Publications by authors named "Nasrin Mesaeli"

12 Publications

  • Page 1 of 1

Endoplasmic reticulum stress enhances endocytosis in calreticulin deficient cells.

Biochim Biophys Acta Mol Cell Res 2019 04 5;1866(4):727-736. Epub 2018 Dec 5.

Department of Biochemistry, Weill Cornell Medicine in Qatar, Doha, Qatar. Electronic address:

Calreticulin an endoplasmic reticulum (ER) chaperone that is involved in the quality control process and plays an important role as a regulator of intracellular calcium homeostasis. Previously, we illustrated that loss of calreticulin (crt-/-) results in the activation of ubiquitin-proteasome pathway facilitating the increased resistance to apoptosis. Our preliminary data illustrated a significant increase in the endocytosis in the calreticulin knockout mouse embryonic fibroblast cells (crt-/-). Therefore, we hypothesized that the mechanism for this increased endocytosis in the crt-/- cells is due to onset of ER stress. To test this hypothesis, we measured endocytosis in the wild type (wt) and crt-/- cells using uptake of fluorescent dextran and showed a significant increase in the rate of its uptake in crt-/- cells as compared to wt cells. To determine the endocytic pathway involved we examined both clathrin and caveolin-1 dependent endocytosis. Our results illustrated no change in the expression of clathrin heavy chain while there was a significant increase in the expression of caveolin-1 in the crt-/- cells as compared to the wt cells. Furthermore, using shRNA we illustrated that knockdown of clathrin heavy chain had no effect on endocytosis in the crt-/- cells. While knock-down of caveolin-1 significantly reduced endocytosis in the crt-/- cells. Finally, we illustrated that a chemical chaperone, 4‑phenylbutyrate significantly reduced both the endoplasmic reticulum stress and endocytosis in the crt-/- cells. Our data shows for the first time, that ER stress led to enhanced caveolin-1 mediated endocytosis and reversal of ER stress reduces endocytosis.
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http://dx.doi.org/10.1016/j.bbamcr.2018.12.003DOI Listing
April 2019

Fourier-Transform Infrared Imaging Spectroscopy and Laser Ablation -ICPMS New Vistas for Biochemical Analyses of Ischemic Stroke in Rat Brain.

Front Neurosci 2018 19;12:647. Epub 2018 Sep 19.

Department of Chemistry and Earth Sciences, Qatar University, Doha, Qatar.

Stroke is the main cause of adult disability in the world, leaving more than half of the patients dependent on daily assistance. Understanding the post-stroke biochemical and molecular changes are critical for patient survival and stroke management. The aim of this work was to investigate the photo-thrombotic ischemic stroke in male rats with particular focus on biochemical and elemental changes in the primary stroke lesion in the somatosensory cortex and surrounding areas, including the corpus callosum. FT-IR imaging spectroscopy and LA-ICPMS techniques examined stroke brain samples, which were compared with standard immunohistochemistry studies. The FTIR results revealed that in the lesioned gray matter the relative distribution of lipid, lipid acyl and protein contents decreased significantly. Also at this locus, there was a significant increase in aggregated protein as detected by high-levels Aβ. Areas close to the stroke focus experienced decrease in the lipid and lipid acyl contents associated with an increase in lipid ester, olefin, and methyl bio-contents with a novel finding of Aβ in the PL-GM and L-WM. Elemental analyses realized major changes in the different brain structures that may underscore functionality. In conclusion, FTIR bio-spectroscopy is a non-destructive, rapid, and a refined technique to characterize oxidative stress markers associated with lipid degradation and protein denaturation not characterized by routine approaches. This technique may expedite research into stroke and offer new approaches for neurodegenerative disorders. The results suggest that a good therapeutic strategy should include a mechanism that provides protective effect from brain swelling (edema) and neurotoxicity by scavenging the lipid peroxidation end products.
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http://dx.doi.org/10.3389/fnins.2018.00647DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6157330PMC
September 2018

SIRT1 Limits Adipocyte Hyperplasia through c-Myc Inhibition.

J Biol Chem 2016 Jan 11;291(5):2119-35. Epub 2015 Dec 11.

