Paxillin nuclear-cytoplasmic localization is regulated by phosphorylation of the LD4 motif: evidence that nuclear paxillin promotes cell proliferation

2009 ◽  
Vol 418 (1) ◽  
pp. 173-184 ◽  
Author(s):  
Jing-Ming Dong ◽  
Lei-Shong Lau ◽  
Yuen-Wai Ng ◽  
Louis Lim ◽  
Ed Manser
Keyword(s):  
Cell Proliferation ◽  
Focal Adhesion ◽  
Nuclear Export ◽  
Nuclear Export Signal ◽  
Pcr Analysis ◽  
Surface Receptors ◽  
Lim Domains ◽  
P21 Activated Kinase ◽  
Nih 3T3 ◽  
G Protein Coupled

Paxillin, a major focal-adhesion complex component belongs to the subfamily of LIM domain proteins and participates in cell adhesion-mediated signal transduction. It is implicated in cell-motility responses upon activation of cell-surface receptors and can recruit, among others, the GIT1 [GRK (G-protein-coupled-receptor kinase)-interacting ARF (ADP-ribosylation factor) GAP (GTPase-activating protein)]–PIX [PAK (p21-activated kinase)-interacting exchange factor]–PAK1 complex. Several adhesion proteins including zyxin, Hic5 and Trip6 are also nuclear and can exert transcriptional effects. In the present study we show that endogenous paxillin shuttles between the cytoplasm and nucleus, and we have used a variety of tagged paxillin constructs to map the nuclear export signal. This region overlaps an important LD4 motif that binds GIT1 and FAK1 (focal-adhesion kinase 1). We provide evidence that phosphorylation of Ser272 within LD4 blocks nuclear export, and we show that this modification also reduces GIT1, but not FAK1, binding; however, Ser272 phosphorylation does not appear to be mediated by PAK1 as previously suggested. Expression of nuclear-localized paxillin LIM domains stimulate DNA synthesis and cell proliferation. By real-time PCR analysis we have established that overexpression of either full-length paxillin or a truncated nuclear form suppresses expression of the parental imprinted gene H19, and modulation of this locus probably affects the rate of NIH-3T3 cell proliferation.

scholarly journals Bioinformatic Analysis of Structure and Function of LIM Domains of Human Zyxin Family Proteins

2021 ◽  
Vol 22 (5) ◽  
pp. 2647
Author(s):  
M. Quadir Siddiqui ◽  
Maulik D. Badmalia ◽  
Trushar R. Patel
Keyword(s):  
Nucleic Acid ◽  
Nuclear Export ◽  
Consensus Sequence ◽  
Nuclear Export Signal ◽  
Bioinformatic Analysis ◽  
Nucleic Acid Binding ◽  
Protein Protein Interaction ◽  
Lim Domains ◽  
Protein Nucleic Acid ◽  
And Function

Members of the human Zyxin family are LIM domain-containing proteins that perform critical cellular functions and are indispensable for cellular integrity. Despite their importance, not much is known about their structure, functions, interactions and dynamics. To provide insights into these, we used a set of in-silico tools and databases and analyzed their amino acid sequence, phylogeny, post-translational modifications, structure-dynamics, molecular interactions, and functions. Our analysis revealed that zyxin members are ohnologs. Presence of a conserved nuclear export signal composed of LxxLxL/LxxxLxL consensus sequence, as well as a possible nuclear localization signal, suggesting that Zyxin family members may have nuclear and cytoplasmic roles. The molecular modeling and structural analysis indicated that Zyxin family LIM domains share similarities with transcriptional regulators and have positively charged electrostatic patches, which may indicate that they have previously unanticipated nucleic acid binding properties. Intrinsic dynamics analysis of Lim domains suggest that only Lim1 has similar internal dynamics properties, unlike Lim2/3. Furthermore, we analyzed protein expression and mutational frequency in various malignancies, as well as mapped protein-protein interaction networks they are involved in. Overall, our comprehensive bioinformatic analysis suggests that these proteins may play important roles in mediating protein-protein and protein-nucleic acid interactions.

scholarly journals Bioinformatic Analysis of Structure and Function of Lim Domains of Zyxin Family Proteins

