Small-subunit rRNA processome proteins are translationally regulated during differentiation of Trypanosoma cruzi.
We used differential display to select genes differentially expressed during differentiation of epimastigotes into metacyclic trypomastigotes in the protozoan parasite Trypanosoma cruzi. One of the selected clones had a sequence similar to that of the small-subunit (SSU) processome protein Sof1p, which is involved in rRNA processing. The corresponding T. cruzi protein, TcSof1, displayed a nuclear localization and is downregulated during metacyclogenesis. Heterologous RNA interference assays showed that depletion of this protein impaired growth but did not affect progression through the cell cycle, suggesting that ribosome synthesis regulation and the cell cycle are uncoupled in this parasite. Quantitative PCR (qPCR) assays of several SSU processome-specific genes in T. cruzi also showed that most of them were regulated posttranscriptionally. This process involves the accumulation of mRNA in the polysome fraction of metacyclic trypomastigotes, where TcSof1 cannot be detected. Metacyclic trypomastigote polysomes were purified and separated by sucrose gradient sedimentation. Northern blot analysis of the sucrose gradient fractions showed the association of TcSof1 mRNA with polysomes, confirming the qPCR data. The results suggest that the mechanism of regulation involves the blocking of translation elongation and/or termination.
Nardelli
Sheila Cristina
SC
Instituto de Biologia Molecular do Paraná, Rua Algacyr Munhoz Mader 3775, Curitiba 81350-010, Paraná, Brazil.
Avila
Andréa Rodrigues
AR
Freund
Aline
A
Motta
Maria Cristina
MC
Manhães
Lauro
L
de Jesus
Teresa Cristina Leandro
TC
Schenkman
Sergio
S
Fragoso
Stenio Perdigão
SP
Krieger
Marco Aurélio
MA
Goldenberg
Samuel
S
Dallagiovanna
Bruno
B
D000818:Animals; D015152:Blotting, Northern; D002454:Cell Differentiation; D005455:Fluorescent Antibody Technique; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D047468:Immunoprecipitation; D009154:Mutation; D014176:Protein Biosynthesis; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism*; D012322:RNA Precursors / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D016053:RNA, Protozoan / Q000235:genetics / Q000378:metabolism; D012335:RNA, Ribosomal / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017411:Ribonucleoproteins, Small Nuclear; D029701:Saccharomyces cerevisiae Proteins; D014349:Trypanosoma cruzi / Q000235:genetics / Q000254:growth & development*
D015800:Protozoan Proteins; D012322:RNA Precursors; D012333:RNA, Messenger; D016053:RNA, Protozoan; D012335:RNA, Ribosomal; D017411:Ribonucleoproteins, Small Nuclear; C081686:SOF1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins