DOCSLIB.ORG

Karyopherin Enrichment at the Nuclear Pore Complex Attenuates Ran Permeability Suncica Barbato*, Larisa E

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Karyopherin Enrichment at the Nuclear Pore Complex Attenuates Ran Permeability Suncica Barbato*, Larisa E © 2020. Published by The Company of Biologists Ltd | Journal of Cell Science (2020) 133, jcs238121. doi:10.1242/jcs.238121 SHORT REPORT Karyopherin enrichment at the nuclear pore complex attenuates Ran permeability Suncica Barbato*, Larisa E. Kapinos*, Chantal Rencurel and Roderick Y. H. Lim‡ ABSTRACT of it. Selective Kap transport is underpinned by multivalent Ran is a small GTPase whose nucleotide-bound forms cycle through interactions with numerous phenylalanine-glycine (FG)-repeat-rich, nuclear pore complexes (NPCs) to direct nucleocytoplasmic transport intrinsically disordered nucleoporins (FG Nups) that line the NPC (NCT). Generally, Ran guanosine triphosphate (RanGTP) binds cargo- channel (Sakiyama et al., 2016). For instance, the classical 97 kDa β carrying karyopherin receptors (Kaps) in the nucleus and releases them import receptor karyopherin subunit 1 (KPNB1, hereafter referred to β into the cytoplasm following hydrolysis to Ran guanosine diphosphate as Kap 1) (Cingolani et al., 1999) engages up to ten FG repeats (RanGDP). This generates a remarkably steep Ran gradient across the (Bayliss et al., 2000, 2002; Bednenko et al., 2003; Isgro and Schulten, nuclear envelope that sustains compartment-specific cargo delivery 2005). Otherwise, the FG Nups are considered to adopt barrier-like and accumulation. However, because NPCs are permeable to small characteristics, such as polymer brushes (Lim et al., 2007; Rout et al., molecules of comparable size, it is unclear how an uncontrolled mixing 2000), gel-like meshworks (Frey and Görlich, 2007; Labokha et al., of RanGTP and RanGDP is prevented. Here, we find that an NPC- 2013) or variations of these (Yamada et al., 2010). Still, Kap-cargo enriched pool of karyopherin subunit beta 1 (KPNB1, hereafter referred complexes are considerably larger than the non-specific size cut-off. β to as Kapβ1) selectively mediates Ran diffusion across the pore but not Furthermore, Kap 1 recruits adaptor proteins from the karyopherin α α passive molecules of similar size (e.g. GFP). This is due to RanGTP subunit family (KPNA, hereafter referred to as Kap ) that bind having a stronger binding interaction with Kapβ1 than RanGDP. For this directly to a diverse repertoire of NLS-cargos such as transcription reason, the RanGDP importer, nuclear transport factor 2, facilitates the factors (Pumroy and Cingolani, 2015; Xu and Massagué, 2004). return of RanGDP into the nucleus following GTP hydrolysis. Hence, our understanding of how NPCs reconcile physical size Accordingly, the enrichment of Kapβ1 at NPCs may function as a exclusion with biochemical selectivity to mediate NCT remains retention mechanism that preserves the sharp transition of RanGTP incomplete. and RanGDP in the nucleus and cytoplasm, respectively. One peculiarity concerns Ran (Melchior et al., 1993; Moore and Blobel, 1993), which controls the site of cargo release, accumulation KEY WORDS: RanGTP, Karyopherin, Nucleocytoplasmic transport, and recycling of Kaps to underpin NCT directionality across Nuclear pore complex the nuclear envelope (Weis, 2003). This is sustained by the interconversion of its two nucleotide-bound forms, RanGTP and INTRODUCTION RanGDP, which are localized to the nucleus and cytoplasm, Nucleocytoplasmic transport (NCT) describes the selective exchange respectively (Görlich et al., 1996). With a molecular mass of of macromolecules between the nucleus and cytoplasm in eukaryotes 25 kDa, Ran is below the NPC size limit for non-specific (Görlich and Kutay, 1999). This is mediated by conduits of 50–60 nm molecules. Also, neither RanGDP nor RanGTP interact with the FG diameter within the nuclear envelope, known as nuclear pore repeats (Rexach and Blobel, 1995). Yet, the concentration of RanGTP complexes (NPCs) (Eibauer et al., 2015; Kim et al., 2018; von is estimated to