In the Rat Midbrain, SG2NA and DJ-1 have Common Interactome, Including Mitochondrial Electron Transporters that are Comodulated Under Oxidative Stress.

Scaffold proteins Striatin and SG2NA assemble kinases and phosphatases into the signalling complexes called STRIPAK. Dysfunctional STRIPAKs cause cancer, cerebral cavernous malformations, etc. DJ-1, a sensor for oxidative stress, has long been associated with the Parkinson's disease, cancer, and immune disorders. SG2NA interacts with DJ-1 and Akt providing neuroprotection under oxidative stress. To dissect the role of SG2NA and DJ-1 in neuronal pathobiology, rat midbrain extracts were immunoprecipitated with SG2NA and sixty-three interacting proteins were identified. BN-PAGE followed by the LC-MS/MS showed 1030 comigrating proteins as the potential constituents of the multimeric complexes formed by SG2NA. Forty-three proteins were common between those identified by co-immunoprecipitation and the BN-PAGE. Co-immunoprecipitation with DJ-1 identified 179 interacting partners, of which forty-one also interact with SG2NA. Among those forty-one proteins immunoprecipitated with both SG2NA and DJ-1, thirty-nine comigrated with SG2NA in the BN-PAGE, and thus are bonafide constituents of the supramolecular assemblies comprising both DJ-1 and SG2NA. Among those thirty-nine proteins, seven are involved in mitochondrial oxidative phosphorylation. In rotenone-treated rats having Parkinson's like symptoms, the levels of both SG2NA and DJ-1 increased in the mitochondria; and the association of SG2NA with the electron transport complexes enhanced. In the hemi-Parkinson's model, where the rats were injected with 6-OHDA into the midbrain, the occupancy of SG2NA and DJ-1 in the mitochondrial complexes also increased. Our study thus reveals a new family of potential STRIPAK assemblies involving both SG2NA and DJ-1, with key roles in protecting midbrain from the oxidative stress.

The profile of expression of the scaffold protein SG2NA(s) differs between cancer types and its interactome in normal vis-a-vis breast tumor tissues suggests its wide roles in regulating multiple cellular pathways.

Striatin and SG2NA are scaffold proteins that form signaling complexes called STRIPAK. It has been associated with developmental abnormalities, cancer, and several other diseases. Our earlier studies have shown that SG2NA forms a complex with the cancer-associated protein DJ-1 and the signaling kinase Akt, promoting cancer cell survival. In the present study, we used bioinformatics analyses to confirm the existence of two isoforms of human SG2NA, i.e., 78 and 87 kDas. In addition, several smaller isoforms like 35 kDa were also seen in western blot analyses of human cell lysates. The expression of these isoforms varies between different cancer cell lines of human origin. Also, the protein levels do not corroborate with its transcript levels, suggesting a complex regulation of its expression. In breast tumor tissues, the expression of the 35 and 78 kDa isoforms was higher as compared to the adjacent normal tissues, while the 87 kDa isoform was found in the breast tumor tissues only. With the progression of stages of breast cancer, while the expression of 78 kDa isoform decreased, 87 kDa became undetectable. In co-immunoprecipitation assays, the profile of the SG2NA interactome in breast tumors vis-à-vis adjacent normal breast tissues showed hundreds of common proteins. Also, some proteins were interacted with SG2NA in breast tumor tissues only. We conclude that SG2NA is involved in diverse cellular pathways and has roles in cellular reprogramming during tumorigenesis of the breast.

Ectopic expression of 35 kDa and knocking down of 78 kDa SG2NAs induce cytoskeletal reorganization, alter membrane sialylation, and modulate the markers of EMT.

