Independent mechanisms are utilized for the coordinate and transient accumulation of two differentiation-specific mRNAs during differentiation of Naegleria gruberi amoebae into flagellates.
During the differentiation of Naegleria gruberi amoebae into flagellates, four differentiation-specific (DS) mRNAs are transiently and coordinately accumulated. Three of the four DS mRNAs, Class II, III, and IV, encode alpha-tubulin, beta-tubulin, and flagellar calmodulin, respectively. The protein product of the Class I mRNA has not been identified. We examined the effects of inhibition of protein synthesis on transcription and accumulation of beta-tubulin mRNA and Class I mRNA to understand the mechanism of coordinate regulation. Inhibition of protein synthesis at the beginning of differentiation completely blocked transcription of the beta-tubulin gene. Addition of cycloheximide at 30 or 40 min after initiation of differentiation inactivated transcription of the beta-tubulin gene in less than 10 min as judged by nuclear run-on experiments. However, once differentiation had proceeded for more than 50 min, inhibition of protein synthesis did not inactivate transcription of beta-tubulin mRNA was more active in cycloheximide-treated cells than in control cells. Cycloheximide treatment at the initiation of the differentiation also blocked transcription of the Class I gene. However, addition of the drug after 30 min had no significant effect on the transcription of the Class I gene. Cycloheximide treatment also increased the half-lives of beta-tubulin and Class I mRNA drastically. These data suggest that: (1) the transient accumulation of the two DS mRNAs during differentiation are regulated by changing both the rate of transcription and the stability of the mRNAs; (2) protein synthesis is required for the transcriptional and post-transcriptional regulations; (3) the transcriptional regulation mechanisms of the beta-tubulin gene and that of the Class I gene are distinct; and (4) the transcription of the beta-tubulin gene is regulated by different mechanisms during differentiation.
Bok
J
J
Department of Biology, Yonsei University, Seoul, Korea.
Jin
Y
Y
Lee
J
J
D000818:Animals; D002147:Calmodulin / Q000096:biosynthesis*; D002454:Cell Differentiation; D003513:Cycloheximide / Q000494:pharmacology*; D005407:Flagella / Q000502:physiology; D005786:Gene Expression Regulation / Q000187:drug effects; D007700:Kinetics; D009253:Naegleria / Q000166:cytology / Q000254:growth & development* / Q000378:metabolism*; D014176:Protein Biosynthesis / Q000187:drug effects; D012333:RNA, Messenger / Q000302:isolation & purification / Q000378:metabolism*; D016053:RNA, Protozoan / Q000302:isolation & purification / Q000378:metabolism; D014404:Tubulin / Q000096:biosynthesis*
D002147:Calmodulin; D012333:RNA, Messenger; D016053:RNA, Protozoan; D014404:Tubulin; D003513:Cycloheximide