and RCAN3 whereas only one RCAN is present in the rest of Eukarya. In addition, RCAN genes have been shown to collocate with RUNX and CLIC genes in ACD clusters (ACD21, ACD6 and ACD1). How the RCAN genes and their clustering in ACDs evolved is still unknown. After analysing RCAN gene family evolution using bioinformatic tools, we propose that the three RCAN vertebrate genes within the ACD clusters, which evolved from single copy genes present in invertebrates and lower eukaryotes, are the result of two rounds of whole genome duplication, followed by a segmental duplication. This evolutionary scenario involves the loss or gain of some RCAN genes during evolution. In addition, we have analysed RCAN gene structure and identified the existence of several characteristic features that can be involved in RCAN evolution and gene expression regulation. These included: several transposable elements, CpG islands in the 5′ region of the genes, the existence of antisense transcripts (NAT) associated with the three human genes, and considerable evidence for bidirectional promoters that regulate RCAN gene expression. Furthermore, we show that the CpG island associated with the RCAN3 gene promoter is unmethylated and transcriptionally active. All these results provide timely new insights into the molecular mechanisms underlying RCAN function and a more in depth knowledge of this gene family whose members are obvious candidates for the development of future therapies. [ABSTRACT FROM AUTHOR]

The receptor for stem cell factor (SCF) is expressed on mast cells and hematopoietic progenitors. SCF-induced signaling pathways remain incompletely defined. In this study, we identified calcineurin and regulator of calcineurin 1 (Rcan1) as novel components in SCF signaling. Calcineurin activity was induced in SCF-stimulated primary mouse and human mast cells. NFAT was activated by SCF in bone marrow-derived mast cells (BMMCs) and mouse bone marrow cells, which contain hematopoietic progenitors. SCFmediated activation also induced expression of Rcan1 in BMMCs. Rcan1-deficient BMMCs showed increased calcineurin activity and enhanced transcriptional activity of NF-kB and NFAT, resulting in increased IL-6 and TNF production following SCF stimulation. These results suggest that Rcan1 suppresses SCF-induced activation of calcineurin and NF-kB. We further demonstrated that SCF-induced Rcan1 expression is dependent on the transcription factor early growth response 1 (Egr1). Interestingly, SCF-induced Egr1 was also suppressed by Rcan1, suggesting a negative regulatory loop between Egr1 and Rcan1. Together, our findings revealed that calcineurin contributes to SCF-induced signaling, leading to NFAT activation, which, together with NF-kB and Egr1, is suppressed by Rcan1. Considering the wide range of biological functions of SCF, these novel regulatory mechanisms in SCF signaling may have broad implications. [ABSTRACT FROM AUTHOR]

Calcineurin B‐like protein‐interacting protein kinases (CIPKs) are components of Ca2+ signaling in responses to abiotic stresses. In this work, the full‐length cDNA of a novel CIPK gene (TaCIPK14) was isolated from wheat and was found to have significant sequence similarity to OsCIPK14/15. Subcellular localization assay revealed the presence of TaCIPK14 throughout the cell. qRT‐PCR analysis showed that TaCIPK14 was upregulated under cold conditions or when treated with salt, PEG or exogenous stresses related signaling molecules including ABA, ethylene and H2O2. Transgenic tobaccos overexpressing TaCIPK14 exhibited higher contents of chlorophyll and sugar, higher catalase activity, while decreased amounts of H2O2 and malondialdehyde, and lesser ion leakage under cold and salt stresses. In addition, overexpression also increased seed germination rate, root elongation and decreased Na+ content in the transgenic lines under salt stress. Higher expression of stress‐related genes was observed in lines overexpressing TaCIPK14 compared to controls under stress conditions. In summary, these results suggested that TaCIPK14 is an abiotic stress‐responsive gene in plants. [ABSTRACT FROM AUTHOR]

Highlights: [•] Green light provided protection against the deleterious effects of UV irradiation upon yeast cells. [•] High density green photons protected the yeast cells against UV in synergism with unfolded protein response inducers. [•] Photoprotective action of green light against UV required intact UPR pathway. [•] The cnb1Δ cells, lacking the regulatory subunit of calcineurin, became more sensitive to UV after green light exposure. [ABSTRACT FROM AUTHOR]

Highlights: [•] RCAN1 self-associates and forms multimers. [•] RCAN1 multimer formation is promoted by the Dyrk1A-mediated phosphorylation. [•] Phospho-RCAN1 expression is lower in aged Dyrk1A TG mice than in littermates. [•] Dyrk1A plays a role in the formation of insoluble RCAN1 aggregates upon aging. [ABSTRACT FROM AUTHOR]

Regulator of calcineurin 1 (RCAN1) controls the activity of calcium/calmodulin-dependent phosphatase calcineurin (CaN), which has been implicated in human anxiety disorders. Previously, we reported that RCAN1 functioned as an inhibitor of CaN activity in the brain. However, we now find enhanced phosphorylation of a CaN substrate, cAMP response element-binding protein (CREB), in the brains of Rcan1 knock-out (KO) mice. Consistent with enhanced CREB activation, we also observe enhanced expression of a CREB transcriptional target, brain-derived neurotrophic factor (BDNF) in Rcan1KOmice. We also discovered thatRCAN1deletion or blockade ofRCAN1-CaN interaction reduced CaN and protein phosphatase-1 localization to nuclear-enriched protein fractions and promoted CREB activation. Because of the potential links between CREB, BDNF, and anxiety, we examined the role of RCAN1 in the expression of innate anxiety. Rcan1 KO mice displayed reduced anxiety in several tests of unconditioned anxiety. Acute pharmacological inhibition of CaN rescued these deficits while transgenic overexpression of human RCAN1 increased anxiety. Finally, we found that Rcan1KOmice lacked the early anxiogenic response to the selective serotonin reuptake inhibitor (SSRI) fluoxetine and had improved latency for its therapeutic anxiolytic effects. Together, our study suggests that RCAN1 plays an important role in the expression of anxiety-related and SSRI-related behaviors through CaN-dependent signaling pathways. These results identify RCAN1 as a mediator of innate emotional states and possible therapeutic target for anxiety. [ABSTRACT FROM AUTHOR]

