Bundling of microtubules in transfected cells does not involve an autonomous dimerization site on the MAP2 molecule.

Burgin, Kelly E.; Ludin, Beat; Ferralli, Jacqueline; Matus, Andrew

doi:10.1091/mbc.5.5.511

Cited by 27 publications

(16 citation statements)

References 34 publications

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“…In this situation, mitosis appears to be inhibited by the increased stabilization of the microtubule network (Fig. 8, pathway 1), a feature which is common to the overexpression of many other microtubule-stabilizing proteins (2,21,26,34,38). However, in another population of VP22-expressing cells, VP22 remains diffuse in the cytoplasm and upon entry into mitosis immediately associates with the condensed cellular chromatin where it remains throughout M phase and into G 1 of the next cell cycle.…”

Section: Discussionmentioning

confidence: 99%

Cytoplasm-to-Nucleus Translocation of a Herpesvirus Tegument Protein during Cell Division

Elliott¹,

O’Hare

2000

J Virol

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We have previously shown that the herpes simplex virus tegument protein VP22 localizes predominantly to the cytoplasm of expressing cells. We have also shown that VP22 has the unusual property of intercellular spread, which involves the movement of VP22 from the cytoplasm of these expressing cells into the nuclei of nonexpressing cells. Thus, VP22 can localize in two distinct subcellular patterns. By utilizing time-lapse confocal microscopy of live cells expressing a green fluorescent protein-tagged protein, we now report in detail the intracellular trafficking properties of VP22 in expressing cells, as opposed to the intercellular trafficking of VP22 between expressing and nonexpressing cells. Our results show that during interphase VP22 appears to be targeted exclusively to the cytoplasm of the expressing cell. However, at the early stages of mitosis VP22 translocates from the cytoplasm to the nucleus, where it immediately binds to the condensing cellular chromatin and remains bound there through all stages of mitosis and chromatin decondensation into the G 1 stage of the next cycle. Hence, in VP22-expressing cells the subcellular localization of the protein is regulated by the cell cycle such that initially cytoplasmic protein becomes nuclear during cell division, resulting in a gradual increase over time in the number of nuclear VP22-expressing cells. Importantly, we demonstrate that this process is a feature not only of VP22 expressed in isolation but also of VP22 expressed during virus infection. Thus, VP22 utilizes an unusual pathway for nuclear targeting in cells expressing the protein which differs from the nuclear targeting pathway used during intercellular trafficking.Herpesviruses have a well-defined replication phase within the nucleus, where they are known to exploit many of the cellular processes performed there. Upon virus entry into the host cell, the viral DNA genome is directed into the nucleus by an as-yet-undefined mechanism and is subsequently transcribed and replicated by a combination of host cell machinery and virus gene products (1,17,22). At later stages in the replication cycle, assembly of the herpesvirus particle is initiated within the nucleus as the newly replicated virus DNA genome is packaged into assembling capsids (39, 41). As a consequence, herpesviruses must target several classes of their gene products, including transcription factors, DNA replication factors, scaffold proteins, and capsid proteins, to the nucleus. A number of virus proteins, such as the immediate-early proteins ICP0 (13, 30) and ICP27 (19,28), the DNA replication protein encoded by gene UL9 (27), and the capsid protein VP19C (40), have been shown to contain classical nuclear localization signals (NLSs), which are defined in the primary amino acid sequence of these proteins (15, 32). Such NLScontaining proteins are translocated from the cytoplasm into the nucleus through the nuclear pores, a process mediated by cellular proteins typified by the heterodimeric complex of importin ␣ and ␤ proteins (15, 32). Th...

