A monopartite nuclear localization sequence regulates nuclear targeting of the actin binding protein myopodin

Type II phosphatidylinositol phosphate kinases (PIPkins) have recently been found to be primarily phosphatidylinositol 5-phosphate 4-kinases, and their physiological role remains unclear. We have previously shown that a Type II PIPkin [isoform(s) unknown], is localized partly in the nucleus [Divecha, Rhee, Letcher and Irvine (1993) Biochem. J. 289, 617-620], and here we show, by transfection of HeLa cells with green-fluorescent-protein-tagged Type II PIPkins, that this is likely to be the Type IIβ isoform. Type IIβ PIPkin has no obvious nuclear localization sequence, and a detailed analysis of the localization of chimaeras and mutants of the α (cytosolic) and β PIPkins shows that the nuclear localization requires the presence of a 17-amino-acid length of α-helix (α-helix 7) that is specific to the β isoform, and that this helix must be present in its entirety, with a precise orientation. This resembles the nuclear targeting of the HIV protein Vpr, and Type IIβ PIPkin is apparently therefore the first example of a eukaryotic protein that uses the same mechanism.

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Identification of the Nuclear Localization Signal inXenopus Cyclin E and Analysis of Its Role in Replication and Mitosis

Molecular Biology of the Cell ◽

10.1091/mbc.e02-07-0449 ◽

2002 ◽

Vol 13 (12) ◽

pp. 4388-4400 ◽

Cited By ~ 26

Author(s):

Jonathan D. Moore ◽

Sally Kornbluth ◽

Tim Hunt

Keyword(s):

Dna Replication ◽

Nuclear Localization ◽

Cyclin E ◽

T Antigen ◽

Cyclin B ◽

Nuclear Accumulation ◽

Nuclear Localization Sequence ◽

Nuclear Targeting ◽

Sv40 Large T Antigen ◽

Localization Sequence

Cyclin-dependent kinase (Cdk)2/cyclin E is imported into nuclei assembled in Xenopus egg extracts by a pathway that requires importin-α and -β. Here, we identify a basic nuclear localization sequence (NLS) in the N-terminus ofXenopus cyclin E. Mutation of the NLS eliminated nuclear accumulation of both cyclin E and Cdk2, and such versions of cyclin E were unable to trigger DNA replication. Addition of a heterologous NLS from SV40 large T antigen restored both nuclear targeting of Cdk2/cyclin E and DNA replication. We present evidence indicating that Cdk2/cyclin E complexes must become highly concentrated within nuclei to support replication and find that cyclin A can trigger replication at much lower intranuclear concentrations. We confirmed that depletion of endogenous cyclin E increases the concentration of cyclin B necessary to promote entry into mitosis. In contrast to its inability to promote DNA replication, cyclin E lacking its NLS was able to cooperate with cyclin B in promoting mitotic entry.

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Actin dynamics regulate subcellular localization of the F-actin-binding protein PALLD in mouse Sertoli cells

Reproduction ◽

10.1530/rep-14-0147 ◽

2014 ◽

Vol 148 (4) ◽

pp. 333-341 ◽

Cited By ~ 6

Author(s):

Bryan A Niedenberger ◽

Vesna A Chappell ◽

Carol A Otey ◽

Christopher B Geyer

Keyword(s):

