The Euplotes raikovi pheromone family: Identification of a sequence segment of potential importance for a distinction into subfamilies

NMR structure of the Euplotes raikovi pheromone E r -23 and identification of its five disulfide bonds 1 1Edited by M. F. Summers

“…7,9 The biological signi®cance of the present work lies in the observation that there is a new pheromone with major global architectural differences from all other Erpheromones, which nonetheless participates in cellcell signaling with cells expressing the short GA group pheromone Er-20. 8,11,32 This implies that the previous attempts at rationalizing apparent structure-function correlations in Er-pheromones will need to be re-evaluated critically. The expanded range of structural motifs including the, so far, unique Er-23 can then serve as a new platform for investigations of the mode of action of these pheromones.…”

A Structurally Deviant Member of the Euplotes raikovi Pheromone Family: Er‐23

“…The Er‐23 cloned macronuclear gene, derived from the RATE‐PCR strategy (see Materials and Methods) was found to comprise 989 nucleotides, and was thus about 100 nucleotides shorter than any other of the presently known E. raikovi pheromone genes (Miceli, La Terza, and Melli 1989; Vallesi et al 1996). It was proven to be complete by a Southern blot analysis of undigested macronuclear DNA preparations carried out using the insert of the Er‐23 cDNA clone as probe (Fig.…”

“…In addition, studies on the structure of Stylonychia tubulin genes (Helftenbein et al 1989) suggest that polyadenylation is specified by inverted repeats, which are located close to the polyadenylation site and contain the motif TTTATTG (Fig. 3), a conserved motif in all E. raikovi pheromone genes (Vallesi et al 1996; G. D. G. and C. M., unpubl. data).…”

“…This is illustrated, for instance, by two groups of E. raikovi strains, one denoted PR and the other GA (with reference to the names of their collection sites, Porto Recanati and Gaeta on the Eastern and Western coasts of Italy, respectively). As a general rule, a strain of the PR group can be induced to form mating pairs by pheromones of members of its group, but does not respond to pheromones of GA strains; similarly, a strain of the GA group can form mating pairs after interaction with pheromones of other members of the GA group, but does not respond to pheromones of PR strains (Vallesi et al 1996). In any case, PR and GA strains, although unable to mate with each other, are not genetically separated.…”

“…All the pheromones analyzed to date (i.e. E r ‐1 (= E r ‐3), E r ‐2 (= E r ‐9), E r ‐7, E r ‐10, and E r ‐11 from PR cells; and E r ‐20, E r ‐21, and E r ‐22 from GA cells) are proteins of 37 and 40 amino acids, with an aspartic acid at the amino‐terminus and six cysteines paired into three highly conserved disulfide bonds (Raffioni et al 1992; Stewart et al 1992; Vallesi et al 1996). Their three‐dimensional conformations are all based on a bundle of three α helices arranged with nearly parallel axes and an up‐down‐up topology (Brown et al 1993; Luginbühl et al 1994, 1996).…”