Microtubule Binding Proteins Are Not Necessarily Microtubule-Associated Proteins

1994 ◽  
Vol 6 (12) ◽  
pp. 1696
Author(s):  
Louis C. Morejohn
Keyword(s):  
Binding Proteins ◽  
Microtubule Binding ◽  
Microtubule Associated Proteins ◽  
Associated Proteins

scholarly journals KIF18A's neck linker permits navigation of microtubule-bound obstacles within the mitotic spindle

2019 ◽  
Vol 2 (1) ◽  
pp. e201800169 ◽  
Author(s):  
Heidi LH Malaby ◽  
Dominique V Lessard ◽  
Christopher L Berger ◽  
Jason Stumpff
Keyword(s):  
Single Molecule ◽  
Mitotic Spindle ◽  
Binding Proteins ◽  
Mitotic Chromosome ◽  
Chromosome Movement ◽  
Wild Type ◽  
Microtubule Associated Proteins ◽  
Chromosome Alignment ◽  
Associated Proteins ◽  
Fiber Dynamics

KIF18A (kinesin-8) is required for mammalian mitotic chromosome alignment. KIF18A confines chromosome movement to the mitotic spindle equator by accumulating at the plus-ends of kinetochore microtubule bundles (K-fibers), where it functions to suppress K-fiber dynamics. It is not understood how the motor accumulates at K-fiber plus-ends, a difficult feat requiring the motor to navigate protein dense microtubule tracks. Our data indicate that KIF18A's relatively long neck linker is required for the motor's accumulation at K-fiber plus-ends. Shorter neck linker (sNL) variants of KIF18A display a deficiency in accumulation at the ends of K-fibers at the center of the spindle. Depletion of K-fiber–binding proteins reduces the KIF18A sNL localization defect, whereas their overexpression reduces wild-type KIF18A's ability to accumulate on this same K-fiber subset. Furthermore, single-molecule assays indicate that KIF18A sNL motors are less proficient in navigating microtubules coated with microtubule-associated proteins. Taken together, these results support a model in which KIF18A's neck linker length permits efficient navigation of obstacles to reach K-fiber ends during mitosis.

scholarly journals Cytoplasmic Dynein Mediates Adenovirus Binding to Microtubules

2004 ◽  
Vol 78 (18) ◽  
pp. 10122-10132 ◽  
Author(s):  
Samir A. Kelkar ◽  
K. Kevin Pfister ◽  
Ronald G. Crystal ◽  
Philip L. Leopold
Keyword(s):  
Molecular Motor ◽  
Retrograde Transport ◽  
Cytoplasmic Dynein ◽  
Bovine Brain ◽  
Microtubule Binding ◽  
Microtubule Associated Proteins ◽  
Primary Mode ◽  
Cell Lysate ◽  
Brain Maps ◽  
Associated Proteins

ABSTRACT During infection, adenovirus (Ad) capsids undergo microtubule-dependent retrograde transport as part of a program of vectorial transport of the viral genome to the nucleus. The microtubule-associated molecular motor, cytoplasmic dynein, has been implicated in the retrograde movement of Ad. We hypothesized that cytoplasmic dynein constituted the primary mode of association of Ad with microtubules. To evaluate this hypothesis, an Ad-microtubule binding assay was established in which microtubules were polymerized with taxol, combined with Ad in the presence or absence of microtubule-associated proteins (MAPs), and centrifuged through a glycerol cushion. The addition of purified bovine brain MAPs increased the fraction of Ad in the microtubule pellet from 17.3% ± 3.5% to 80.7% ± 3.8% (P < 0.01). In the absence of tubulin polymerization or in the presence of high salt, no Ad was found in the pellet. Ad binding to microtubules was not enhanced by bovine brain MAPs enriched for tau protein or by the addition of bovine serum albumin. Enhanced Ad-microtubule binding was also observed by using a fraction of MAPs purified from lung A549 epithelial cell lysate which contained cytoplasmic dynein. Ad-microtubule interaction was sensitive to the addition of ATP, a hallmark of cytoplasmic dynein-dependent microtubule interactions. Immunodepletion of cytoplasmic dynein from the A549 cell lysate abolished the MAP-enhanced Ad-microtubule binding. The interaction of Ad with both dynein and dynactin complexes was demonstrated by coimmunoprecipitation. Partially uncoated capsids isolated from cells 40 min after infection also exhibited microtubule binding. In summary, the primary mode of Ad attachment to microtubules occurs though cytoplasmic dynein-mediated binding.