From the Departments of Microbiology and Immunology,

The expansion of fat mass in the obese state is due to increased adipocyte hypertrophy and hyperplasia. The molecular mechanism that drives adipocyte hyperplasia remains unknown. The NAD(+)-dependent protein deacetylase sirtuin 1 (SIRT1), a key regulator of mammalian metabolism, maintains proper metabolic functions in many tissues, counteracting obesity. Here we report that differentiated adipocytes are hyperplastic when SIRT1 is knocked down stably in mouse 3T3-L1 preadipocytes. This phenotype is associated with dysregulated adipocyte metabolism and enhanced inflammation. We also demonstrate that SIRT1 is a key regulator of proliferation in preadipocytes. Quantitative proteomics reveal that the c-Myc pathway is altered to drive enhanced proliferation in SIRT1-silenced 3T3-L1 cells. Moreover, c-Myc is hyperacetylated, levels of p27 are reduced, and cyclin-dependent kinase 2 (CDK2) is activated upon SIRT1 reduction. Remarkably, differentiating SIRT1-silenced preadipocytes exhibit enhanced mitotic clonal expansion accompanied by reduced levels of p27 as well as elevated levels of CCAAT/enhancer-binding protein β (C/EBPβ) and c-Myc, which is also hyperacetylated. c-Myc activation and enhanced proliferation phenotype are also found to be SIRT1-dependent in proliferating mouse embryonic fibroblasts and differentiating human SW872 preadipocytes. Reducing both SIRT1 and c-Myc expression in 3T3-L1 cells simultaneously does not induce the adipocyte hyperplasia phenotype, confirming that SIRT1 controls adipocyte hyperplasia through c-Myc regulation. A better understanding of the molecular mechanisms of adipocyte hyperplasia will open new avenues toward understanding obesity.
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http://dx.doi.org/10.1074/jbc.M115.675645DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4732199PMC
January 2016

Loss of calreticulin function decreases NFκB activity by stabilizing IκB protein.

Biochim Biophys Acta 2014 Nov 3;1843(11):2385-93. Epub 2014 Jul 3.

Department of Biochemistry and Medical Genetics, University of Manitoba, Winnipeg, Manitoba, Canada; Department of Biochemistry, Weill Cornell Medical College Qatar, Doha, Qatar. Electronic address:

Transcription factor NFκB is activated by several processes including inflammation, endoplasmic-reticulum (ER) stress, increase in Akt signaling and enhanced proteasomal degradation. Calreticulin (CRT) is an ER Ca(2+)-binding chaperone that regulates many cellular processes. Gene-targeted deletion of CRT has been shown to induce ER stress that is accompanied with a significant increase in the proteasome activity. Loss of CRT function increases the resistance of CRT-deficient (crt-/-) cells to UV- and drug-induced apoptosis. Based on these reports we hypothesized that loss of CRT will activate NFκB signaling thus contributing to enhanced resistance to apoptosis. In contrast to our hypothesis, we observed a significant decrease in the basal transcriptional activity of NFκB in CRT-deficient cells. Treatment with lipopolysaccharide failed to increase the transcriptional activity of NFκB in the crt-/- cells to the same level as in the wt cells. Our data illustrate that the mechanism of decreased NFκB activity in CRT-deficient cells is mediated by a significant increase in IκB protein expression. Furthermore, we showed a significant increase in protein phosphatase 2A activity inhibition which resulted in decreased IκBα protein level in CRT-deficient cells. Based on our data we concluded that loss of CRT increases the stability of IκB protein thus reducing NFκB activity.
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http://dx.doi.org/10.1016/j.bbamcr.2014.06.017DOI Listing
November 2014

Extracellular K+ concentration controls cell surface density of IKr in rabbit hearts and of the HERG channel in human cell lines.

J Clin Invest 2009 Sep 24;119(9):2745-57. Epub 2009 Aug 24.

Department of Physiology, Queen's University, Kingston, Ontario, Canada.