be at least 200 times higher in the nucleus than in the Appen et al., 2015). Given their considerable size, NPCs are cytoplasm (Görlich et al., 2003; Kalab et al., 2002; Smith et al., 2002). permeable to passive molecules below ∼40 kDa, whereas larger non- Thus, how an uncontrolled mixing of RanGTP and RanGDP is specific macromolecules are generally withheld (Popken et al., 2015; prevented at NPCs remains unknown. Importantly, a disruption in the Timney et al., 2016). Meanwhile, three main groups of protein are Ran gradient results in the loss of NCT directionality (Nachury and selectively trafficked across the NPC central channel to sustain NCT. Weis, 1999) and has been linked to apoptosis (Wong et al., 2009), These are transport receptors known as karyopherins (Kaps), signal- hyperosmotic stress (Kelley and Paschal, 2007) and disease specific cargos and the Ran GTPase that harmonizes the process. (Eftekharzadeh et al., 2018). Aprioriexclusive NPC access is reserved for Kaps (Kimura and In the nucleus, RanGTP binds Kapβ1 to disassemble NLS-cargo– Imamoto, 2014; Tran et al., 2007). These include importins that Kapα–Kapβ1 complexes (Chi et al., 1996; Görlich et al., 1996; deliver cargos bearing nuclear localization signals (NLS) (Boulikas, Rexach and Blobel, 1995). This serves to facilitate the nuclear 1994; Cokol et al., 2000) into the nucleus, and exportins that usher retention of NLS-cargos whose return to the cytoplasm is hindered in cargos containing nuclear export signals (NES) (Xu et al., 2012) out the absence of FG Nup binding. On the other hand, RanGTP–Kapβ1 retains its interactions with the FG Nups to return through NPCs (Kapinos et al., 2017). At the cytoplasmic periphery, RanGTP is Biozentrum & The Swiss Nanoscience Institute, University of Basel, 4056 Basel, hydrolyzed to RanGDP by RanGTPase-activating protein 1 Switzerland. *These authors contributed equally to this work (RanGAP1) together with the Ran-binding proteins RanBP1 and RanBP2 (Lounsbury and Macara, 1997; Vetter et al., 1999). This ‡ Author for correspondence ([email protected]) frees Kapβ1, which is then able to seek out the next NLS-cargo. Still, R.Y.H.L., 0000-0001-5015-6087 Ran seems to accumulate at NPCs (Abu-Arish et al., 2009; Smith et al., 2002; Wong et al., 2009; Yang and Musser, 2006), suggesting Received 19 August 2019; Accepted 13 December 2019 that it does not freely diffuse through the NPC like other non-specific Journal of Cell Science 1 SHORT REPORT Journal of Cell Science (2020) 133, jcs238121. doi:10.1242/jcs.238121 molecules of similar size (Timney et al., 2016). RanGDP then recruits against samples re-populated with exoKapα–Kapβ1 (20 µM:10 µM) a dedicated import factor, i.e. nuclear transport factor 2 (NUTF2, (Fig. 2A). Unexpectedly, exoRanGDP retention was the lowest in hereafter referred to as NTF2) (Ribbeck et al., 1998; Smith et al., nuclei harboring exoKapα–Kapβ1, i.e. 25% less than in control 1998), which returns RanGDP to the nucleus. Upon re-entry, the samples that lacked exoKaps (Fig. 2B,C). In comparison, nuclei that chromatin-bound enzyme regulator of chromosome condensation 1 contained exoKapβ1 alone showed 20% more exoRanGDP retention (RCC1, also referred to as Ran guanine nucleotide exchange factor or than control cells. This signified that exoKapα–Kapβ1 does not RanGEF) (Klebe et al., 1995b; Renault et al., 2001) recharges impede exoRanGDP outflow at the NPC as effectively as exoKapβ1 RanGDP to RanGTP to complete the cycle. In this manner, NCT alone. Nevertheless, we did obtain exoRanGDP fluorescence at the cargo delivery and the recycling of Kaps are regulated by RanGAP1 nuclear envelope of control cells, which suggested residual binding and RanGEF as well as the controlled exchange of RanGTP and with either endogenous transport receptors or other NPC components RanGDP across NPCs (Abu-Arish et al., 2009; Izaurralde et al., 1997; (Görlich et al., 1996; Partridge and Schwartz, 2009; Schrader et al., Kalab et al., 2006, 2002). 