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Section: Discussionmentioning

confidence: 99%

Cytoplasm-to-Nucleus Translocation of a Herpesvirus Tegument Protein during Cell Division

Elliott¹,

O’Hare

2000

J Virol

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“…In mammalian cell lines, expression of MAP2 remodels endogenous microtubule structure, leading to the formation of bundles of microtubules near the perimeter of the cell (Weisshaar et al, 1992). MAP2 is not thought to physically cross-link microtubules (Burgin et al, 1994). Rather, the bundles are comprised of microtubules that have become hyperstabilized and aligned in parallel arrays.…”

Section: Pka Activity Regulates Map2c Interactions With Microtubules mentioning

confidence: 99%

Phosphorylation-dependent Localization of Microtubule-associated Protein MAP2c to the Actin Cytoskeleton

Ozer

Halpain

2000

MBoC

109

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Microtubule-associated protein 2 (MAP2) is a neuronal phosphoprotein that promotes net microtubule growth and actin cross-linking and bundling in vitro. Little is known about MAP2 regulation or its interaction with the cytoskeleton in vivo. Here we investigate the in vivo function of three specific sites of phosphorylation on MAP2. cAMP-dependent protein kinase activity disrupts the MAP2-microtubule interaction in living HeLa cells and promotes MAP2c localization to peripheral membrane ruffles enriched in actin. cAMP-dependent protein kinase phosphorylates serines within three KXGS motifs, one within each tubulin-binding repeat. These highly conserved motifs are also found in homologous proteins tau and MAP4. Phosphorylation at two of these sites was detected in brain tissue. Constitutive phosphorylation at these sites was mimicked by single, double, and triple mutations to glutamic acid. Biochemical and microscopy-based assays indicated that mutation of a single residue was adequate to disrupt the MAP2-microtubule interaction in HeLa cells. Double or triple point mutation promoted MAP2c localization to the actin cytoskeleton. Specific association between MAP2c and the actin cytoskeleton was demonstrated by retention of MAP2c-actin colocalization after detergent extraction. Specific phosphorylation states may enhance the interaction of MAP2 with the actin cytoskeleton, thereby providing a regulated mechanism for MAP2 function within distinct cytoskeletal domains.

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“…4B, top). In contrast, MTs were thin and short in the presence of the MAP2 protein, which is unable to support MT bundling in vitro (5,28). Thin and short MTs were also observed when BSA was used as a negative control (data not shown).…”

mentioning

confidence: 93%

The Dual-Specificity Phosphatase CDC14B Bundles and Stabilizes Microtubules

Cho

Liu

Gomez

et al. 2005

Molecular and Cellular Biology

101

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The Cdc14 dual-specificity phosphatases regulate key events in the eukaryotic cell cycle. However, little is known about the function of mammalian CDC14B family members. Here, we demonstrate that subcellular localization of CDC14B protein is cell cycle regulated. CDC14B can bind, bundle, and stabilize microtubules in vitro independently of its catalytic activity. Basic amino acid residues within the nucleolar targeting domain are important for both retaining CDC14B in the nucleolus and preventing microtubule bundling. Overexpression of CDC14B resulted in the formation of cytoplasmic CDC14B and microtubule bundles in interphase cells. These microtubule bundles were resistant to microtubule depolymerization reagents and enriched in acetylated ␣-tubulin. Expression of cytoplasmic forms of CDC14B impaired microtubule nucleation from the microtubule organization center. CDC14B is thus a novel microtubule-bundling and -stabilizing protein, whose regulated subcellular localization may help modulate spindle and microtubule dynamics in mitosis.

show abstract

Bundling of microtubules in transfected cells does not involve an autonomous dimerization site on the MAP2 molecule.

Cited by 27 publications

References 34 publications

Cytoplasm-to-Nucleus Translocation of a Herpesvirus Tegument Protein during Cell Division

Cytoplasm-to-Nucleus Translocation of a Herpesvirus Tegument Protein during Cell Division

Phosphorylation-dependent Localization of Microtubule-associated Protein MAP2c to the Actin Cytoskeleton

The Dual-Specificity Phosphatase CDC14B Bundles and Stabilizes Microtubules

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