Although the modulation of ion channel gating by hormones and drugs has been extensively studied, much less is known about how cell surface ion channel expression levels are regulated. Here, we demonstrate that the cell surface density of both the heterologously expressed K+ channel encoded by the human ether-a-go-go-related gene (HERG) and its native counterpart, the rapidly activating delayed rectifier K+ channel (IKr), in rabbit hearts in vivo is precisely controlled by extracellular K+ concentration ([K+]o) within a physiologically relevant range. Reduction of [K+]o led to accelerated internalization and degradation of HERG channels within hours. Confocal analysis revealed colocalization between HERG and ubiquitin during the process of HERG internalization, and overexpression of ubiquitin facilitated HERG degradation under low [K+]o. The HERG channels colocalized with a marker of multivesicular bodies during internalization, and the internalized HERG channels were targeted to lysosomes. Our results provide the first evidence to our knowledge that the cell surface density of a voltage-gated K+ channel, HERG, is regulated by a biological factor, extracellular K+. Because hypokalemia is known to exacerbate long QT syndrome (LQTS) and Torsades de pointes tachyarrhythmias, our findings provide a potential mechanistic link between hypokalemia and LQTS.
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http://dx.doi.org/10.1172/JCI39027DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2735921PMC
September 2009

Calreticulin regulates insulin receptor expression and its downstream PI3 Kinase/Akt signalling pathway.

Biochim Biophys Acta 2008 Dec 17;1783(12):2344-51. Epub 2008 Sep 17.

Institute of Cardiovascular Sciences, St. Boniface General Hospital Research Centre 351 Tache Avenue, Winnipeg, Manitoba, Canada R2H 2A6.

Defects in insulin signalling and glucose metabolism are associated with the development of diabetes. Insulin signalling is initiated by the binding of insulin to its receptor and triggering cascades of events including activation of PI3kinase/Akt signalling pathway. Calreticulin (CRT) is a calcium binding chaperone molecule located in the endoplasmic reticulum. Targeted deletion of CRT in mice is embryonic lethal as a result of developmental and metabolic abnormalities. Rescued CRT null mice develop severe hypoglycemia the reason for which is not known. In addition, ventricular cardiomyocytes isolated from CRT null (crt-/-) mice have increased glycogen deposits. Therefore, the aim of this study was to investigate the changes in the glucose uptake and insulin signalling pathway (mainly PI3 kinase/Akt) in the absence of CRT. Here we show a significant increase in the glucose uptake by the crt-/- cells. This increase was accompanied by a significant increase in both insulin receptor beta expression, Insulin receptor substrate-1 phosphorylation, GLUT-1 expression and in insulin stimulated Akt phosphorylation and kinase activity in the crt-/- cells. Intriguingly, the increased expression of insulin receptor beta in the crt-/- was due to decreased levels of p53 protein. The current study is the first evidence for the up-regulation of insulin receptor density and activity in the absence of CRT function.
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http://dx.doi.org/10.1016/j.bbamcr.2008.08.014DOI Listing
December 2008

Enhanced ubiquitin-proteasome activity in calreticulin deficient cells: a compensatory mechanism for cell survival.

Biochim Biophys Acta 2008 Jun 20;1783(6):1237-47. Epub 2008 Mar 20.

Institute of Cardiovascular Sciences, St. Boniface General Hospital Research Centre, Winnipeg, Canada.

Calreticulin is a lectin chaperone essential for intracellular calcium homeostasis. Deletion of calreticulin gene compromises the overall quality control within the endoplasmic reticulum (ER) leading to activation of the unfolded protein response. However, the ER structure of calreticulin deficient cells (crt-/-) is not altered due to accumulation of misfolded proteins. Therefore, the aim of this study was to determine whether the ubiquitin-proteasome pathway is activated in crt-/- cells as a compensatory mechanism for cell survival. Here we show a significant increase in the expression of genes involved in ER associated degradation and activation of the ubiquitin-proteasome system in crt-/- cells. We also demonstrated that the ubiquitination of two proteins processed in ER, connexin 43 and A1AT NHK (alpha1-antitrypsin mutant) are increased in crt-/- cells. Furthermore, we showed that the increased proteasome activity in the crt-/- cells could be rescued upon re-introduction of calreticulin or calsequestrin (a muscle calcium binding protein). We also illustrated that increased cytosolic Ca2+ enhances the proteasome activity. Interestingly, suppression of calnexin function using siRNA further elevated the proteasome activity in crt-/- cells. This is the first report to show that loss of calreticulin function enhances the ubiquitin-proteasome activity which could function as a compensatory mechanism for cell survival.
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http://dx.doi.org/10.1016/j.bbamcr.2008.03.004DOI Listing
June 2008

Differential expression and activity of matrix metalloproteinase-2 and -9 in the calreticulin deficient cells.

