The localization of the divergent beta 2-tubulin isotype in the microtubular arrays of Physarum polycephalum

1989 ◽  
Vol 94 (2) ◽  
pp. 217-226
Author(s):  
M. Diggins-Gilicinski ◽  
L. Solnica-Krezel ◽  
T.G. Burland ◽  
E.C. Paul ◽  
W.F. Dove
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Polyclonal Antibody ◽  
Mitotic Spindle ◽  
Subcellular Distribution ◽  
Physarum Polycephalum ◽  
Mitotic Spindles ◽  
Tubulin Isotypes ◽  
Tubulin Isotype ◽  
Beta 2

The beta 2-tubulin isotype of Physarum polycephalum is only 83% identical in amino acid sequence with the constitutively expressed beta 1B-tubulin and the myxamoeba-specific beta 1A-tubulin isotypes. A polyclonal antibody specific for beta 2-tubulin was used to monitor the subcellular distribution of the beta 2-tubulin antigen in the mitotic spindle of the mature plasmodium - the sole microtubular array in that stage of Physarum. By immunofluorescence, the beta 2-tubulin antigen was detected throughout this anastral mitotic spindle, at all stages of mitosis. Physarum myxamoebae contain astral mitotic spindles and cytoskeletal microtubules. No beta 2-tubulin antigen was detected in the myxamoebal stage. However, as cultures of myxamoebae developed into plasmodia, the beta 2-tubulin antigen was found in the astral mitotic spindles and cytoskeletons in developing cells. Thus, the presence of the plasmodial beta 2-tubulin isotype in a mitotic spindle does not determine a closed, anastral mitosis.

scholarly journals Variable pathways for developmental changes of mitosis and cytokinesis in Physarum polycephalum.

1991 ◽  
Vol 113 (3) ◽  
pp. 591-604 ◽  
Author(s):  
L Solnica-Krezel ◽  
T G Burland ◽  
W F Dove
Keyword(s):  
Cell Cycle ◽  
Mitotic Spindle ◽  
Physarum Polycephalum ◽  
Developmental Changes ◽  
Binucleate Cell ◽  
Mitotic Spindles ◽  
Tubulin Isotypes ◽  
Binucleate Cells ◽  
Microscopic Studies ◽  
Mitotic Figures

The development of a uninucleate ameba into a multinucleate, syncytial plasmodium in myxomycetes involves a change from the open, astral mitosis of the ameba to the intranuclear, anastral mitosis of the plasmodium, and the omission of cytokinesis from the cell cycle. We describe immunofluorescence microscopic studies of the amebal-plasmodial transition (APT) in Physarum polycephalum. We demonstrate that the reorganization of mitotic spindles commences in uninucleate cells after commitment to plasmodium formation, is completed by the binucleate stage, and occurs via different routes in individual developing cells. Most uninucleate developing cells formed mitotic spindles characteristic either of amebae or of plasmodia. However, chimeric mitotic figures exhibiting features of both amebal and plasmodial mitoses, and a novel star microtubular array were also observed. The loss of the ameba-specific alpha 3-tubulin and the accumulation of the plasmodium-specific beta 2-tubulin isotypes during development were not sufficient to explain the changes in the organization of mitotic spindles. The majority of uninucleate developing cells undergoing astral mitoses (amebal and chimeric) exhibited cytokinetic furrows, whereas cells with the anastral plasmodial mitosis exhibited no furrows. Thus, the transition from astral to anastral mitosis during the APT could be sufficient for the omission of cytokinesis from the cell cycle. However, astral mitosis may not ensure cytokinesis: some cells undergoing amebal or chimeric mitosis contained unilateral cytokinetic furrows or no furrow at all. These cells would, most probably, fail to divide. We suggest that a uninucleate committed cell undergoing amebal or chimeric mitosis can either divide or else form a binucleate cell. In contrast, a uninucleate cell with a mitotic spindle of the plasmodial type gives rise only to a binucleate cells. Further, the decision to enter mitosis after commitment to the APT is independent of the developmental changes in the organization of the mitotic spindle and cytokinesis.