SG2NA is a protein of the striatin family that organizes STRIPAK complexes. It has splice variants expressing differentially in tissues. Its 78 kDa isoform regulates cell cycle, maintains homeostasis in the endoplasmic reticulum, and prevents oxidative injuries. The 35 kDa variant is devoid of the signature WD-40 repeats in the carboxy terminal, and its function is unknown. We expressed it in NIH 3T3 cells that otherwise express 78 kDa variant only. These cells (35 EE) have altered morphology, faster rate of migration, and enhanced growth as measured by the MTT assay. Similar phenotypes were also seen in cells where the endogenous 78 kDa isoform was downregulated by siRNA (78 KD). Proteomic analyses showed that several cancer-associated proteins are modulated in both 35 EE and 78 KD cells. The 35 EE cells have diffused actin fibers, distinctive ultrastructure, reduced sialylation, and increased expression of MMP2 & 9. The 78 KD cells also had diffused actin fibers and an upregulated expression of MMP2. In both cells, markers epithelial to mesenchymal transition (EMT) viz, E- & N-cadherins, ß-catenin, slug, vimentin, and ZO-1 were modulated partially in tune with the EMT process. Since NIH 3T3 cells are mesenchymal, we also expressed 35 kDa SG2NA in MCF-7 cells of epithelial origin. In these cells (MCF-7-35), the actin fibers were also diffused and the modulation of the markers was more in tune with the EMT process. However, unlike in 35 EE cells, in MCF-7-35 cells, membrane sialylation rather increased. We infer that ectopic expression of 35 kDa and downregulation of 78 kDa SG2NAs partially induce transformed phenotypes.

Mitoapocynin, a mitochondria targeted derivative of apocynin induces mitochondrial ROS generation and apoptosis in multiple cell types including cardiac myoblasts: a potential constraint to its therapeutic use.

Mitoapocynin is a triphenylphosphonium conjugated derivative of apocynin that specifically locates to the mitochondria. It has been developed as a mitochondrially targeted therapeutic antioxidant. We attempted to attenuate the mitochondrial ROS induced in H9c2 cardiac myoblast cells treated with norepinephrine. Mitoapocynin was a poor quencher of total ROS as detected by the fluoroprobe DCFH-DA. Using mitochondrial superoxide specific probe MitoSoxRed, we found that 5-10 µM mitoapocynin itself induces superoxide over and above that is generated by the norepinephrine treatment. A supposedly control molecule to mitoapocynin, the synthetic compound PhC11TPP, having the triphenylphosphonium group and a benzene moiety with C11 aliphatic chain spacer was also found to be a robust inducer of mitochondrial ROS. Subsequent assays with several cell lines viz., NIH3T3, HEK293, Neuro2A, MCF-7 and H9c2, showed that prolonged exposure to mitoapocynin induces cell death by apoptosis that can be partially prevented by the general antioxidant N-acetyl cysteine. Analyses of mitochondrial electron transport complexes by Blue Native Polyacrylamide gel electrophoresis showed that both mitoapocynin and PhC11TPP disrupt the mitochondrial Complex I and V, and in addition, PhC11TPP also damages the Complex IV. Our data thus highlights the limitations of the therapeutic use of mitoapocynin as an antioxidant.

Catestatin reverses the hypertrophic effects of norepinephrine in H9c2 cardiac myoblasts by modulating the adrenergic signaling.

Catestatin (CST) is a catecholamine release-inhibitory peptide secreted from the adrenergic neurons and the adrenal glands. It regulates the cardiovascular functions and it is associated with cardiovascular diseases. Though its mechanisms of actions are not known, there are evidences of cross-talk between the adrenergic and CST signaling. We hypothesized that CST moderates the adrenergic overdrive and studied its effects on norepinephrine-mediated hypertrophic responses in H9c2 cardiac myoblasts. CST alone regulated the expression of a number of fetal genes that are induced during hypertrophy. When cells were pre-treated CST, it blunted the modulation of those genes by norepinephrine. Norepinephrine (2 µM) treatment also increased cell size and enhanced the level of Troponin T in the sarcomere. These effects were attenuated by the treatment with CST. CST attenuated the immediate generation of ROS and the increase in glutathione peroxidase activity induced by norepinephrine treatment. Expression of fosB and AP-1 promoter-reporter constructs was used as the endpoint readout for the interaction between the CST and adrenergic signals at the gene level. It showed that CST largely attenuates the stimulatory effects of norepinephrine and other mitogenic signals through the modulation of the gene regulatory modules in a characteristic manner. Depending upon the dose, the signaling by CST appears to be disparate, and at 10-25 nM doses, it primarily moderated the signaling by the ß1/2-adrenoceptors. This study, for the first time, provides insights into the modulation of adrenergic signaling in the heart by CST.