Subcellular Localization ◽

Sertoli Cell ◽

Nuclear Localization ◽

Sertoli Cells ◽

Binding Protein ◽

Terminal Differentiation ◽

Mutant Mice ◽

Actin Binding ◽

Actin Binding Protein ◽

Receptor Mutant

Sertoli cells undergo terminal differentiation at puberty to support all phases of germ cell development, which occurs in the mouse beginning in the second week of life. By ∼18 dayspostpartum(dpp), nearly all Sertoli cells have ceased proliferation. This terminal differentiation is accompanied by the development of unique and regionally concentrated filamentous actin (F-actin) structures at the basal and apical aspects of the seminiferous epithelium, and this reorganization is likely to involve the action of actin-binding proteins. Palladin (PALLD) is a widely expressed F-actin-binding and bundling protein recently shown to regulate these structures, yet it is predominantly nuclear in Sertoli cells at puberty. We found that PALLD localized within nuclei of primary Sertoli cells grown in serum-free media but relocalized to the cytoplasm upon serum stimulation. We utilized this system within vivorelevance to Sertoli cell development to investigate mechanisms regulating nuclear localization of this F-actin-binding protein. Our results indicate that PALLD can be shuttled from the nucleus to the cytoplasm, and that this relocalization occurred following depolymerization of the F-actin cytoskeleton in response to cAMP signaling. Nuclear localization was reduced inHpg-mutant testes, suggesting the involvement of gonadotropin signaling. We found that PALLD nuclear localization was unaffected in testis tissues from LH receptor and androgen receptor-mutant mice. However, PALLD nuclear localization was reduced in the testes of FSH receptor-mutant mice, suggesting that FSH signaling during Sertoli cell maturation regulates this subcellular localization.

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Characterization of nuclear localization and nuclear export signals of yeast actin-binding protein Pan1

FEBS Letters ◽

10.1016/j.febslet.2007.10.036 ◽

2007 ◽

Vol 581 (28) ◽

pp. 5371-5376 ◽

Cited By ~ 4

Author(s):

Joanna Kamińska ◽

Magdalena Sędek ◽

Monika Wysocka-Kapcińska ◽

Teresa Żołądek

Keyword(s):

Nuclear Localization ◽

Nuclear Export ◽

Binding Protein ◽

Actin Binding ◽

Actin Binding Protein

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Identification and characterization of a nuclear localization sequence-binding protein in yeast.

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.86.22.8808 ◽

1989 ◽

Vol 86 (22) ◽

pp. 8808-8812 ◽

Cited By ~ 26

Author(s):

W. C. Lee ◽

T. Melese

Keyword(s):

Nuclear Localization ◽

Binding Protein ◽

Nuclear Localization Sequence ◽

Localization Sequence ◽

Identification And Characterization

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Actin binding to WH2 domains regulates nuclear import of the multifunctional actin regulator JMY

Molecular Biology of the Cell ◽

10.1091/mbc.e11-12-0992 ◽

2012 ◽

Vol 23 (5) ◽

pp. 853-863 ◽

Cited By ~ 41

Author(s):

J. Bradley Zuchero ◽

Brittany Belin ◽

R. Dyche Mullins

Keyword(s):

Dna Damage ◽

Actin Filament ◽

Nuclear Localization ◽

Nuclear Import ◽

Actin Polymerization ◽

Regulatory Protein ◽

Actin Binding ◽

Nuclear Localization Sequence ◽

Filament Assembly ◽

Localization Sequence

Junction-mediating and regulatory protein (JMY) is a regulator of both transcription and actin filament assembly. In response to DNA damage, JMY accumulates in the nucleus and promotes p53-dependent apoptosis. JMY's actin-regulatory activity relies on a cluster of three actin-binding Wiskott–Aldrich syndrome protein homology 2 (WH2) domains that nucleate filaments directly and also promote nucleation activity of the Arp2/3 complex. In addition to these activities, we find that the WH2 cluster overlaps an atypical, bipartite nuclear localization sequence (NLS) and controls JMY's subcellular localization. Actin monomers bound to the WH2 domains block binding of importins to the NLS and prevent nuclear import of JMY. Mutations that impair actin binding, or cellular perturbations that induce actin filament assembly and decrease the concentration of monomeric actin in the cytoplasm, cause JMY to accumulate in the nucleus. DNA damage induces both cytoplasmic actin polymerization and nuclear import of JMY, and we find that damage-induced nuclear localization of JMY requires both the WH2/NLS region and importin β. On the basis of our results, we propose that actin assembly regulates nuclear import of JMY in response to DNA damage.

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