2020 ◽  
Author(s):  
Mohammad Quadir Siddiqui ◽  
Maulik D. Badmalia ◽  
Trushar R. Patel
Keyword(s):  
Protein Interactions ◽  
Nuclear Export ◽  
Nuclear Export Signal ◽  
Bioinformatic Analysis ◽  
Elastic Network Model ◽  
Cellular Functions ◽  
Consensus Analysis ◽  
Mutational Frequency ◽  
Lim Domains ◽  
And Function

AbstractLim domains are one of the most abundant types of zinc-finger domains and are linked with diverse cellular functions ranging from cytoskeleton maintenance to gene regulation. Zyxin family Lim domains perform the critical cellular functions and are indispensable for cellular integrity. Despite having these important functions the fundamental nature of the sequence, structure, functions, and interactions of the Zyxin family Lim domains are largely unknown. Therefore, we have used a set of in-silico tools and bioinformatics databases to distill the fundamental properties of the Zyxin family proteins/Lim domains from their amino acid sequence, phylogeny, biochemical analysis, post-translational modifications, structure dynamics, molecular interactions, and functions. Consensus analysis of the nuclear export signal suggests a conserved Leucine-rich motif composed of LxxLxL/LxxxLxL. Molecular modeling and structural analysis demonstrate that Lim domains of the members of the Zyxin family proteins share similarities with transcriptional regulators, suggesting they could interact with nucleic acids as well. Normal mode, Covariance, and Elastic Network Model analysis of Zyxin family Lim domains suggest only the Lim1 region has similar internal dynamics properties, compared to Lim2/3. Protein expression/mutational frequency studies of the Zyxin family demonstrated higher expression and mutational frequency rates in various malignancies. Protein-protein interactions indicate that these proteins could facilitate metabolic rewiring and oncogenic addiction paradigm. Our comprehensive analysis of the Zyxin family proteins indicates that the Lim domains function in a variety of ways and could be implicated in rational protein engineering and might lead as a better therapeutic target for various diseases, including cancers.

scholarly journals Caspase-mediated nuclear pore complex trimming in cell differentiation and endoplasmic reticulum stress

2021 ◽  
Author(s):  
Ukrae H. Cho ◽  
Martin W. Hetzer
Keyword(s):  
Endoplasmic Reticulum ◽  
Focal Adhesion ◽  
Nuclear Pore Complex ◽  
Nuclear Export ◽  
Nuclear Export Signal ◽  
Nuclear Pore ◽  
Genome Regulation ◽  
Focal Adhesion Proteins ◽  
Domain Protein ◽  
Export Signal

Introductory ParagraphDuring programmed cell death, caspases degrade 7 out of ∼30 nucleoporins (Nups) to irreversibly demolish the nuclear pore complex (NPC)1. However, for poorly understood reasons, caspases are also activated in differentiating cells in a non-apoptotic manner2,3. Here, we describe reversible, caspase-mediated NPC “trimming” during early myogenesis. We find that sublethal levels of caspases selectively proteolyze 4 peripheral Nups, Nup358, Nup214, Nup153, and Tpr, resulting in the transient block of nuclear export pathways. Several nuclear export signal (NES)-containing focal adhesion proteins concomitantly accumulate in the nucleus where they function as transcription cofactors4. We show that one such protein, FAK (focal adhesion kinase), drives a global reconfiguration of MBD2 (methyl CpG binding domain protein 2)-mediated genome regulation. We also observe caspase-mediated NPC trimming during neurogenesis and endoplasmic reticulum (ER) stress. Our results illustrate that the NPC can be proteolytically regulated in response to non-apoptotic cues, and call for a reassessment of the death-centric view of caspases.

scholarly journals A Nuclear Export Signal and Phosphorylation Regulate Dok1 Subcellular Localization and Functions

2006 ◽  
Vol 26 (11) ◽  
pp. 4288-4301 ◽  
Author(s):  
Yamei Niu ◽  
François Roy ◽  
Frédéric Saltel ◽  
Charlotte Andrieu-Soler ◽  
Wen Dong ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Nuclear Export ◽  
Nuclear Export Signal ◽  
Specific Inhibitor ◽  
Adaptor Protein ◽  
Cell Spreading ◽  
Nuclear Accumulation ◽  
Serum Starvation ◽  
Cytoplasmic Localization ◽  
Export Signal