Subcellular dynamics of variants of SG2NA in NIH3T3 fibroblasts.

SG2NA, a WD40 repeat protein of the Striatin subfamily, has four splicing and one messenger RNA edit variants. It is fast emerging as a scaffold for multimeric signaling complexes with roles in tissue development and disease. The green fluorescent protein (GFP)-tagged variants of SG2NA were ectopically expressed in NIH3T3 cells and their modulation by serum and GSK3ß-ERK signaling were monitored. The 87, 78, and 35 kDa variants showed a biphasic modulation by serum till 24 h but the 52 kDa variant remained largely unresponsive. Inhibition of phosphatases by okadaic acid increased the levels of the endogenous 78 kDa and the ectopically expressed GFP-tagged 87 and 78 kDa SG2NAs. Contrastingly, okadaic acid treatment reduced the level of GFP-tagged 35 kDa SG2NA, suggesting differential modes of their stability through phosphorylation-dephosphorylation. The inhibition of GSK3ß by LiCl showed a gradual decrease in the levels of 78 kDa. In the case of the other variants viz, GFP-tagged 35, 52, and 87 kDa, inhibition of GSK3ß caused an initial increase followed by a decrease with a subtle difference in kinetics and intensities. Similar results were also seen upon inhibition of GSK3ß by small interfering RNA. All the variants showed an increase followed by a decrease upon inhibition of extracellular-signal-regulated-kinase (ERK). These variants are localized in the plasma membrane, endoplasmic reticulum, mitochondria, and the nucleus with different propensities and no discernable subcellular distribution was seen upon stimulation by serum and the inhibition of phosphatases, GSK3ß, and ERK. Taken together, the variants of SG2NA are modulated by the kinase-phosphatase network in a similar but characteristic manner.

Apocynin prevents isoproterenol-induced cardiac hypertrophy in rat.

Oxidative stress is implicated in the pathogenesis of a plethora of cardiovascular diseases including interstitial fibrosis, contractile dysfunction, ischemia-reperfusion injury, and cardiac remodeling. However, antioxidant therapies targeting oxidative stress in the progression of those diseases have largely been unsuccessful. The current study evaluated the effects of a NADPH oxidase inhibitor, apocynin (Apo), on the production of reactive oxygen species and the development of pathological cardiac hypertrophy under sustained ß-adrenergic stimulation in male Wistar rats. As evident from the HW/BW ratio, HW/TL ratio, echocardiography, and histopathology, hypertrophic responses induced by isoproterenol (Iso; 5 mg/Kg body weight, subcutaneous) were blocked by Apo (10 mg/Kg body weight, intraperitoneal). Iso treatment increased the transcript levels of cybb and p22-phox, the two subunits of Nox. Iso treatment also caused a decrease in reduced glutathione level that was restored by Apo. Increase in mRNA levels of a number of markers of hypertrophy, viz., ANP, BNP, ß-MHC, and ACTA-1 by Iso was either partially or completely prevented by Apo. Activation of key signaling kinases such as PKA, Erk, and Akt by Iso was also prevented by Apo treatment. Our study thus provided hemodynamic, biochemical, and molecular evidences supporting the therapeutic value of Apo in ameliorating adrenergic stress-induced cardiac hypertrophy.

Activation of adrenergic receptor in H9c2 cardiac myoblasts co-stimulates Nox2 and the derived ROS mediate the downstream responses.

