Identification of the tubulin gene family and sequence determination of one beta-tubulin gene in a cold-poikilotherm protozoan, the antarctic ciliate Euplotes focardii.
Four different tubulin genes were identified in the somatic nucleus (macronucleus) of Euplotes focardii, a strictly cold-adapted, Antarctic ciliate: one of 1,800 bp for alpha-tubulin and three of 2,150, 1,900, and 1,600 bp, respectively, for beta-tubulin. Preliminarily analysed for restriction fragment length polymorphisms, these genes showed remarkable differences in organisation from tubulin genes of other ciliates which live in temperate areas and were analysed in parallel with E. focardii. The complete coding sequence of the 1,600 bp beta-tubulin gene was then determined and shown to contain unique structural features of potential importance for E. focardii microtubule organization and activity. Of eight unique substitutions detected, seven were concentrated in the large amino terminal domain of the molecule that directly interacts with the carboxy terminal region of alpha-tubulin for heterodimer formation. Sequence analysis of the cloned gene revealed, in addition, a potential new exception in the use of the genetic code by ciliates. A TAG codon was aligned in correspondence with Trp-21 which is strictly conserved in every tubulin sequence so far determined.
Miceli
C
C
Dipartimento di Biologia Molecolare, Cellulare ed Animale Università di Camerino, Italy.
Ballarini
P
P
Di Giuseppe
G
G
Valbonesi
A
A
Luporini
P
P
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern; D003001:Cloning, Molecular; D016054:DNA, Protozoan / Q000032:analysis / Q000096:biosynthesis / Q000235:genetics*; D016812:Euplotes / Q000235:genetics* / Q000302:isolation & purification; D017125:Genes, Protozoan; D016678:Genome; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D015183:Restriction Mapping; D014404:Tubulin / Q000032:analysis / Q000235:genetics*
D016054:DNA, Protozoan; D014404:Tubulin