Matrix Biol 2007 Jul 6;26(6):463-72. Epub 2007 Mar 6.

Division of Stroke and Vascular Disease, St. Boniface General Hospital Research Centre, Department of Biochemistry and Medical Genetics, Faculty of Medicine, University of Manitoba, Winnipeg, Canada.

Calreticulin is an endoplasmic reticulum protein important in cardiovascular development. Deletion of the calreticulin gene leads to defects in the heart and the formation of omphaloceal. These defects could both be due to changes in the extracellular matrix composition. Matrix metalloproteinases (MMP)-2 and MMP-9 are two of the MMPs which are essential for cardiovascular remodelling and development. Here, we tested the hypothesis that the defects observed in the heart and body wall of the calreticulin null embryos are due to alterations in MMP-2 and MMP-9 activity. Our results demonstrate that there is a significant decrease in the MMP-9 and increase in the MMP-2 activity and expression in the calreticulin deficient cells. We also showed that there is a significant increase in the expression level of membrane type-1 matrix metalloproteinase (MT1-MMP). In contrast, there was no change in the tissue inhibitor of matrix metalloproteinase (TIMP)-1 or -2 in the calreticulin deficient cells as compared to the wild type cells. Interestingly, the inhibition of the MEK kinase pathway using PD98059 attenuated the decrease in the MMP-9 mRNA with no effect on the MMP-2 mRNA level in the calreticulin deficient cells. Furthermore, PI3 kinase inhibitor decreased the expression of both the MMP-2 and MMP-9. This study is the first report on the role of calreticulin in regulating MMP activity.
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http://dx.doi.org/10.1016/j.matbio.2007.02.005DOI Listing
July 2007

Upregulation of the alpha1-adrenoceptor-induced phosphoinositide and inotropic response in hypothyroid rat heart.

Mol Cell Biochem 2006 Feb;283(1-2):93-100

Division of Stroke and Vascular Disease, St. Boniface General Hospital Research Centre, Winnipeg, Canada.

In this study, we examined changes in the biochemical and inotropic events of the alpha(1)-adrenoceptor signaling pathway in hypothyroid rat hearts. Hypothyroidism was induced by treating experimental animals with 0.05% 6-n-propyl-2-thiouracil (PTU) in drinking water for 7 weeks. A significant decrease of beta- and an increase in alpha(1)-adrenoceptor density as well as an increase in the basal activity of the phosphoinositide (4,5) bisphosphate hydrolyzing phospholipase C was observed in sarcolemmal membranes purified from hypothyroid hearts as compared to age-matched euthyroid controls. Following stimulation with 10 microM phenylephrine (in the presence of 10 microM atenolol), the increase of contractile parameters over baseline values was significantly higher in hypo- than euthyroid hearts, while the opposite occurred under beta-stimulation with 0.1 microM isoproterenol. Interestingly, the increase in phenylephrine-mediated positive inotropy was accompanied by a significant increase in the sarcolemmal phospholipase C activity and in the inositol 1,4,5-trisphosphate content in hypothyroid as compared to euthyroid controls. Our results suggest that cardiac alpha(1)-adrenoceptor and its associated phosphoinositide signaling pathway may act as a reserve for catecholamine inotropic response in hypothyroidism, where the beta-adrenoceptors are compromised.
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http://dx.doi.org/10.1007/s11010-006-2348-9DOI Listing
February 2006

Cells expressing unique Na+/Ca2+ exchange (NCX1) splice variants exhibit different susceptibilities to Ca2+ overload.

Am J Physiol Heart Circ Physiol 2006 May 6;290(5):H2155-62. Epub 2006 Jan 6.

Division of Stroke and Vascular Disease, St. Boniface Hospital Research Centre, Winnipeg, MB, Canada R2H 2A6.