In recent years, NADPH oxidases (Noxes) have emerged as an important player in cardiovascular pathophysiology. Despite the growing evidences on the role of specific Nox isoforms, mechanisms of their activation, targets of reactive oxygen species (ROS) generated, and their downstream effects are poorly understood as yet. In this study, we treated H9c2 cardiac myoblasts with norepinephrine (NE, 2 µM), inducing ROS generation that was inhibited by Nox2-specific peptide inhibitor gp91ds-tat. Organelle-specific hydrogen peroxide-sensitive probe HyPer showed that the site of ROS generation is primarily in the cytosol, to some extent in the endoplasmic reticulum (ER) but not the mitochondria. Modulation of mRNAs of marker genes of cardiac hypertrophy i.e. induction in ANP and ß-MHC, and reduction in α-MHC by NE treatment was prevented by specific inhibition of Nox2 by gp91ds-tat. Induction of ANP and ß-MHC at the protein level were also attenuated by the inhibition of Nox2. Induction of c-Jun and FosB, the two members of the transcription factor family AP-1, were also blocked by the inhibition of Nox2 by gp91ds-tat. Induction of promoter-reporter constructs harboring multiple AP-1 elements and the upstream of FosB and ANP genes by NE were also blocked by the inhibition of Nox2 by gp91ds-tat and a dominant negative mutant of p22phox, a constituent of Nox2 that prevents its activation. This study for the first time establishes the significant role of Nox2 in mediating the NE-induced pathological adrenergic signaling in cardiac myoblasts.

SG2NA enhances cancer cell survival by stabilizing DJ-1 and thus activating Akt.

SG2NA in association with striatin and zinedin forms a striatin family of WD-40 repeat proteins. This family of proteins functions as scaffold in different signal transduction pathways. They also act as a regulatory subunit of protein phosphatase 2A. We have shown that SG2NA which evolved first in the metazoan evolution among the striatin family members expresses different isoforms generated out of alternative splicing. We have also shown that SG2NA protects cells from oxidative stress by recruiting DJ-1 and Akt to mitochondria and membrane in the post-mitotic neuronal cells. DJ-1 is both cancer and Parkinson's disease related protein. In the present study we have shown that SG2NA protects DJ-1 from proteasomal degradation in cancer cells. Hence, downregulation of SG2NA reduces DJ-1/Akt colocalization in cancer cells resulting in the reduction of anchorage dependent and independent growth. Thus SG2NA enhances cancer cell survival. Reactive oxygen species enhances SG2NA, DJ-1 and Akt trimerization. Removal of the reactive oxygen species by N-acetyl-cysteine thus reduces cancer cell growth.

Novel role of a family of major facilitator transporters in biofilm development and virulence of Candida albicans.

The QDR (quinidine drug resistance) family of genes encodes transporters belonging to the MFS (major facilitator superfamily) of proteins. We show that QDR transporters, which are localized to the plasma membrane, do not play a role in drug transport. Hence, null mutants of QDR1, QDR2 and QDR3 display no alterations in susceptibility to azoles, polyenes, echinocandins, polyamines or quinolines, or to cell wall inhibitors and many other stresses. However, the deletion of QDR genes, individually or collectively, led to defects in biofilm architecture and thickness. Interestingly, QDR-lacking strains also displayed attenuated virulence, but the strongest effect was observed with qdr2∆, qdr3∆ and in qdr1/2/3∆ strains. Notably, the attenuated virulence and biofilm defects could be reversed upon reintegration of QDR genes. Transcripts profiling confirmed differential expression of many biofilm and virulence-related genes in the deletion strains as compared with wild-type Candida albicans cells. Furthermore, lipidomic analysis of QDR-deletion mutants suggests massive remodelling of lipids, which may affect cell signalling, leading to the defect in biofilm development and attenuation of virulence. In summary, the results of the present study show that QDR paralogues encoding MFS antiporters do not display conserved functional linkage as drug transporters and perform functions that significantly affect the virulence of C. albicans.

