Purification and characterization of a pheromone that induces sexual cell division in the unicellular green alga Closterium ehrenbergii

Closterium ehrenbergii is a unicellular charophycean alga consisting of two sexes: mating type plus (mt+) and minus (mt–). The sexual reproductive process consists of five steps: formation of sexual pairs, cell division of each member of a pair, formation of conjugation papillae, release of protoplasts from gametangial cells, and fusion of protoplasts to form a zygote. The second step, called sexual cell division (SCD), produces two gametangial cells from one vegetative mother cell. The SCD of mt+ cell is mediated by a diffusible sex pheromone, named SCD-inducing pheromone (SCD-IP). This pheromone is released from mt– cells in the light, and the presence of mt+ cells stimulates its secretion from mt– cells. SCD-IP was purified by sequential column-chromatographic fractionation from culture medium in which both mating type cells had been co-cultured. Purified SCD-IP is a glycoprotein with an apparent molecular mass of 20 kDa. The molecular mass of the SCD-IP was estimated to be 18 kDa by mass spectrometry. Amino-terminal and two internal amino acid sequences of the pheromone revealed significant similarity to another Closterium pheromone, protoplast release-inducing protein (PR-IP) inducer of Closterium peracerosum-strigosum-littorale complex (C. pslc). These two pheromones induced different morphological reactions in each Closterium species. Based on these results, the diversity of sex pheromones is discussed.