scholarly journals Functionally distinct isoforms of dynactin are expressed in human neurons.

1996 ◽  
Vol 7 (8) ◽  
pp. 1167-1180 ◽  
Author(s):  
M K Tokito ◽  
D S Howland ◽  
V M Lee ◽  
E L Holzbaur
Keyword(s):  
Cytoplasmic Dynein ◽  
Binding Motif ◽  
Binding Assays ◽  
Microtubule Binding ◽  
Microtubule Associated Proteins ◽  
Human Neurons ◽  
Alternative Mrna Splicing ◽  
Associated Proteins ◽  
Dynactin Complex

P150Glued is the largest subunit of dynactin, which binds to cytoplasmic dynein and activates vesicle transport along microtubules. We have isolated human cDNAs encoding p150Glued as well as a 135-kDa isoform; these isoforms are expressed in human brain by alternative mRNA splicing of the human DCTN1 gene. The p135 isoform lacks the consensus microtubule-binding motif shared by members of the p150Glued/Glued/CLIP-170/BIK1 family of microtubule-associated proteins and, therefore, is predicted not to bind directly to microtubules. We used transient transfection assays and in vitro microtubule-binding assays to demonstrate that the p150 isoform binds to microtubules, but the p135 isoform does not. However, both isoforms bind to cytoplasmic dynein, and both partition similarly into cytosolic and membrane cellular fractions. Sequential immunoprecipitations with an isoform-specific antibody for p150 followed by a pan-isoform antibody revealed that, in brain, these polypeptides assemble to form distinct complexes, each of which sediments at approximately 20 S. On the basis of these observations, we hypothesize that there is a conserved neuronal function for a distinct form of the dynactin complex that cannot bind directly to cellular microtubules.

scholarly journals Domains of Neuronal Microtubule-associated Proteins and Flexural Rigidity of Microtubules

1997 ◽  
Vol 138 (5) ◽  
pp. 1067-1075 ◽  
Author(s):  
Harald Felgner ◽  
Rainer Frank ◽  
Jacek Biernat ◽  
Eva-Maria Mandelkow ◽  
Eckhard Mandelkow ◽  
...  
Keyword(s):  
Optical Tweezers ◽  
Flexural Rigidity ◽  
Microtubule Binding ◽  
Fourfold Increase ◽  
Microtubule Associated Proteins ◽  
Relative Contribution ◽  
Low Concentrations ◽  
Neuronal Processes ◽  
Associated Proteins ◽  
And Function

Microtubules are flexible polymers whose mechanical properties are an important factor in the determination of cell architecture and function. It has been proposed that the two most prominent neuronal microtubule-associated proteins (MAPs), tau and MAP2, whose microtubule binding regions are largely homologous, make an important contribution to the formation and maintenance of neuronal processes, putatively by increasing the rigidity of microtubules. Using optical tweezers to manipulate single microtubules, we have measured their flexural rigidity in the presence of various constructs of tau and MAP2c. The results show a three- or fourfold increase of microtubule rigidity in the presence of wild-type tau or MAP2c, respectively. Unexpectedly, even low concentrations of MAPs promote a substantial increase in microtubule rigidity. Thus at ∼20% saturation with full-length tau, a microtubule exhibits &gt;80% of the rigidity observed at near saturating concentrations. Several different constructs of tau or MAP2 were used to determine the relative contribution of certain subdomains in the microtubule-binding region. All constructs tested increase microtubule rigidity, albeit to different extents. Thus, the repeat domains alone increase microtubule rigidity only marginally, whereas the domains flanking the repeats make a significant contribution. Overall, there is an excellent correlation between the strength of binding of a MAP construct to microtubules (as represented by its dissociation constant Kd) and the increase in microtubule rigidity. These findings demonstrate that neuronal MAPs as well as constructs derived from them increase microtubule rigidity, and that the changes in rigidity observed with different constructs correlate well with other biochemical and physiological parameters.