2008). Following these observations, we rationalized that NPCs More recently, a steady-state enrichment of Kapβ1 was uncovered might effectively impede exoRanGDP movement based on its in FG Nup layers (Kapinos et al., 2014; Schoch et al., 2012; Vovk biochemical interactions with exoKapβ1. et al., 2016; Wagner et al., 2015; Zahn et al., 2016) and in NPCs Subsequently, binding affinity measurements between exoKapβ1 (Görlich et al., 1995; Kapinos et al., 2017; Lowe et al., 2015). We also and either exoRanGDP or a GDP-bound, non-hydrolyzable Ran found that depleting this Kapβ1 pool abolishes NPC barrier function mutant comprising a Gln69 to Leu point mutation (RanQ69L-GDP) against large non-specific cargos (see Kapinos et al., 2017), which resulted in Kd values of 0.5±0.07 µM or 1.3±0.15 µM, respectively suggests that Kaps serve as bona fide constituents of the NPC barrier (Fig. S1D), consistent with literature values (Forwood et al., 2008; mechanism. This motivated our present study, in which we show that Lonhienne et al., 2009). In comparison, binding between Kapα and NPC-bound Kapβ1 selectively retains RanGTP and RanGDP at the Kapβ1isstronger(Kd=0.2 µM) (Bednenko et al., 2003; Catimel et al., NPC but not passive molecules of similar size (e.g. GFP). This is due 2001; Kapinos et al., 2017). For this reason, exoRanGDP is unable to to binding interactions with Kapβ1 at the pore, which are stronger for outcompete exoKapα for exoKapβ1. Indeed, the nuclear and nuclear RanGTP and weaker for RanGDP. In comparison, NPCs that lack envelope exoRanGDP signals are at least ∼30% higher for exoKapβ1 Kaps show unrestricted Ran movement, i.e. a Ran ‘leak’. Therefore, than exoKapα·Kapβ1 (Fig. 2B,C). Therefore, we reasoned that RanGTP outflow depends on the hydrolysis of RanGTP to RanGDP, exoKapβ1 in the NPC might serve to restrict exoRanGDP outflow. By whereas RanGDP import requires NTF2. These results explain how contrast, a lack of binding of exoRanGDP to exoKapα–Kapβ1results Kaps might serve to maintain the Ran gradient by regulating the in a higher outflow of exoRanGDP from the nucleus. movement of RanGTP/GDP through NPCs. To validate the latter, we used GFP (∼26 kDa), which is similar in size compared with Ran but does not bind to FG Nups or Kaps and, RESULTS AND DISCUSSION therefore should not be retained at the nuclear envelope.
Load more

Recommended publications
  • In Vitro Differentiation of Bone Marrow Mesenchymal Stem Cells Into Endometrial Epithelial Cells in Mouse: a Proteomic Analysis
    Int J Clin Exp Pathol 2014;7(7):3662-3672 www.ijcep.com /ISSN:1936-2625/IJCEP0000322 Original Article In vitro differentiation of bone marrow mesenchymal stem cells into endometrial epithelial cells in mouse: a proteomic analysis Qing Cong1,2, Bin Li1,2, Yisheng Wang1,2, Wenbi Zhang1,2, Mingjun Cheng1,2, Zhiyong Wu1,2, Xiaoyan Zhang1,2, Wei Jiang1,2, Congjian Xu1,2,3,4 1Obstetrics and Gynecology Hospital of Fudan University, 2Shanghai Key Laboratory of Female Reproductive Endocrine Related Diseases, 3Department of Obstetrics and Gynecology of Shanghai Medical School, 4Institute of Biomedical Sciences, Fudan University, Shanghai, P.R. China Received March 24, 2014; Accepted June 23, 2014; Epub June 15, 2014; Published July 1, 2014 Abstract: Objective: Mouse bone marrow mesenchymal stem cells (BMSCs) have been demonstrated to differenti- ate into female endometrial epithelial cells (EECs) in vivo. Our previous studies demonstrated that BMSCs can differentiate in the direction of EECs when co-cultured with endometrial stromal cells in vitro. Here, we obtain and analyse differential proteins and their relevant pathways in the process of BMSCs differentiating into EECs by iso- baric tags for relative and absolute quantitation (iTRAQ) proteomic analysis. Methods: A 0.4-µm pore size indirect co- culture system was established with female mice endometrial stromal cells (EStCs) restricted in the upper Transwell chamber and BMSCs in the lower well plate. After indirect co-culture for several days, the BMSCs were revealed to progressively differentiate towards EECs in vitro. Then, four groups were divided according to different co-culture days with single culture groups of BMSCs as controls.