The Na+/Ca2+ exchanger (NCX) NCX1 exhibits tissue-specific alternative splicing. Such NCX splice variants as NCX1.1 and NCX1.3 are also differentially regulated by Na+ and Ca2+, although the physiological implications of these regulatory characteristics are unclear. On the basis of their distinct regulatory profiles, we hypothesized that cells expressing these different splice variants might exhibit unique responses to conditions promoting Ca2+ overload, such as during exposure to cardiac glycosides or simulated ischemia. NCX1.1 or NCX1.3 was expressed in human embryonic kidney (HEK)-293 cells or rat neonatal ventricular cardiomyocytes (NVC), and expression was confirmed by Western blotting and immunocytochemical analyses. HEK-293 cells lacked NCX1 protein before transfection. With use of adenoviral vectors, neonatal cardiomyocytes were induced to overexpress the NCX1.1 splice variant by nearly twofold, whereas the NCX1.3 isoform was expressed on the endogenous NCX1.1 background. Total expression was comparable for NCX1.1 and NCX1.3. Exposure of NVC to ouabain induced a significant increase in cellular Ca2+, an effect that was exaggerated in cells overexpressing NCX1.1, but not NCX1.3. The increase in intracellular Ca2+ was inhibited by 5 microM KB-R7943. Cardiomyocytes overexpressing NCX1.1 also exhibited a greater accumulation of intracellular Ca2+ in response to simulated ischemia than did cells expressing NCX1.3. Similar responses were observed in HEK-293 cells where NCX1.1 was expressed. We conclude that expression of the NCX1.3 splice variant protects against severe Ca2+ overload, whereas NCX1.1 promotes Ca2+ overload in response to cardiac glycosides and ischemic challenges. These results highlight the importance of ionic regulation in controlling NCX1 activity under conditions that promote Ca2+ overload.
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http://dx.doi.org/10.1152/ajpheart.00958.2005DOI Listing
May 2006

Impaired p53 expression, function, and nuclear localization in calreticulin-deficient cells.

Mol Biol Cell 2004 Apr 6;15(4):1862-70. Epub 2004 Feb 6.

Division of Stroke and Vascular Disease, St. Boniface General Hospital Research Centre, University of Manitoba, Winnipeg, Canada R2H 2A6.

The tumor suppressor protein, p53 is a transcription factor that not only activates expression of genes containing the p53 binding site but also can repress the expression of some genes lacking this binding site. Previous studies have shown that overexpression of wild-type p53 leads to apoptosis and cell cycle arrest. DNA damage, such as that caused by UV irradiation, results in p53 stabilization and nuclear localization that subsequently induces apoptosis. Recently, the level of calreticulin (CRT) has been correlated with the rate of apoptosis. Therefore, the aim of this study was to investigate the role of CRT in the regulation of apoptosis via modulating p53 function and expression. Here we show a significant decrease in both basal and DNA damage induced p53 functions in the CRT-deficient cells (crt-/-). This study is the first to demonstrate that CRT function is required for the stability and localization of the p53 protein. By using immuonocytochemical techniques, we showed that observed changes in p53 in the crt-/- cells are due to the nuclear accumulation of Mdm2 (murine double minute gene). These results, lead us to conclude that CRT regulates p53 function by affecting its rate of degradation and nuclear localization.
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http://dx.doi.org/10.1091/mbc.e03-04-0251DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC379282PMC
April 2004

Compromised calnexin function in calreticulin-deficient cells.

Biochem Biophys Res Commun 2003 May;304(4):661-6

Department of Biochemistry, Canadian Institutes of Health Research Membrane Protein Research Group, University of Alberta, Edmonton, Alberta, Canada T6G 2H7.

Calnexin and calreticulin are molecular chaperones, which are involved in the protein folding, assembly, and retention/retrieval. We know that calreticulin-deficiency is lethal in utero, but do not understand the contribution of chaperone function to this phenotype. Here we studied protein folding and chaperone function of calnexin in the absence of calreticulin. We show that protein folding is accelerated and quality control is compromised in calreticulin-deficient cells. Calnexin-substrate association is severely reduced, leading to accumulation of unfolded proteins and a triggering of the unfolded protein response (UPR). PERK and Ire1alpha and eIF2alpha are also activated in calreticulin-deficient cells. We show that the absence of calreticulin can have devastating effects on the function of the others, compromising overall quality control of the secretory pathway and activating UPR-dependent pathways.
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http://dx.doi.org/10.1016/s0006-291x(03)00643-0DOI Listing
May 2003
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