scholarly journals A novel microtubule-binding motif identified in a high molecular weight microtubule-associated protein from Trypanosoma brucei

1992 ◽  
Vol 117 (1) ◽  
pp. 95-103 ◽  
Author(s):  
A Hemphill ◽  
M Affolter ◽  
T Seebeck
Keyword(s):  
Molecular Weight ◽  
Amino Acid ◽  
Trypanosoma Brucei ◽  
Binding Motif ◽  
Amino Acid Repeat ◽  
Microtubule Binding ◽  
Microtubule Associated Proteins ◽  
Repeat Units ◽  
Associated Proteins

The major component of the cytoskeleton of the parasitic hemoflagellate Trypanosoma brucei is a membrane skeleton which consists of a single layer of tightly spaced microtubules. This array encloses the entire cell body, and it is apposed to, and connected with, the overlying cell membrane. The microtubules of this array contain numerous microtubule-associated proteins. Prominent among those is a family of high molecular weight, repetitive proteins which consist to a large extent of tandemly arranged 38-amino acid repeat units. The binding of one of these proteins, MARP-1, to microtubules has now been characterized in vitro and in vivo. MARP-1 binds to microtubules via tubulin domains other than the COOH-termini used by microtubule-associated proteins from mammalian brain, e.g., MAP2 or Tau. In vitro binding assays using recombinant protein, as well as transfection of mammalian cell lines, have established that the repetitive 38-amino acid repeat units represent a novel microtubule-binding motif. This motif is very similar in length to those of the mammalian microtubule-associated proteins Tau, MAP2, and MAP-U, but both its sequence and charge are different. The observation that the microtubule-binding motifs both of the neural and the trypanosomal proteins are of similar length may reflect the fact that both mediate binding to the same repetitive surface, the microtubule, while their sequence and charge differences are in agreement with the observation that they interact with different domains of the tubulins.

scholarly journals A Common Mechanism for Cytoplasmic Dynein-Dependent Microtubule Binding Shared among Adeno-Associated Virus and Adenovirus Serotypes

2006 ◽  
Vol 80 (15) ◽  
pp. 7781-7785 ◽  
Author(s):  
Samir Kelkar ◽  
Bishnu P. De ◽  
Guangping Gao ◽  
James M. Wilson ◽  
Ronald G. Crystal ◽  
...  
Keyword(s):  
Retrograde Transport ◽  
Cytoplasmic Dynein ◽  
Common Mechanism ◽  
Dependent Manner ◽  
Adeno Associated Virus ◽  
Microtubule Binding ◽  
Microtubule Associated Proteins ◽  
Associated Proteins ◽  
Serotype 5 ◽  
Vectorial Transport

ABSTRACT During infection, adenovirus-associated virus (AAV) undergoes microtubule-dependent retrograde transport as part of a program of vectorial transport of viral genome to the nucleus. A microtubule binding assay was used to evaluate the hypothesis that cytoplasmic dynein mediates AAV interaction with microtubules. Binding of AAV serotype 2 (AAV2) was enhanced in a nucleotide-dependent manner by the presence of total cellular microtubule-associated proteins (MAPs) but not cytoplasmic dynein-depleted MAPs. Excess AAV2 capsid protein prevented microtubule binding by AAV serotypes 2, 5, and rh.10, as well as adenovirus serotype 5, indicating that similar binding sites are used by these viruses.

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