    [Show full text]
  • (HMGB1) Deletion Leads to Small Heart and Glycolipid Metabolic
    Yu et al. Cell Death Discovery (2020) 6:106 https://doi.org/10.1038/s41420-020-00340-9 Cell Death Discovery ARTICLE Open Access Cardiomyocyte-restricted high-mobility group box 1 (HMGB1) deletion leads to small heart and glycolipid metabolic disorder through GR/PGC-1α signalling Peng Yu 1, Ming Liu2,BaoliZhang3,YingYu2,EnyongSu3,ShiyaoXie3,LeiZhang3,XueYang3,HongJiang 3, Ruizhen Chen3, Yunzeng Zou3 and Junbo Ge3 Abstract Cardiac growth and remodelling are key biological processes influencing the physiological performance of the heart, and a previous study showed a critical role for intracellular HMGB1 in vitro. However, the in vivo study, which used conditional Hmgb1 ablation, did not show a significant effect on cellular or organic function. We have demonstrated the extracellular effect of HMGB1 as a pro-inflammatory molecule on cardiac remodelling. In this study, we found that HMGB1 deletion by cTnT-Cre in mouse hearts altered glucocorticoid receptor (GR) function and glycolipid metabolism, eventually leading to growth retardation, small heart and heart failure. The subcellular morphology did not show a significant change caused by HMGB1 knockout. The heart showed significant elevation of glycolysis, free fatty acid deposition and related enzyme changes. Transcriptomic analysis revealed a list of differentially expressed genes that coincide with glucocorticoid receptor function in neonatal mice and a significant increase in inflammatory genes in 1234567890():,; 1234567890():,; 1234567890():,; 1234567890():,; adult mice. Cardiac HMGB1 knockout led to a series of changes in PGC-1α, UCP3 and GyK, which were the cause of metabolic changes and further impacted cardiac function. Ckmm-Cre Hmgb1fl/fl mice did not show a specific phenotype, which was consistent with the reported negative result of cardiomyocyte-specific Hmgb1 deletion via MHC-Cre.
    [Show full text]
  • (KPNA7), a Divergent Member of the Importin a Family of Nuclear Import
    Kelley et al. BMC Cell Biology 2010, 11:63 http://www.biomedcentral.com/1471-2121/11/63 RESEARCH ARTICLE Open Access Karyopherin a7 (KPNA7), a divergent member of the importin a family of nuclear import receptors Joshua B Kelley1, Ashley M Talley1, Adam Spencer1, Daniel Gioeli2, Bryce M Paschal1,3* Abstract Background: Classical nuclear localization signal (NLS) dependent nuclear import is carried out by a heterodimer of importin a and importin b. NLS cargo is recognized by importin a, which is bound by importin b. Importin b mediates translocation of the complex through the central channel of the nuclear pore, and upon reaching the nucleus, RanGTP binding to importin b triggers disassembly of the complex. To date, six importin a family members, encoded by separate genes, have been described in humans. Results: We sequenced and characterized a seventh member of the importin a family of transport factors, karyopherin a 7 (KPNA7), which is most closely related to KPNA2. The domain of KPNA7 that binds Importin b (IBB) is divergent, and shows stronger binding to importin b than the IBB domains from of other importin a family members. With regard to NLS recognition, KPNA7 binds to the retinoblastoma (RB) NLS to a similar degree as KPNA2, but it fails to bind the SV40-NLS and the human nucleoplasmin (NPM) NLS. KPNA7 shows a predominantly nuclear distribution under steady state conditions, which contrasts with KPNA2 which is primarily cytoplasmic. Conclusion: KPNA7 is a novel importin a family member in humans that belongs to the importin a2 subfamily. KPNA7 shows different subcellular localization and NLS binding characteristics compared to other members of the importin a family.