Cellular redox, epigenetics and diseases.

Since the Central dogma of Molecular Biology was proposed about 40 years ago; our understanding of the intricacies of gene regulation has undergone tectonic shifts almost every decade. It is now widely accepted that the complexity of an organism is not directed by the sheer number of genes it carries but how they are decoded by a myriad of regulatory modules. Over the years, it has emerged that the organizations chromatins and its remodeling; splicing and polyadenylation of pre-mRNAs, stability and localization of mRNAs and modulation of their expression by non-coding and miRNAs play pivotal roles in metazoan gene expression. Nevertheless, in spite of tremendous progress in our understanding of all these mechanisms of gene regulation, the way these events are coordinated leading towards a highly defined proteome of a given cell type remains enigmatic. In that context, the structures of many metazoan genes cannot fully explain their pattern of expression in different tissues, especially during embryonic development and progression of various diseases. Further, numerous studies done during the past quarter of a century suggested that the heritable states of transcriptional activation or repression of a gene can be influenced by the covalent modifications of constituent bases and associated histones; its chromosomal context and long-range interactions between various chromosomal elements (Holliday 1987; Turner 1998; Lyon 1993). However, molecular dissection of these phenomena is largely unknown and is an exciting topic of research under the sub-discipline epigenetics (Gasser et al. 1998).

Cholesterol depletion enhances adrenergic signaling in cardiac myocytes.

Cardiac myocytes endogenously express α and ß adrenergic receptors, prototypes of the G-protein coupled receptor superfamily. Depending upon the dose of norepinephrine (agonist) exposure, hypertrophy and apoptosis are initiated by differential induction of two discrete constituents of the transcription factor AP-1, i.e., FosB and Fra-1. We explored differential adrenergic signaling as a paradigm for understanding how cholesterol dictates cells to choose hypertrophy or apoptosis. For this, we used fosB and fra-1 promoter-reporter constructs for monitoring adrenergic signaling. We show that cholesterol depletion enhances norepinephrine-mediated signaling in cardiac myocytes. Importantly, this increased signaling is reduced to original level upon cholesterol replenishment. We used specific ligands for α and ß adrenergic receptors and show that the enhanced signaling upon cholesterol depletion is a combined effect of both α and ß adrenergic receptors. These results constitute the first report demonstrating the effect of cholesterol on adrenergic signaling using a direct end-point gene expression.

In cardiac muscle cells, both adrenergic agonists and antagonists induce reactive oxygen species from NOX2 but mutually attenuate each other's effects.

In cardiac muscle cells adrenergic agonists stimulate the generation of reactive oxygen species, followed by redox signaling. We postulated that the antagonists would attenuate such reactive oxygen species generation by the agonists. H9c2 cardiac myoblasts, neonatal rat cardiac myocytes, and HEK293 cells expressing ß1/ß2 adrenoceptors were stimulated with several agonists and antagonists. All the agonists and antagonists independently generated reactive oxygen species; but its generation was minimum whenever an agonists was added together with an antagonist. We monitored the Ca++ signaling in the treated cells and obtained similar results. In all treatment sets, superoxide and H2O2 were generated in the mitochondria and the cytosol respectively. NOX2 inhibitor gp91ds-tat blocked reactive oxygen species generation by both the agonists and the antagonists. The level of p47phox subunit of NOX2 rapidly increased upon treatment, and it translocated to the plasma membrane, confirming NOX2 activation. Inhibitor studies showed that the activation of NOX2 involves ERK, PI3K, and tyrosine kinases. Recombinant promoter-reporter assays showed that reactive oxygen species generated by both the agonists and antagonists modulated downstream gene expression. Mice injected with the ß-adrenergic agonist isoproterenol and fed with the antagonist metoprolol showed a robust induction of p47phox in the heart. We conclude that both the agonism and antagonism of adrenoceptors initiate redox signaling but when added together, they mutually counteract each other's effects. Our study thus highlights the importance of reactive oxygen species in adrenoceptor agonism and antagonism with relevance to the therapeutic use of the ß blockers.

