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 1-tubulin cDNA clone, 13105, which is shown to correspond to the trnscript of the beiC P-tubulin locus and to encode 12 tubulin, the 1 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 (2 tubulin is only 83% identical to the two ,3 tubulins expressed in amoebae. This compares with 70 to 83% identity between Physarum P12 tubulin and the 1 tubulins of yeasts, fungi, alga, trypanosome, fruit fly, chicken, and mouse. On the other hand, Physarum 12 tubulin is no more similar to, for example, Aspergillus 1 tubulins than it is to those of Drosophila melanogaster or mammals. Several eucaryotes express at least one widely diverged 1 tubulin as well as one or more 1 tubulins that conform more closely to a consensus 1-tubulin sequence. We suggest that ,B-tubulins diverge more when their expression pattern is restricted, especially when this restriction results in their use in fewer functions. This divergence among P tubulins could have resulted through neutral drift. For example, exclusive use of Physarum P2 tubulin in the spindle may have allowed more amino acid substitutions than would be functionally tolerable in the 1 tubulins that are utilized in multiple microtubular organelles.Alternatively, restricted use of P tubulins may allow positive selection to operate more freely to refine 3-tubulin function.Most eucaryotes express multiple genes each for a and K tubulins, the principal protein components of microtubules. The products of these genes usually have distinct sequences. For 13 tubulins, identity between multiple polypeptides within a single organism varies from 100% for Chlamydomonas reinhardtii (38) to 78% for mammals (34). The varied functions of microtubules in the cytoskeleton, centrioles, spindles, and flagella, together with the observed tubulin sequence differences, have inspired the conclusion that the sequence differences between multiple tubulins within an organism have been positively selected during evolution (32). The murine i tubulin M,1l is only 78% identical to several other murine i tubulins and is expressed specifically in hematopoietic tissues, prompting the speculation that at least some of the sequence differences reflect functional specialization of individual 1 tubulins (34).In the myxomycete Physarum polycephalum, threetubulin genes are known; one gene is expressed specifically in the amoebal cell type, another gene is expressed specifically in the plasmodial cell type, and the third gene, betB, is expressed in both amoebae and plasmodia (4, 28). The multinucleate plasmodium of P. polycephalum is unusual among eucaryotic cells in that it uses tubulins o...

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.