    [Show full text]
  • Supplementary Table 2
    Supplementary Table 2. Differentially Expressed Genes following Sham treatment relative to Untreated Controls Fold Change Accession Name Symbol 3 h 12 h NM_013121 CD28 antigen Cd28 12.82 BG665360 FMS-like tyrosine kinase 1 Flt1 9.63 NM_012701 Adrenergic receptor, beta 1 Adrb1 8.24 0.46 U20796 Nuclear receptor subfamily 1, group D, member 2 Nr1d2 7.22 NM_017116 Calpain 2 Capn2 6.41 BE097282 Guanine nucleotide binding protein, alpha 12 Gna12 6.21 NM_053328 Basic helix-loop-helix domain containing, class B2 Bhlhb2 5.79 NM_053831 Guanylate cyclase 2f Gucy2f 5.71 AW251703 Tumor necrosis factor receptor superfamily, member 12a Tnfrsf12a 5.57 NM_021691 Twist homolog 2 (Drosophila) Twist2 5.42 NM_133550 Fc receptor, IgE, low affinity II, alpha polypeptide Fcer2a 4.93 NM_031120 Signal sequence receptor, gamma Ssr3 4.84 NM_053544 Secreted frizzled-related protein 4 Sfrp4 4.73 NM_053910 Pleckstrin homology, Sec7 and coiled/coil domains 1 Pscd1 4.69 BE113233 Suppressor of cytokine signaling 2 Socs2 4.68 NM_053949 Potassium voltage-gated channel, subfamily H (eag- Kcnh2 4.60 related), member 2 NM_017305 Glutamate cysteine ligase, modifier subunit Gclm 4.59 NM_017309 Protein phospatase 3, regulatory subunit B, alpha Ppp3r1 4.54 isoform,type 1 NM_012765 5-hydroxytryptamine (serotonin) receptor 2C Htr2c 4.46 NM_017218 V-erb-b2 erythroblastic leukemia viral oncogene homolog Erbb3 4.42 3 (avian) AW918369 Zinc finger protein 191 Zfp191 4.38 NM_031034 Guanine nucleotide binding protein, alpha 12 Gna12 4.38 NM_017020 Interleukin 6 receptor Il6r 4.37 AJ002942
    [Show full text]
  • KPNA2 Promotes Metabolic Reprogramming in Glioblastomas By
    Li et al. Journal of Experimental & Clinical Cancer Research (2018) 37:194 https://doi.org/10.1186/s13046-018-0861-9 RESEARCH Open Access KPNA2 promotes metabolic reprogramming in glioblastomas by regulation of c-myc Jie Li1†, Qian Liu2†, Zihao Liu3, Qian Xia3, Zihao Zhang3, Rui Zhang1, Taihong Gao1, Guangyan Gu2, Yanan Wang2, Dan Wang2, Xiuyang Chen1, Yihang Yang1, Dong He1 and Tao Xin1,4* Abstract Background: Cancer cells maintain energy metabolism mainly by glycolysis, even under sufficient oxygen conditions. It gives cancer cells better growth advantages under complicated internal environment. KPNA2 is a novel oncogene that has received much attention in recent years, but the exact mechanisms of KPNA2 in tumorigenesis and progression are largely unknown. Especially its potential roles in the metabolic transformation of tumors still remain to be explored. Methods: The expressions of KPNA2 in glioblastoma and normal human brain samples were analyzed by immunohistochemical analysis. The activities of key enzymes in glycolysis, the production of lactate acid and glucose uptake were investigated by colorimetry. GLUT-1 expression was measured by flow cytometry. CCK8 was used to examine the cell viability in vitro, and the xenograft models in nude mice were established to explore the roles of KPNA2 in vivo. In addition, Co-IP, subcellular fractionation, western blot, immunofluorescence and luciferase assay were used to investigate the internal connection between KPNA2, c-myc and E2F1. Results: In the present study, we found that KPNA2 was highly expressed in the glioma compared to the normal brain tissues. Level of KPNA2 was an independent predictor of prognosis in the glioma patients.
    [Show full text]
  • Supplemental Table 1
    Symbol Gene name MIN6.EXO MIN6.M1 MIN6.M2 MIN6.M3 MIN6.M4 A2m alpha-2-macroglobulin A2m Acat1 acetyl-Coenzyme A acetyltransferase 1 Acat1 Acly ATP citrate lyase Acly Acly Acly Act Actin Act Act Act Act Aga aspartylglucosaminidase Aga Ahcy S-adenosylhomocysteine hydrolase Ahcy Alb Albumin Alb Alb Alb Aldoa aldolase A, fructose-bisphosphate Aldoa Anxa5 Annexin A5 Anxa5 AP1 Adaptor-related protein complex AP1 AP2 Adaptor protein complex AP2 Arf1 ADP-ribosylation factor 1 Arf1 Atp1a1 ATPase Na/K transpoting Atp1a1 ATP1b1 Na/K ATPase beta subunit ATP1b1 ATP6V1 ATPase, H+ transporting.. ATP6V1 ATP6v1 ATP6v1 Banf1 Barrier to autointegration factor Banf1 Basp1 brain abundant, memrane signal protein 1 Basp1 C3 complement C3 C3 C3 C3 C4 Complement C4 C4 C4 C4 Calm2 calmodulin 2 (phosphorylase kinase, delta) Calm2 Capn5 Calpain 5 Capn5 Capn5 Cct5 chaperonin subunit 5 Cct5 Cct8 chaperonin subunit 8 Cct8 CD147 basigin CD147 CD63 CD63 CD63 CD81 CD81 CD81 CD81 CD81 CD81 CD81 CD82 CD82 CD82 CD82 CD90.2 thy1.2 CD90.2 CD98 Slc3a2 CD98 CD98 Cdc42 Cell division cycle 42 Cdc42 Cfl1 Cofilin 1 Cfl1 Cfl1 Chmp4b chromatin modifying protein 4B Chmp4b Chmp5 chromatin modifying protein 5 Chmp5 Clta clathrin, light polypeptide A Clta Cltc Clathrin Hc Cltc Cltc Cltc Cltc Clu clusterin Clu Col16a1 collagen 16a1 Col16a1 Col2 Collagen type II Col2a1 Col2 Col6 Collagen type VI alpha 3 Col6a3 Col6 CpE carboxypeptidase E CpE CpE CpE, CpH CpE CpE Cspg4 Chondroitin sulfate proteoglycan 4 Cspg4 CyCAP Cyclophilin C-associated protein CyCAP CyCAP Dnpep aspartyl aminopeptidase Dnpep Dstn destrin Dstn EDIL3 EGF-like repeat discoidin.
    [Show full text]
  • Regulation of Protein Kinase Cδ Nuclear Import and Apoptosis by Mechanistic Target of Rapamycin Complex-1 Antonio Layoun1,5, Alexander A
    www.nature.com/scientificreports Corrected: Publisher Correction OPEN Regulation of protein kinase Cδ Nuclear Import and Apoptosis by Mechanistic Target of Rapamycin Complex-1 Antonio Layoun1,5, Alexander A. Goldberg1,5, Ayesha Baig1, Mikaela Eng1, Ortal Attias1, Kristof Nelson1, Alexandra Carella1, Nahomi Amberber1, Jill A. Fielhaber1, Kwang-Bo Joung1, T. Martin Schmeing 2, Yingshan Han1, Jefrey Downey3, Maziar Divangahi3, Philippe P. Roux 4 & Arnold S. Kristof1* Inactivation of the protein complex ‘mechanistic target of rapamycin complex 1’ (mTORC1) can increase the nuclear content of transcriptional regulators of metabolism and apoptosis. Previous studies established that nuclear import of signal transducer and activator of transcription-1 (STAT1) requires the mTORC1-associated adaptor karyopherin-α1 (KPNA1) when mTORC1 activity is reduced. However, the role of other mTORC1-interacting proteins in the complex, including ‘protein kinase C delta’ (PKCδ), have not been well characterized. In this study, we demonstrate that PKCδ, a STAT1 kinase, contains a functional ‘target of rapamycin signaling’ (TOS) motif that directs its interaction with mTORC1. Depletion of KPNA1 by RNAi prevented the nuclear import of PKCδ in cells exposed to the mTORC1 inhibitor rapamycin or amino acid restriction. Mutation of the TOS motif in PKCδ led to its loss of regulation by mTORC1 or karyopherin-α1, resulting in increased constitutive nuclear content. In cells expressing wild-type PKCδ, STAT1 activity and apoptosis were increased by rapamycin or interferon-β. Those expressing the PKCδ TOS mutant exhibited increased STAT1 activity and apoptosis; further enhancement by rapamycin or interferon-β, however, was lost. Therefore, the TOS motif in PKCδ is a novel structural mechanism by which mTORC1 prevents PKCδ and STAT1 nuclear import, and apoptosis.