Morphogenic regulator EFG1 affects the drug susceptibilities of pathogenic Candida albicans.

This study shows that the morphogenic regulator EFG1 level affects the drug susceptibilities of Candida albicans when grown on solid growth media. The Deltaefg1 mutant showed sensitivity particularly to those drugs that target ergosterol or its metabolism. Efg1p disruption showed a gene-dosage effect on drug susceptibilities and resulted in enhanced susceptibility to drugs in the homozygous mutant as compared with the wild type, heterozygous and revertant strains. The enhanced sensitivity to drugs was independent of the status of ATP-binding cassette and MFS multidrug efflux pumps of C. albicans. The Deltaefg1 mutant displayed increased membrane fluidity that coincided with the downregulation of ERG11 and upregulation of OLE1 and ERG3, leading to enhanced passive diffusion of drugs. Interestingly, Deltaefg1 mutant cells displayed enhanced levels of endogenous ROS levels. Notably, the higher levels of ROS in the Deltaefg1 mutant could be reversed by the addition of antioxidants. However, the restoration of ROS levels did not reverse the drug sensitivities of the Deltaefg1 mutant. Taken together, we, for the first time, establish a new role to EFG1 in affecting the drug susceptibilities of C. albicans cells, independent of ROS and known drug efflux mechanisms.

Transcriptomic Validation of the Protective Effects of Aqueous Bark Extract of Terminalia arjuna (Roxb.) on Isoproterenol-Induced Cardiac Hypertrophy in Rats.

Aqueous extract of the bark of Terminalia arjuna (TA) is used by a large population in the Indian subcontinent for treating various cardiovascular conditions. Animal experiments have shown its anti-atherogenic, anti-hypertensive, and anti-inflammatory effects. It has several bioactive ingredients with hemodynamic, ROS scavenging, and anti-inflammatory properties. Earlier we have done limited proteomic and transcriptomic analysis to show its efficacy in ameliorating cardiac hypertrophy induced by isoproterenol (ISO) in rats. In the present study we have used high-throughput sequencing of the mRNA from control and treated rat heart to further establish its efficacy. ISO (5 mg/kg/day s.c.) was administered in male adult rats for 14 days to induce cardiac hypertrophy. Standardized aqueous extract TA bark extract was administered orally. Total RNA were isolated from control, ISO, ISO + TA, and TA treated rat hearts and subjected to high throughput sequence analysis. The modulations of the transcript levels were then subjected to bio-informatics analyses using established software. Treatment with ISO downregulated 1,129 genes and upregulated 204 others. Pre-treatment with the TA bark extracts markedly restored that expression pattern with only 97 genes upregulated and 85 genes downregulated. The TA alone group had only 88 upregulated and 26 downregulated genes. The overall profile of expression in ISO + TA and TA alone groups closely matched with the control group. The genes that were modulated included those involved in metabolism, activation of receptors and cell signaling, and cardiovascular and other diseases. Networks associated with those genes included those involved in angiogenesis, extracellular matrix organization, integrin binding, inflammation, drug metabolism, redox metabolism, oxidative phosphorylation, and organization of myofibril. Overlaying of the networks in ISO and ISO_TA group showed that those activated in ISO group were mostly absent in ISO_TA and TA group, suggesting a global effect of the TA extracts. This study for the first time reveals that TA partially or completely restores the gene regulatory network perturbed by ISO treatment in rat heart; signifying its efficacy in checking ISO-induced cardiac hypertrophy.

WD-40 repeat protein SG2NA has multiple splice variants with tissue restricted and growth responsive properties.

