betaTubulin56D (ß1 tubulin): Biological Overview | Evolutionary Homologs | Regulation | Protein Interactions | Developmental Biology | Effects of Mutation | References

Gene name - betaTubulin56D

Synonyms - ß1 tubulin

Cytological map position - 56C

Function - component of microtubules

Keyword(s) - cytoskeleton

Symbol - betaTub56D

FlyBase ID:FBgn0003887

Genetic map position - 2-[88]

Classification - beta tubulin

Cellular location - cytoplasmic



NCBI links: Precomputed BLAST | Entrez Gene
BIOLOGICAL OVERVIEW

Drosophila has four ß-tubulin genes: each encodes a distinct isoform. In the oocyte, the microtubule cytoskeleton plays diverse roles both in the prepositioning of developmental cues (see also Bicoid, Oskar and Nanos) and in cytoplasmic reorganization (see cytoskeleton) of the oocyte (Theurkauf, 1993 and 1994). These early oocyte functions are supported by microtubules assembled from the maternally supplied ß1-tubulin isoform. Beta tubulins are found dimerized with alpha tubulins for which there are also four isoforms in Drosophila. In most cases an alpha tubulin - beta tubulin heterodimer is the functional form of tubulin in the cell.

One remarkable microtubule function is a change in microtubule polarity that takes place between stage 6 and stage 7 of oogenesis. At stage 6, a microtubule organizing center (MTOC) is present at the posterior pole. By stage 7 this center is no longer detected, and new microtubules are present at the anterior margin of the oocyte; this area appears to act as a microtubule nucleating region. Protein Kinase A (PKA) mutations act in the germ line to disrupt both microtubule distribution and RNA localization along the AP axis. In normal oocytes, the site of microtubule nucleation shifts immediately prior to polarized localization of Bicoid and Oskar mRNAs.

The shift in polarity can be detected using using a fusion protein of kinesin with lacZ. Kinesin is a motor protein that "walks" along microtubules from the minus end (where tubulin polymerization starts) to the plus end (Dyneins travel from plus ends to minus ends). LacZ is an enzyme whose reaction produces a colored product, allowing a visualization of Kinesin movement along microtubules. In PKA-deficient oocytes, the Kinesin fusion protein fails to accumulate at the posterior pole by stage 9, although this is the expected location in normal eggs. In PKA mutants, posterior microtubules are present during this transition; OskarmRNA fails to accumulate at the posterior, while BicoidmRNA accumulates at both ends of the oocyte. Similar RNA mislocalization patterns for Notch and Delta mutants suggest that PKA transduces a signal for microtubule reorganization that is sent by posteriorly located follicle cells (Lane, 1994).

Maternally encoded ß1 tubulin is incorporated into zygotic mitotic spindles. Thus ß1 tubulin is the primary ß tubulin component of the mitotic apparatus. Gamma tubulin has an important role, however as a nucleating substrate for ß1 tubulin polymerization in the mitotic apparatus. Later in development a strong expression of ß1 is observed in the central nervous system and in the epidermis. All chordotonal organs and apodemes (muscle attachment sites within the epidermis) are marked by ß1 tubulin. In Drosophila testis stem cells, ß1 tubulin is found in mitotically active germ cells and all somatic parts of the testis, but starting with early spermatocytes, the ß1 isotype is switched off and all microtubular arrays contain ß2 tubulin (Raff, 1982, Bialojan, 1984, Gasch, 1988, Buttgereit, 1991 and Buttgereit, 1993b).

The complex expression pattern of ß1 tubulin suggests an equally complex regulation. At least four regulatory regions are involved. Maternal expression is regulated by the sequences located between -2.2 kb and the start site, that is within the first 2.2 kb of ß1 upstream sequences. Enhancer elements are located in the intron (between +0.44 kb and +2.5 kb) that drive expression in the chordotonal organs, the CNS and the apodemes (Buttgereit, 1991). Expression in testis stem cells is regulated by a short sequence located between -45 and -191 upstream from the transcriptional start site, while independent regions in the first intron (+443 to +2525) drive expression in the cyst cells and paragonia, the somatic tissues of the male gonad. The tubulin isotype switch between ß1 and ß2 in the germ line can be attributed mostly to transcriptional control. Since ß1 mRNA and/or protein is very rapidly degraded in the transition from early to mid-spermatogenesis, other factors besides transcriptional control may be involved in the isotypes switch (Buttgereit, 1993b).