    [Show full text]
  • Nucleocytoplasmic Transport: Regulatory Mechanisms and the Implications in Neurodegeneration
    International Journal of Molecular Sciences Review Nucleocytoplasmic Transport: Regulatory Mechanisms and the Implications in Neurodegeneration Baojin Ding * and Masood Sepehrimanesh Department of Biology, University of Louisiana at Lafayette, 410 East Saint Mary Boulevard, Lafayette, LA 70503, USA; [email protected] * Correspondence: [email protected] Abstract: Nucleocytoplasmic transport (NCT) across the nuclear envelope is precisely regulated in eukaryotic cells, and it plays critical roles in maintenance of cellular homeostasis. Accumulating evidence has demonstrated that dysregulations of NCT are implicated in aging and age-related neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD), Alzheimer’s disease (AD), and Huntington disease (HD). This is an emerging research field. The molecular mechanisms underlying impaired NCT and the pathogenesis leading to neurodegener- ation are not clear. In this review, we comprehensively described the components of NCT machinery, including nuclear envelope (NE), nuclear pore complex (NPC), importins and exportins, RanGTPase and its regulators, and the regulatory mechanisms of nuclear transport of both protein and transcript cargos. Additionally, we discussed the possible molecular mechanisms of impaired NCT underlying aging and neurodegenerative diseases, such as ALS/FTD, HD, and AD. Keywords: Alzheimer’s disease; amyotrophic lateral sclerosis; Huntington disease; neurodegenera- tive diseases; nuclear pore complex; nucleocytoplasmic transport; Ran GTPase Citation: Ding, B.; Sepehrimanesh, M. Nucleocytoplasmic Transport: Regulatory Mechanisms and the Implications in Neurodegeneration. 1. Introduction Int. J. Mol. Sci. 2021, 22, 4165. As a hallmark of eukaryotic cells, the genetic materials are separated from the cyto- https://doi.org/10.3390/ijms plasmic contents by a highly regulated membrane, called nuclear envelope (NE), which 22084165 has two concentric bilayer membranes, the inner nuclear membrane (INM), and outer nuclear membrane (ONM).
    [Show full text]
  • Thesis Was to Elucidate the Function of Recently Discovered Nuclear IGF-1R As Well As to Investigate Its Nuclear Translocation Pathway
    From the Department of Oncology-Pathology Karolinska Institutet, Stockholm, Sweden FROM MEMBRANE TO NUCLEUS: NEW ROLES AND FUNCTIONS OF SUMOYLATED IGF-1R AND EGFR Sylvia Packham Stockholm 2015 All previously published papers were reproduced with permission from the publisher. Published by Karolinska Institutet. Printed by AJ E-print AB, Stockholm, Sweden © Sylvia Packham, 2015 ISBN 978-91-7549-877-5 To my beloved Justin and Family ABSTRACT Cell surface receptor tyrosine kinases (RTKs) role in cell signaling have been studied for decades and their role in cancer progression are undisputable. The insulin-like growth factor 1 receptor, IGF-1R, has been demonstrated to play a critical part in tumorigenesis; downregulation of the IGF-1R in tumor xenografts results in complete tumor regression. Previously, RTK research has focused on the canonical signaling pathways activated by ligand binding at the plasma membrane. However, strong evidence keeps emerging that several RTKs have a second functionally mechanism, inside the cell nucleus, where the receptors reside after ligand stimulation. The aim of this thesis was to elucidate the function of recently discovered nuclear IGF-1R as well as to investigate its nuclear translocation pathway. Since it was previously shown that SUMOylation of the IGF-1R is essential for its nuclear translocation we also set out to investigate SUMO modification of the epidermal growth factor receptor (EGFR). In paper I, we present a functional role for nuclear IGF-1R in gene transcription. Inside the nucleus, IGF-1R functions as a co-activator to LEF-1/TCF transcription factor. Nuclear IGF-1R enhances transcription of cyclin D1 and axin2, and we show that it is enriched in the cyclin D1 promoter region.
    [Show full text]
  • Nuclear Localization Signal-Independent and Importin/Karyopherin-Independent Nuclear Import of B-Catenin François Fagotto*, Ursula Glück and Barry M
    View metadata, citation and similar papers at core.ac.uk brought to you by CORE Researchprovided byPaper Elsevier -181 Publisher Connector Nuclear localization signal-independent and importin/karyopherin-independent nuclear import of b-catenin François Fagotto*, Ursula Glück and Barry M. Gumbiner Background: Control of the nuclear localization of specific proteins is an Address: Cellular Biochemistry and Biophysics important mechanism for regulating many signal transduction pathways. Upon Program, Memorial Sloan–Kettering Cancer Center, 1275 York Avenue, Box 564, New York 10021, activation of the Wnt signaling pathway, β-catenin localizes into the nucleus and USA. interacts with TCF/LEF-1 (T-cell factor/lymphocyte enhancer factor-1) transcription factors, triggering activation of downstream genes. The role of *Present address: Max-Plank Institute for regulated nuclear localization in β-catenin signaling is still unclear. β-catenin has Developmental Biology, Division for Cell Biology, Spemannstrasse 35-V, 72076 Tübingen, Germany. no nuclear localization sequence (NLS). Although it has been reported that β- catenin can piggyback into the nucleus by binding to TCF/LEF-1, there is Correspondence: Barry M. Gumbiner evidence that its import is independent of TCF/LEF-1 in vivo. Therefore, the E-mail: [email protected] mechanism for β-catenin nuclear localization remains to be established. Received: 22 October 1997 15 January 1998 β Revised: Results: We have analyzed -catenin nuclear import in an in vitro assay using Accepted: 15 January 1998 permeabilized cells. β-catenin docks specifically onto the nuclear envelope in the absence of other cytosolic factors. Docking is not inhibited by an NLS Published: 23 January 1998 peptide and does not require importins/karyopherins, the receptors for classical Current Biology 1998, 8:181–190 β β NLS substrates.