Background: Zinc, an essential trace element, inhibits osteoclast differentiation in vitro and in vivo. The molecular mechanism for the inhibitory effect of zinc, however, is poorly understood. The purpose of this study was to investigate the effect of zinc and determine its molecular mechanism on receptor activator of NF-κB ligand (RANKL)-induced osteoclastogenesis in mouse bone marrow-derived monocyte cells (BMMs) and RAW264.7 cells. Results: In BMMs, zinc treatment during osteoclast differentiation decreased RANKL-induced osteoclast formation in a dose-dependent manner. We show that zinc suppressed the mRNA levels of nuclear factor of activated T-cells, cytoplasmic 1 (Nfatc1). Zinc also accumulated phospho-Nfatc1 (p-Nfatc1) in the cytosol in a dose-dependent manner and inhibited the translocation of Nfatc1 to the nucleus in RAW264.7 cells. Zinc suppressed the activities of Nfatc1 in the nucleus without changing the activities of NF-κB in RAW264.7 cells. In contrast, calcineurin activity decreased in response to zinc but its protein level was unchanged. RANKL-induced Ca2+ oscillations were inhibited by zinc treatment, but phospho-phospholipase Cγ1 (p-PLCγ1), the upstream signaling molecule of Ca2+ oscillations, was unaffected. Moreover, a constitutively active form of Nfatc1 obviously rescued suppression of osteoclastogenesis by zinc. Conclusions: Taken together, these results demonstrate for the first time that the inhibitory effect of zinc during osteoclastogesis is caused by suppressing the Ca2+-Calcineurin-NFATc1 signaling pathway. Thus, zinc may be a useful therapeutic candidate for the prevention of bone loss caused by NFATc1 activation in osteoclasts. [ABSTRACT FROM AUTHOR]

Abstract: Cyclosporine A and FK506 produce immunosuppression by blocking calcineurin phosphatase activity and consequently activation of cytosolic Nuclear Factor of Activated T-cell (NFATc) transcription factor. Due to the chronic toxicity associated with their administration, the development of more specific immunosuppressants is currently an important unmet medical need. In this context, an immunosuppressant peptide derived from the CIC motif of the human Regulators of Calcineurin (RCAN) proteins has been shown to inhibit NFATc signaling without affecting general phosphatase activity of calcineurin. Here we show that protein kinase CK2 phosphorylates a conserved serine residue within the CIC motif of vertebrate RCANs, which increases its affinity for calcineurin and consequently its inhibition of NFATc-dependent gene expression in activated T-cells. Molecular modeling studies have led us to identify a positively charged interaction site on the surface of calcineurin where the phosphorylated serine residue of the CIC motif would normally locate. Finally, we have also identified RCAN3 as a new phosphoprotein with multiple phosphorylation sites. Therefore, our findings reveal for the first time a novel molecular mechanism underlying the regulation of calcineurin–NFATc signaling by means of phosphorylation of the CIC motif of RCAN proteins. The knowledge of how RCAN proteins modulate the calcineurin–NFATc pathway paves the way for the development of potent novel selective immunosuppressant drugs. [Copyright &y& Elsevier]

Abstract: Background: Tacrolimus (FK506) has a superior immunosuppressive effect compared with cyclosporine (CSA) without a significant increase in generalized infectious complications. Differences in specific infections such as Clostridium difficile (CDI) have not been reported. We investigated the relationship between calcineurin inhibitors and CDI, hypothesizing that choice of calcineurin inhibitor (CSA or FK506) after lung transplantation would have no effect on the incidence of CDI. Methods: We performed a retrospective chart review of lung transplant recipients between June 1, 2000, and December 31, 2005, at a single institution. Positive CDI assays through December 11, 2011, were also recorded. We used Student’s t- and chi-squared tests (α = 0.05) to compare CSA and FK506 groups. We calculated adjusted hazard ratios for CDI using Cox proportional hazard models. Results: We identified 217 lung transplant recipients: 106 patients in the CSA group and 111 patients in the FK506 group. A total of 31 patients (27.9%) in the FK506 group developed CDI postoperatively compared with 20 patients (18.9%) in the CSA group (P = 0.16). The adjusted hazard ratio for CDI in the FK506 group was not significantly higher (1.53; 95% confidence interval, 0.78–2.98). There was no significant difference in the intensive care unit or total length of stay, in-hospital incidence rate, time to first CDI episode, or recurrence rate between groups. Conclusions: The CDI rates were not significantly higher in the FK506 group than the CSA group in our study. These data are consistent with previous studies on FK506 that show no increase in infectious complications over CSA, and demonstrate its continued safety in lung transplantation. [Copyright &y& Elsevier]