ABSTRACT Dok1 is believed to be a mainly cytoplasmic adaptor protein which down-regulates mitogen-activated protein kinase activation, inhibits cell proliferation and transformation, and promotes cell spreading and cell migration. Here we show that Dok1 shuttles between the nucleus and cytoplasm. Treatment of cells with leptomycin B (LMB), a specific inhibitor of the nuclear export signal (NES)-dependent receptor CRM1, causes nuclear accumulation of Dok1. We have identified a functional NES (348LLKAKLTDPKED359) that plays a major role in the cytoplasmic localization of Dok1. Src-induced tyrosine phosphorylation prevented the LMB-mediated nuclear accumulation of Dok1. Dok1 cytoplasmic localization is also dependent on IKKβ. Serum starvation or maintaining cells in suspension favor Dok1 nuclear localization, while serum stimulation, exposure to growth factor, or cell adhesion to a substrate induce cytoplasmic localization. Functionally, nuclear NES-mutant Dok1 had impaired ability to inhibit cell proliferation and to promote cell spreading and cell motility. Taken together, our results provide the first evidence that Dok1 transits through the nucleus and is actively exported into the cytoplasm by the CRM1 nuclear export system. Nuclear export modulated by external stimuli and phosphorylation may be a mechanism by which Dok1 is maintained in the cytoplasm and membrane, thus regulating its signaling functions.

scholarly journals Hic-5 Communicates between Focal Adhesions and the Nucleus through Oxidant-Sensitive Nuclear Export Signal

2003 ◽  
Vol 14 (3) ◽  
pp. 1158-1171 ◽  
Author(s):  
Motoko Shibanuma ◽  
Joo-ri Kim-Kaneyama ◽  
Keiko Ishino ◽  
Nobuko Sakamoto ◽  
Tomoko Hishiki ◽  
...  
Keyword(s):  
Gene Expression ◽  
Focal Adhesion ◽  
Nuclear Export ◽  
Nuclear Export Signal ◽  
Focal Adhesions ◽  
Dominant Negative ◽  
Nuclear Accumulation ◽  
Adhesion Protein ◽  
Focal Adhesion Protein ◽  
Export Signal

hic-5 was originally isolated as an H2O2-inducible cDNA clone whose product was normally found at focal adhesions. In this study, we found that Hic-5 accumulated in the nucleus in response to oxidants such as H2O2. Other focal adhesion proteins including paxillin, the most homologous to Hic-5, remained in the cytoplasm. Mutation analyses revealed that the C- and N-terminal halves of Hic-5 contributed to its nuclear localization in a positive and negative manner, respectively. After the finding that leptomycin B (LMB), an inhibitor of nuclear export signal (NES), caused Hic-5 to be retained in the nucleus, Hic-5 was demonstrated to harbor NES in the N-terminal, which was sensitive to oxidants, thereby regulating the nuclear accumulation of Hic-5. NES consisted of a leucine-rich stretch and two cysteines with a limited similarity to Yap/Pap-type NES. In the nucleus, Hic-5 was suggested to participate in the gene expression of c-fos. Using dominant negative mutants, we found that Hic-5 was actually involved in endogenous c-fos gene expression upon H2O2 treatment. Hic-5 was thus proposed as a focal adhesion protein with the novel aspect of shuttling between focal adhesions and the nucleus through an oxidant-sensitive NES, mediating the redox signaling directly to the nucleus.

111-OR: Pancreatic Elastase Regulates Human Beta-Cell Proliferation via the Focal Adhesion and PAR2 Signaling Pathways

Diabetes ◽  
2019 ◽  
Vol 68 (Supplement 1) ◽  
pp. 111-OR
Author(s):  
GIORGIO BASILE ◽  
AMEDEO VETERE ◽  
KA-CHEUK LIU ◽  
JIANG HU ◽  
OLOV ANDERSSON ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Beta Cell ◽  
Signaling Pathways ◽  
Focal Adhesion ◽  
Beta Cell Proliferation ◽  
Pancreatic Elastase ◽  
Human Beta Cell

Comparative Study on Mitogen Activated Protein Kinase of Plasmodium Species by Using in silico Method

2012 ◽  
Vol 9 (1) ◽  
pp. 1
Author(s):  
Mohd Fakharul Zaman Raja Yahya ◽  
Hasidah Mohd Sidek
Keyword(s):  
Amino Acid ◽  
In Silico ◽  
Nuclear Export ◽  
Nuclear Export Signal ◽  
Plasmodium Species ◽  
Mitogen Activated Protein Kinase ◽  
Multiple Sequence ◽  
Wide Range ◽  
Mitogen Activated Protein ◽  
Human Plasmodium