    [Show full text]
  • Comparative Gene Expression Analysis to Identify Common Factors in Multiple Cancers
    COMPARATIVE GENE EXPRESSION ANALYSIS TO IDENTIFY COMMON FACTORS IN MULTIPLE CANCERS DISSERTATION Presented in Partial Fulfillment of the Requirements for the Degree Doctor of Philosophy in the Graduate School of The Ohio State University By Leszek A. Rybaczyk, B.A. ***** The Ohio State University 2008 Dissertation Committee: Professor Kun Huang, Adviser Professor Jeffery Kuret Approved by Professor Randy Nelson Professor Daniel Janies ------------------------------------------- Adviser Integrated Biomedical Science Graduate Program ABSTRACT Most current cancer research is focused on tissue-specific genetic mutations. Familial inheritance (e.g., APC in colon cancer), genetic mutation (e.g., p53), and overexpression of growth receptors (e.g., Her2-neu in breast cancer) can potentially lead to aberrant replication of a cell. Studies of these changes provide tremendous information about tissue-specific effects but are less informative about common changes that occur in multiple tissues. The similarity in the behavior of cancers from different organ systems and species suggests that a pervasive mechanism drives carcinogenesis, regardless of the specific tissue or species. In order to detect this mechanism, I applied three tiers of analysis at different levels: hypothesis testing on individual pathways to identify significant expression changes within each dataset, intersection of results between different datasets to find common themes across experiments, and Pearson correlations between individual genes to identify correlated genes within each dataset. By comparing a variety of cancers from different tissues and species, I was able to separate tissue and species specific effects from cancer specific effects. I found that downregulation of Monoamine Oxidase A is an indicator of this pervasive mechanism and can potentially be used to detect pathways and functions related to the initiation, promotion, and progression of cancer.
    [Show full text]
  • Transcriptomic Analysis of Patients with Tetralogy of Fallot Reveals the Effect of Chronic Hypoxia on Myocardial Gene Expression
    Ghorbel et al Congenital Heart Disease Transcriptomic analysis of patients with tetralogy of Fallot reveals the effect of chronic hypoxia on myocardial gene expression Mohamed T. Ghorbel, PhD, Myriam Cherif, PhD, Emma Jenkins, PhD, Amir Mokhtari, MRCS, Damien Kenny, MRCPCH, Gianni D. Angelini, FRCS, and Massimo Caputo, MD Objectives: In cyanotic patients undergoing repair of heart defects, chronic hypoxia is thought to lead to greater susceptibility to ischemia and reoxygenation injury. We sought to find an explanation to such a hypothesis by investigating the cardiac gene expression in patients with tetralogy of Fallot undergoing cardiac surgery. CHD Methods: The myocardial gene profile was investigated in right ventricular biopsy specimens obtained from 20 patients with a diagnosis of cyanotic (n ¼ 11) or acyanotic (n ¼ 9) tetralogy of Fallot undergoing surgical repair. Oligonucleotide microarray analyses were performed on the samples, and the array results were validated with Western blotting and enzyme-linked immunosorbent assay. Results: Data revealed 795 differentially expressed genes in cyanotic versus acyanotic hearts, with 198 upregu- lated and 597 downregulated. Growth/morphogenesis, remodeling, and apoptosis emerged as dominant func- tional themes for the upregulated genes and included the apoptotic gene TRAIL (tumor necrosis factor–related apoptosis-inducing ligand), the remodeling factor OPN (osteopontin), and the mitochondrial function gene COX11 (cytochrome-c oxidase 11). In contrast, transcription, mitogen-activated protein kinase signaling, and contractile machinery were the dominant functional classes for the downregulated genes, which included the calcium-handling gene NCX1 (sodium-calcium exchanger). Protein levels of COX11, NCX1, OPN, and LYZ (ly- sozyme) in the myocardium followed the same pattern obtained by means of transcriptomics.
    [Show full text]