Increased transcript levels of a methionine synthase during adhesion-induced activation of Chlamydomonas reinhardtii gametes

Venkatesh Kurvari, Fang Qian, William J. Snell

Research output: Contribution to journalArticle

23 Scopus citations

Abstract

Chlamydomonas gametes of opposite mating types interact through flagellar adhesion molecules called agglutinins leading to a signal transduction cascade that induces cell wall loss and activation of mating structures along with other cellular responses that ultimately result in zygote formation. To identify molecules involved in these complex cellular events, we have employed subtractive and differential hybridization with cDNA from mt+ gametes activated for fertilization and non-signaling, vegetative (non-gametic) cells. We identified 55 cDNA clones whose transcripts were regulated in activated gametes. Here we report the molecular cloning and characterization of the complementary DNA (cDNA) for one clone whose transcripts in activated gametes were several-fold higher than in normal gametes. Regulation of the transcript was not related simply to protein synthesis because it was not increased in cells synthesizing new cell wall proteins. The cDNA contained a single open reading frame (ORF) of 815 amino acids encoding a polypeptide of calculated relative mass of 87 kDa. Database search analysis and sequence alignment indicated that the deduced amino acid sequence exhibited 42% identity and 62% similarity to a class of prokaryotic methyl transferases (5-methyltetrahydrofolate-homocysteine methyl transferase; EC 2.1.1.14) known to be involved in the terminal step of de novo biosynthesis of methionine. This enzyme catalyzes transfer of a methyl group from 5-methyltetrahydrofolate to homocysteine resulting in methionine formation. Affinity-purified polyclonal antibodies raised against a bacterially produced GST-fusion protein identified a 85 kDa soluble protein in Chlamydomonas gametes. Southern blot hybridization indicated that the enzyme is encoded by a single-copy gene. The evidence presented in this paper raises the possibility that, in addition to its participation in de novo biosynthesis and regeneration of methionine, Chlamydomonas methionine synthase may play a role in adhesion-induced events during fertilization.

Original languageEnglish (US)
Pages (from-to)1235-1252
Number of pages18
JournalPlant Molecular Biology
Volume29
Issue number6
DOIs
StatePublished - Dec 1 1995

Keywords

  • Chlamydomonas
  • methionine synthase
  • signal transduction
  • subtractive and differential hybridization
  • transcriptional up-regulation

ASJC Scopus subject areas

  • Agronomy and Crop Science
  • Genetics
  • Plant Science

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