Variable pathways for developmental changes in composition and organization of microtubules in Physarum polycephalum

1990 ◽  
Vol 96 (3) ◽  
pp. 383-393
Author(s):  
L. Solnica-Krezel ◽  
M. Diggins-Gilicinski ◽  
T.G. Burland ◽  
W.F. Dove
Keyword(s):  
Physarum Polycephalum ◽  
Microtubule Organizing Center ◽  
Developmentally Regulated ◽  
Tubulin Isotypes ◽  
Cellular Events ◽  
Organizing Center ◽  
Cytoplasmic Microtubules ◽  
Beta 2 ◽  
Rare Cells ◽  
Microtubular Structures

The development of uninucleate amoebae into multinucleate plasmodia in myxomycetes is called the amoebal-plasmodial transition (APT). During the APT in Physarum polycephalum the ability to form flagellar axonemes is lost; the astral, open mitosis is replaced by the anastral, closed mitosis; and cytoskeletal microtubules disappear. These changes are accompanied by alterations in the repertoire of expressed tubulins. Using immunofluorescence microscopy we have studied the timing of loss and accumulation of developmentally regulated tubulin isotypes in relation to other cellular events during the APT. We specifically asked whether changes in the composition of microtubules are correlated with changes in their organization. The plasmodium-specific beta 2-tubulin can first be detected in microtubules of uninucleate cells after they become committed to plasmodium formation. However, rare cells are observed that exhibit beta 2-tubulin at earlier or only at later stages of development. Amoeba-specific acetylated alpha 3-tubulin disappears gradually during development. Individual cells differ in the timing of loss of this isotype: alpha 3-tubulin is present in the majority of uninucleate cells, in a fraction of binucleate and quadrinucleate cells, and is absent from larger multinucleate cells. Cytoplasmic microtubules in uninucleate cells are organized by a single microtubule-organizing center (MTOC) juxtaposed to the nucleus. Binucleate cells and quadrinucleate cells exhibit variable numbers of MTOCs. Cytoplasmic microtubules persist during the APT until the stage of plasmodia containing at least 100 nuclei. The lack of a strict correlation between the changes in tubulin composition and changes in organization of microtubular structures indicates that accumulation of beta 2-tubulin and disappearance of alpha 3-tubulin isotypes are not sufficient to bring about reorganization of microtubules during development. Individual cells in a developing population differ not only in the succession of accumulation and loss of developmentally regulated tubulins, but also in the sequences of other cellular changes occurring during the APT.

scholarly journals A gene encoding the major beta tubulin of the mitotic spindle in Physarum polycephalum plasmodia.

1988 ◽  
Vol 8 (3) ◽  
pp. 1275-1281 ◽  
Author(s):  
T G Burland ◽  
E C Paul ◽  
M Oetliker ◽  
W F Dove
Keyword(s):  
Mitotic Spindle ◽  
Physarum Polycephalum ◽  
Fruit Fly ◽  
Beta Tubulin ◽  
Gene Encoding ◽  
Neutral Drift ◽  
Beta 2 ◽  
Restricted Use ◽  
Microtubular Organelles ◽  
Plasmodium Of Physarum Polycephalum

The multinucleate plasmodium of Physarum polycephalum is unusual among eucaryotic cells in that it uses tubulins only in mitotic-spindle microtubules; cytoskeletal, flagellar, and centriolar microtubules are absent in this cell type. We have identified a beta-tubulin cDNA clone, beta 105, which is shown to correspond to the transcript of the betC beta-tubulin locus and to encode beta 2 tubulin, the beta tubulin expressed specifically in the plasmodium and used exclusively in the mitotic spindle. Physarum amoebae utilize tubulins in the cytoskeleton, centrioles, and flagella, in addition to the mitotic spindle. Sequence analysis shows that beta 2 tubulin is only 83% identical to the two beta tubulins expressed in amoebae. This compares with 70 to 83% identity between Physarum beta 2 tubulin and the beta tubulins of yeasts, fungi, alga, trypanosome, fruit fly, chicken, and mouse. On the other hand, Physarum beta 2 tubulin is no more similar to, for example, Aspergillus beta tubulins than it is to those of Drosophila melanogaster or mammals. Several eucaryotes express at least one widely diverged beta tubulin as well as one or more beta tubulins that conform more closely to a consensus beta-tubulin sequence. We suggest that beta-tubulins diverge more when their expression pattern is restricted, especially when this restriction results in their use in fewer functions. This divergence among beta tubulins could have resulted through neutral drift. For example, exclusive use of Physarum beta 2 tubulin in the spindle may have allowed more amino acid substitutions than would be functionally tolerable in the beta tubulins that are utilized in multiple microtubular organelles. Alternatively, restricted use of beta tubulins may allow positive selection to operate more freely to refine beta-tubulin function.