SG2NA is a member of the striatin family of WD-40 repeat proteins with potential scaffolding functions. It was originally identified as a tumor antigen with increased expression during S to G2 phase of cell cycle. We report here that mouse SG2NA has at least five novel splice variants of which two are devoid of the carboxyl terminal WD-40 repeats. The variants of SG2NA are generated by alternative splicing at the exon 7-9 regions and differ in their expression profiles in various tissues tested. While the 83, 78, 38 and 35 kDa variants are present in both brain and heart, the 87 kDa form is brain specific. Also, the expression of 35 kDa variant is more in neonatal than in adult tissues. Western analysis suggests that the SG2NA isoforms differentially respond to growth stimuli. Upon serum stimulation, while the 35 kDa variant is increased, the 78 kDa form is diminished. Splicing variation of SG2NA is conserved in metazoan evolution. In embryonic chicken there are at least four variants of which one is present in brain but absent in heart. Taken together, splicing variation of SG2NA might have some critical roles in differentiation and maturation in metazoan cells.

Regulation of ceruloplasmin in human hepatic cells by redox active copper: identification of a novel AP-1 site in the ceruloplasmin gene.

Cp (ceruloplasmin), a copper containing plasma protein, mainly synthesized in the liver, is known to be functional between the interface of iron and copper metabolism. We have reported previously that Cp is regulated by cellular iron status, but the process of the regulation of Cp by copper still remains a subject for investigation. In the present paper, we show that PDTC (pyrrolidine dithiocarbamate), a thiol compound widely known to increase intracellular redox copper, regulates Cp expression in hepatic cells by a copper-dependent transcriptional mechanism. To find out the mechanism of induction, chimeric constructs of the Cp 5'-flanking region driving luciferase were transfected into human hepatic cells. Deletion and mutational analyses showed the requirement of a novel APRE [AP-1 (activator protein-1) responsive element] present about 3.7 kb upstream of the translation initiation site. The role of AP-1 was confirmed by electrophoretic mobility-shift analysis. Western blot and overexpression studies detected the AP-1 as a heterodimer of c-jun and c-fos proteins. The activation of AP-1 was found to be copper-dependent as a specific extracellular chelator bathocuproine disulfonic acid blocked PDTC-mediated induction of AP-1-DNA binding and increased reporter gene activity. Whereas, in a copper-free medium, PDTC failed to activate either AP-1 or Cp synthesis, supplementation of copper could reverse AP-1 activation and Cp synthesis. Our finding is not only the first demonstration of regulation of Cp by redox copper but may also explain previous findings of increased Cp expression in cancers like hepatocarcinoma, where the intracellular copper level is higher in a redox compromised environment.

Author Correction: GSK3ß and ERK regulate the expression of 78 kDa SG2NA and ectopic modulation of its level affects phases of cell cycle.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has been fixed in the paper.

Biophysical Characterization of SG2NA Variants and their Interaction with DJ-1 and Calmodulin in vitro.

SG2NA was first discovered as nuclear autoantigen in lung and bladder cancer patient. It was named SG2NA as its expression increases during S to G2 phase of cell cycle. SG2NA/Striatin3 was classified as a member of Striatin family along with Straitin and Zinedin due to its structural and functional relatedness. At the molecular level, SG2NA is characterized by the presence of multiple protein-protein interaction domains viz., a caveolin binding motif, a coiled coil structure, Ca2+-calmodulin binding domain and a large WD-40 repeat domain in the same order from amino to the carboxyl termini. Analysis of secondary structures of 87 and 78 kDa SG2NA isoforms showed characteristic combinations of α-helix, ß-structure, ß-turns and random coil; suggesting of effective refolding after denaturation. This study for the first time establishes the structural differences between the two prevalent isoforms of SG2NA. Recently we observed that DJ-1 interacts with variants of SG2NA both in vitro and in vivo. The SG2NA isoforms purified from inclusion bodies showed the different secondary structure conformations, stability and interaction pattern for their interacting partners (DJ-1 and calmodulin) which imparts functional diversity of SG2NA. The SG2NA isoforms showed significant differential binding affinity to DJ-1 and Calmodulin.