The two beta-tubulin genes of Chlamydomonas reinhardtii code for identical proteins.

Abstract

The two "-tubulin genes of the unicellular green alga Chiamydomonas reinhardtii are expressed coordinately after deflagellation and produce two transcripts of 2.1 and 2.0 kiobases. Full-length cDNA clones corresponding to the transcript of each gene were isolated. DNA sequences were obtained from the cDNA clones and from cloned tubulin gene fragments. Both genes contained 1,332 base pairs of coding sequence, with only 19 nucleotide differences between the genes. Because all the differences occurred at the third base position of a codon and did not change the predicted amino acid sequence, we concluded that both 0-tubulin genes code for the same protein of 443 amino acids. The predicted amino acid sequence is 89 and 72% homologous with Dtubulins from chicken and yeast cells, respectively. Each gene had three intervening sequences, which occurred at identical positions. Although the first two intervening sequences were not conserved between the two genes, the nucleotide sequence of the third intervening sequence was 89% conserved between the genes. The codon usage in the tubulin genes of C. reinhardhii was very biased: only 37 different codons were used. Striking differences occurred between the codons used in these nuclear genes and C. reinhardtii chloroplast genes.

Molecular cloning and expression of flagellar radial spoke and dynein genes of Chlamydomonas.

“…Transcripts from all four genes appear to accumulate coordinately during flagellar regeneration (5,39,43), and sequence comparisons among these genes have revealed several conserved 5' flanking regions (4) as well as an intriguing homology among all the genes in one of their intervening sequences (38,43). These conserved sequences are potential control regions for the induction of tubulin mRNA synthesis during flagellar regeneration, but because tubulin is used by the cell in organelles other than the flagellum, such as the mitotic spindle, there is a possibility that these sequences function to regulate tubulin gene expression during cellular events other than the production of new flagella.…”

Coordinate and selective beta-tubulin gene expression associated with cilium formation in sea urchin embryos.

“…Triggering mechanisms have been sought to account for the coordinate expression of the duplicate B-and ~-tubulin genes of Chlamydomonas induced by deflagellation (Brunke et al 1984;Youngblom et al 1984;Siflow and Rosenbaum 1985). However, in this case, as well as for the single tubulin genes of Tetrahymena (Callahan et al 1984), the lack of large multigene tubulin families obviates the question of whether certain tubulin genes may be selectively excluded from induction.…”

“…The major proteins of axonemal microtubules, the ~-and ~-tubulins, need to be synthesized for complete flagellum regeneration in Chlamydomonas (Rosenbaum et al 1969) and may be drawn from a preexisting pool, as in the case of cilium regeneration in Tetrahymena (Guttman and Gorovsky 1979). Deflagellation induces the coordinately increased transcription of the two ~-and two f3-tubulin genes of Chlamydomonas (Brunke et al 1984;Youngblom et al 1984;Silflow and Rosenbaum 1985) as well as the stabilizaiton of tubulin mRNA (Baker et al 1984). In the case of the sea urchin embryo, cilium regeneration could be demonstrated to proceed through an initial phase that is independent of new protein synthesis and a second phage that depends on protein synthesis (Bums 1979).…”

The CRY1 gene in Chlamydomonas reinhardtii: structure and use as a dominant selectable marker for nuclear transformation.

“…The CRYl-] allele used for these experiments encompassed the entire transcribed region, plus 250 bp of upstream sequence and 425 bp downstream of the putative polyadenylation signal (TGTAA) (Fig. 3) (49). In cotransformations of plasmid pCRY1-1 with pMN24 (transformation method 1; Materials and Methods), 6% of the Nit' transformants were more resistant to cryptopleurine than the untransformed strain, but the resistance obtained was not as high as that observed for the heterozygous diploid ( Table 1).…”

“…Approximately eight nuclear genomic equivalents (120,000 plaques) of an EMBL4 X library, constructed by using strain 21gr DNA (49), were screened with the CRY1 cDNA insert as described above. Eight X clones were purified, and their restriction maps were compared.…”