scholarly journals Purification and partial amino acid sequence of the actin-fragmin kinase from Physarum polycephalum

1993 ◽  
Vol 214 (1) ◽  
pp. 111-119 ◽  
Author(s):  
Jan GETTEMANS ◽  
Yvette VILLE ◽  
Joel VANDEKERCKHOVE ◽  
Etienne WAELKENS
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Physarum Polycephalum ◽  
Partial Amino Acid Sequence

scholarly journals The mammalian beta-tubulin repertoire: hematopoietic expression of a novel, heterologous beta-tubulin isotype.

1986 ◽  
Vol 103 (5) ◽  
pp. 1903-1910 ◽  
Author(s):  
D Wang ◽  
A Villasante ◽  
S A Lewis ◽  
N J Cowan
Keyword(s):  
Bone Marrow ◽  
Amino Acid ◽  
Cdna Library ◽  
Polypeptide Chain ◽  
Mouse Bone Marrow ◽  
Amino Acid Substitutions ◽  
Beta Tubulin ◽  
Tubulin Isotype ◽  
Beta 2 ◽  
Carboxy Terminal

We describe the structure of a novel and unusually heterologous beta-tubulin isotype (M beta 1) isolated from a mouse bone marrow cDNA library, and a second isotype (M beta 3) isolated from a mouse testis cDNA library. Comparison of M beta 1 and M beta 3 with the completed (M beta 4, M beta 5) or extended (M beta 2) sequence of three previously described beta-tubulin isotypes shows that each includes a distinctive carboxy-terminal region, in addition to multiple amino acid substitutions throughout the polypeptide chain. In every case where a mammalian interspecies comparison can be made, both the carboxy-terminal and internal amino acid substitutions that distinguish one isotype from another are absolutely conserved. We conclude that these characteristic differences are important in determining functional distinctions between different kinds of microtubule. The amino acid homologies between M beta 2, M beta 3, M beta 4, and M beta 5 are in the range of 95-97%; however the homology between M beta 1 and all the other isotypes is very much less (78%). The dramatic divergence in M beta 1 is due to multiple changes that occur throughout the polypeptide chain. The overall level of expression of M beta 1 is low, and is restricted to those tissues (bone marrow, spleen, developing liver and lung) that are active in hematopoiesis in the mouse. We predict that the M beta 1 isotype is functionally specialized for assembly into the mammalian marginal band.

scholarly journals The second subunit of methanol dehydrogenase of Methylobacterium extorquens AM1

1989 ◽  
Vol 260 (3) ◽  
pp. 857-862 ◽  
Author(s):  
D N Nunn ◽  
D Day ◽  
C Anthony
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Methanol Dehydrogenase ◽  
Beta Subunit ◽  
Methylobacterium Extorquens ◽  
Methylobacterium Extorquens Am1 ◽  
Beta 2

The nucleotide and deduced amino acid sequence of a novel small (beta) subunit of methanol dehydrogenase of Methylobacterium extorquens AM1 (previously Pseudomonas AM1) has been determined. Work with the whole protein has shown that is has an alpha 2 beta 2 configuration.