Malaria parasites, Plasmodium can infect a wide range of hosts including humans and rodents. There are two copies of mitogen activated protein kinases (MAPKs) in Plasmodium, namely MAPK1 and MAPK2. The MAPKs have been studied extensively in the human Plasmodium, P. falciparum. However, the MAPKs from other Plasmodium species have not been characterized and it is therefore the premise of presented study to characterize the MAPKs from other Plasmodium species-P. vivax, P. knowlesi, P. berghei, P. chabaudi and P.yoelli using a series of publicly available bioinformatic tools. In silico data indicates that all Plasmodium MAPKs are nuclear-localized and contain both a nuclear localization signal (NLS) and a Leucine-rich nuclear export signal (NES). The activation motifs of TDY and TSH were found to be fully conserved in Plasmodium MAPK1 and MAPK2, respectively. The detailed manual inspection of a multiple sequence alignment (MSA) construct revealed a total of 17 amino acid stack patterns comprising of different amino acids present in MAPKJ and MAPK2 respectively, with respect to rodent and human Plasmodia. It is proposed that these amino acid stack patterns may be useful in explaining the disparity between rodent and human Plasmodium MAPKs. 

Comparative Study on Mitogen Activated Protein Kinase of Plasmodium Species by Using In silico Method

2012 ◽  
Vol 9 (1) ◽  
pp. 1
Author(s):  
Mohd Fakharul Zaman Raja Yahya ◽  
Hasidah Mohd Sidek
Keyword(s):  
Amino Acid ◽  
In Silico ◽  
Nuclear Export ◽  
Nuclear Export Signal ◽  
Plasmodium Species ◽  
Mitogen Activated Protein Kinase ◽  
Multiple Sequence ◽  
Bioinformatic Tools ◽  
Wide Range ◽  
Human Plasmodium

Malaria parasites, Plasmodium can infect a wide range ofhosts including humans and rodents. There are two copies ofmitogen activated protein kinases (MAPKs) in Plasmodium, namely MAPK1 and MAPK2. The MAPKs have been studied extensively in the human Plasmodium, P. falciparum. However, the MAPKs from other Plasmodium species have not been characterized and it is therefore the premise ofpresented study to characterize the MAPKs from other Plasmodium species-P. vivax, P. knowlesi, P. berghei, P. chabaudi and P.yoelli using a series ofpublicly available bioinformatic tools. In silico data indicates that all Plasmodium MAPKs are nuclear-localizedandcontain both a nuclear localization signal (NLS) anda Leucine-rich nuclear export signal (NES). The activation motifs ofTDYand TSH werefound to befully conserved in Plasmodium MAPK1 and MAPK2, respectively. The detailed manual inspection ofa multiple sequence alignment (MSA) construct revealed a total of 17 amino acid stack patterns comprising ofdifferent amino acids present in MAPK1 and MAPK2 respectively, with respect to rodent and human Plasmodia. 1t is proposed that these amino acid stack patterns may be useful in explaining the disparity between rodent and human Plasmodium MAPKs.

scholarly journals Specific inhibition of the Survivin–CRM1 interaction by peptide-modified molecular tweezers

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Annika Meiners ◽  
Sandra Bäcker ◽  
Inesa Hadrović ◽  
Christian Heid ◽  
Christine Beuck ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Rational Design ◽  
Nuclear Export Signal ◽  
Signal Interference ◽  
Receptor Interaction ◽  
Dual Function ◽  
Experimental Proof ◽  
Molecular Tweezers ◽  
Recognition Element ◽  
Dynamics Simulations

AbstractSurvivin’s dual function as apoptosis inhibitor and regulator of cell proliferation is mediated via its interaction with the export receptor CRM1. This protein–protein interaction represents an attractive target in cancer research and therapy. Here, we report a sophisticated strategy addressing Survivin’s nuclear export signal (NES), the binding site of CRM1, with advanced supramolecular tweezers for lysine and arginine. These were covalently connected to small peptides resembling the natural, self-complementary dimer interface which largely overlaps with the NES. Several biochemical methods demonstrated sequence-selective NES recognition and interference with the critical receptor interaction. These data were strongly supported by molecular dynamics simulations and multiscale computational studies. Rational design of lysine tweezers equipped with a peptidic recognition element thus allowed to address a previously unapproachable protein surface area. As an experimental proof-of-principle for specific transport signal interference, this concept should be transferable to any protein epitope with a flanking well-accessible lysine.

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