Coordinate Regulation of Ribosomal Component Synthesis in Acanthamoeba castellanii: 5S RNA Transcription Is Down Regulated during Encystment by Alteration of TFIIIA Activity

Abstract

Transcription of large rRNA precursor and 5S RNA were examined during encystment of Acanthamoeba castellanii. Both transcription units are down regulated almost coordinately during this process, though 5S RNA transcription is not as completely shut down as rRNA transcription. The protein components necessary for transcription of 5S RNA and tRNA were determined, and fractions containing transcription factors comparable to TFIIIA, TFIIIB, and TFIIIC, as well as RNA polymerase III and a 3-end processing activity, were identified. Regulation of 5S RNA transcription could be recapitulated in vitro, and the activities of the required components were compared. In contrast to regulation of precursor rRNA, there is no apparent change during encystment in the activity of the polymerase dedicated to 5S RNA expression. Similarly, the transcriptional and promoter-binding activities of TFIIIC are not altered in parallel with 5S RNA regulation. TFIIIB transcriptional activity is unaltered in encysting cells. In contrast, both the transcriptional and DNA-binding activities of TFIIIA are strongly reduced in nuclear extracts from transcriptionally inactive cells. These results were analyzed in terms of mechanisms for coordinate regulation of rRNA and 5S RNA expression.The number of ribosomes in the cell is regulated to correspond to the quantitative requirement for protein synthesis. Therefore, the rate of biosynthesis of ribosomes varies with the cellular proliferation rate, in response to cellular growth without cell division (e.g., cardiac hypertrophy [1]), and with significant changes in protein synthesis rates in secretory tissues (e.g., the Drosophila paragonial gland [29] [reviewed in reference 22]). The synthesis of the 80 to 90 components of the ribosome must be coordinately up or down regulated to correspond to changes in ribosome elaboration rate. The mechanisms leading to balanced synthesis of these components is poorly understood. For the ribosomal proteins, regulation occurs at several steps in the biosynthetic pathway, including transcription by RNA polymerase II, processing, translation efficiency, and mRNA stability. The critical regulated step depends on the particular ribosomal protein and the species (2,3,(19)(20)(21)30). rRNA and 5S RNA are transcribed by RNA polymerases I and III, respectively. Thus, various ribosomal components are transcribed by each of the three eukaryotic RNA polymerases. Expression of the stable RNAs is regulated at the transcriptional level. Therefore, the problem of coordinating their expression is, at least in part, one of coordinating transcription of their genes by distinct transcriptional systems. This study investigates the mechanisms involved in coordinating precursor rRNA and 5S RNA transcription in Acanthamoeba castellanii.When starved for essential nutrients, A. castellanii leaves the growth/division cycle and undergoes cellular differentiation into a dormant cyst. When this occurs, rRNA transcription, which in the proliferating cell is 75% of pulse-labeled RNA, ceases comple...

Silkworm TFIIIB Binds both Constitutive and Silk Gland-Specific tRNAAla Promoters but Protects Only the Constitutive Promoter from DNase I Cleavage

“…TFIIIB/D and TFIIIB/C/D complexes could be similar in size and shape but differ by a small number of polypeptides. Detection of TFIIIB binding to polymerase III templates by gel retardation or DNase I footprinting has been reported previously only for S. cerevisiae (21,23,24), despite the chromatographic resolution of TFIIIB activity in a variety of other systems: mammalian cells (43,57,66), Xenopus laevis (16,46), Drosophila melanogaster (3), and Acanthamoeba castellanii (30). Two kinds of experiments provide indirect evidence for TFIIIB binding in nonyeast systems, however.…”

Rrn3 Phosphorylation Is a Regulatory Checkpoint for Ribosome Biogenesis

“…Studies on the inactivation of rDNA transcription during encystment of Acanthamoeba castellanii demonstrated that inactivation was the result of a modification of RNA polymerase I (52,53). Similarly, studies on the regulation of rDNA transcription in mammalian cells undergoing serum starvation or P1798 cells exposed to DEX demonstrated that a polymeraseassociated factor was responsible for the inhibition of rDNA transcription.…”

Regulation of the RNA Polymerase I and III Transcription Systems in Response to Growth Conditions

“…In contrast, viral infection and serum factors have been shown to alter the activity of the TFIIIC fraction (33,34). Recently the differential expression of the 5 S rRNA and tRNA genes during encystment in Acanthamoeba castellanii has been attributed to the disappearance of the 5 S rRNA-specific transcription factor TFIIIA (35).…”