Tektin B1 from ciliary microtubules: primary structure as deduced from the cDNA sequence and comparison with tektin A1

1993 ◽  
Vol 106 (3) ◽  
pp. 909-918 ◽  
Author(s):  
R. Chen ◽  
C.A. Perrone ◽  
L.A. Amos ◽  
R.W. Linck
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Sequence Homology ◽  
Molecular Design ◽  
Consensus Sequence ◽  
Coiled Coil ◽  
Predicted Amino Acid Sequence ◽  
Mitotic Spindles ◽  
Intermediate Filament Proteins ◽  
Duplication Events

Tektins are a class of proteins that form filamentous polymers in the walls of ciliary and flagellar microtubules, and they may also be present in centrioles, centrosomes and mitotic spindles. We report here the cloning and sequencing of a cDNA for ciliary tektin B1. Comparison of the predicted amino acid sequence of tektin B1 with the previously published sequence for tektin A1 reveals several features that better define this class of proteins. Like tektin A1, the central region of the tektin B1 polypeptide chain is predicted to form a coiled-coil rod, consisting of four major alpha-helical regions that are separated by non-helical linkers. Between the central rod domains of tektins A and B there is a 34%/20% amino acid sequence identity/similarity, including equivalent 50-residue segments containing 36 identities, and a high probability of long-range structural homology. The tektin polypeptide chains are divided into two major segments that have significant sequence homology to each other, both within a given tektin chain and between tektins A and B, indicative of gene duplication events. The tektins have a secondary structure and molecular design similar to, but a low primary sequence homology with, intermediate filament proteins. Unlike tektin A1, tektin B1 lacks any part of the C-terminal IFP consensus sequence.

Amino acid sequence data of α-tubulin from myxamoebae of Physarum polycephalum

1986 ◽  
Vol 192 (4) ◽  
pp. 919-924 ◽  
Author(s):  
Monika Singhofer-Wowra ◽  
Melvyn Little ◽  
Lesley Clayton ◽  
Peter Dawson ◽  
Keith Gull
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Physarum Polycephalum ◽  
Sequence Data ◽  
Amino Acid Sequence Data

scholarly journals Five mouse tubulin isotypes and their regulated expression during development.

1985 ◽  
Vol 101 (3) ◽  
pp. 852-861 ◽  
Author(s):  
S A Lewis ◽  
M G Lee ◽  
N J Cowan
Keyword(s):  
Developmental Expression ◽  
Beta Tubulin ◽  
Alpha Tubulin ◽  
Tubulin Isotypes ◽  
Tubulin Genes ◽  
Mouse Tissues ◽  
Tubulin Isotype ◽  
Specific Mrnas ◽  
Beta 2 ◽  
Carboxy Terminal

We describe five mouse tubulin cloned cDNAs, two (M alpha 1 and M alpha 2) that encode alpha-tubulin and three (M beta 2, M beta 4, and M beta 5) that encode beta-tubulin. The sequence of these clones reveals that each represents a distinct gene product. Within the sequence common to the two alpha-tubulin cDNAs, the encoded amino acids are identical, though the 3' noncoding regions are wholly dissimilar. In contrast, the three beta-tubulin cDNAs show considerable carboxy-terminal heterogeneity. Two of the beta-tubulin isotypes defined by the cloned sequences are absolutely conserved between mouse and human, and all three beta-tubulin isotypes are conserved between mouse and rat. This result implies the existence of selective constraints that have maintained sequence identity after species divergence. This conclusion is reinforced by the near identity between a third mouse beta-tubulin isotype and a chicken beta-tubulin isotype. The significance of the interspecies conservation of tubulin isotypes is discussed in relationship to microtubule function. We have used non-cross-hybridizing 3' noncoding region probes from the five cloned mouse tubulin cDNAs to study the developmental expression of each isotype in various mouse tissues. M alpha 1 and M beta 2 are expressed in an approximately coordinate fashion, and their transcripts are most abundant in brain and lung. M alpha 2 and M beta 5 are ubiquitously expressed and to a similar extent in each tissue, with the greatest abundance in spleen, thymus, and immature brain. In contrast, M beta 4 is expressed exclusively in brain. Whereas the expression of the latter isotype increases dramatically during postnatal development, transcripts from all four other tubulin genes decline from maximum levels at or before birth. Tissue-specific development changes in the abundance of tubulin isotype-specific mRNAs are discussed in relationship to organogenesis in the mouse.

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