Expression in apodemes is of interest because presence of ß1 tubulin might provide mechanical support for the muscles to which they are attached. Within the apodemes, microtubules play an essential role as stabilizers against the stress forces generated by the inserted muscles. Only a few proteins are known to be involved in cell-cell adhesion and cell-cell communication in muscle attachment sites: integrins are involved in cell-cell adhesion, EGF-R is involved in cell-cell communication, and Laminin A and Glutactin, glycoproteins with sequences similar to serine esterases are present in the basement membrane. Stripe is an epidermal cell specific zinc finger transcription factor; Groovin appears to be the contact protein for myotubules (See Stripe).

The onset of myodeme expression is first detected at late stage 13. By this stage the fusion of myoblasts to mytubes has already occured, and now the muscles elongate and insert at their attachment sites within the epidermis. The first cells positive for ß1 tubulin are the intersegmental apodemes restricted to the dorsal side of the embryo. Ventrally at early stage 15 three types of apodemes stain for ß1: the intersegmental apodeme, and two intrasegmental apodemes. On the basis of classical observations, cells that express ß1 represent tendon cells, characterized by a high number of microtubule bundles. In insect species bundles of microtubules extend from the myoepidermal junction into the epidermis, where they insert in hemidesmosomes. It appears that cell-cell contact is necessary for the activation of a regulatory cascade leading to the activation of ß1 tubulin gene expression, but prior to such activation, the tendon cell fate has already been determined (Buttgereit, 1993a).


GENE STRUCTURE

Exons - 4


PROTEIN STRUCTURE

Amino Acids - 448

Structural Domains

Three of the four ß-tubulins of Drosophila exhibit an 87% homology in overall amino acid sequence. The first and second introns of ß3-tubulin occur in positions identical to those of the first two introns in the vertebrate beta-tubulin genes and are identical to those of ß1 tubulin. The fly ß3-tubulin contains an additional six amino acids not present in any other beta-tubulin. The position of the third intron differs from that of the vertebrate beta-tubulin genes, but is identical in position to the Drosophila testis-specific ß2 isoform. The testis-specific isoform and the ubiquitously expressed ß1 isoform are highly homologous both to each other and to the vertebrate major neural form. The 3' untranslated region of the ß1 mRNA has no homology to the corresponding regions of the ß2 tubulin mRNA. The ß3 isoform, still more divergent, reveals an ancient divergence in beta-tubulin sequences that must have occurred at or before the split between lines leading to insects and vertebrates. For ß tubulins there appear to be two major regions that are different in their secondary structures, that is in the folding of the polypeptide chain. A predicted alpha-helical region is present in the carboxyl third of the molecule, and a predicted beta-pleated sheet region falls primarily in the amino-terminal two-thirds of the molecule (Rudolph, 1987 and Michiels, 1987).

The ß1-, ß2- and ß3-tubulin-specific sequences in the chromosomes of Drosophila auraria are found in the same polytene band in region 32C of the 2L polytene chromosome. In contrast, the three beta tubulin genes in D. melanogaster are not closely linked (Scouras, 1994).

The alphabeta tubulin heterodimer is the structural subunit of microtubules, which are cytoskeletal elements essential for intracellular transport and cell division in all eukaryotes. Each tubulin monomer binds a guanine nucleotide, which is nonexchangeable when it is bound in the alpha subunit, or N site, and exchangeable when bound in the beta subunit, or E site. The alpha- and beta-tubulins share 40% amino-acid sequence identity; both exist in several isotype forms, and both undergo a variety of posttranslational modifications. Limited sequence homology has been found with the proteins FtsZ and Misato, which are involved in cell division in bacteria and Drosophila, respectively. An atomic model is presented of the alphabeta tubulin dimer fitted to a 3.7-A density map obtained by electron crystallography of zinc-induced tubulin sheets. The structures of alpha- and beta-tubulin are basically identical: each monomer is formed by a core of two beta-sheets surrounded by alpha-helices. The monomer structure is very compact, but can be divided into three functional domains: the amino-terminal domain, containing the nucleotide-binding region, an intermediate domain, containing the Taxol-binding site, and the carboxy-terminal domain, which probably constitutes the binding surface for motor proteins (Nogales, 1998).


betaTubulin56D (ß1 tubulin): Evolutionary Homologs | Regulation | Protein Interactions | Developmental Biology | Effects of Mutation | References
date revised: 25 March 98 

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