Insights into the regulation of GPEET procyclin during differentiation from early to late procyclic forms of Trypanosoma brucei.
The procyclic form of Trypanosoma brucei colonises the gut of its insect vector, the tsetse fly. GPEET and EP procyclins constitute the parasite's surface coat at this stage of the life cycle, and the presence or absence of GPEET distinguishes between early and late procyclic forms, respectively. Differentiation from early to late procyclic forms in vivo occurs in the fly midgut and can be mimicked in culture. Our analysis of this transition in vitro delivered new insights into the process of GPEET repression. First, we could show that parasites followed a concrete sequence of events upon triggering differentiation: after undergoing an initial growth arrest, cells lost GPEET protein, and finally late procyclic forms resumed proliferation. Second, we determined the stability of both GPEET and EP mRNA during differentiation. GPEET mRNA is exceptionally stable in early procyclic forms, with a half-life >6h. The GPEET mRNA detected in late procyclic form cultures is a mixture of transcripts from both bona fide late procyclic forms and GPEET-positive 'laggard' parasites present in these cultures. However, its stability was clearly reduced during differentiation and in late procyclic form cultures. Alternatively processed GPEET transcripts were enriched in samples from late procyclic forms, suggesting that altered mRNA processing might contribute to repression of GPEET in this developmental stage. In addition, we detected GPEET transcripts with non-templated oligo(U) tails that were enriched in late procyclic forms. To the best of our knowledge, this is the first study reporting a uridylyl-tailed, nuclear-encoded mRNA species in trypanosomatids or any other protozoa.
Institute of Cell Biology, University of Bern, Baltzerstrasse 4, CH-3012 Bern, Switzerland; Graduate School of Cellular and Biomedical Sciences, University of Bern, Freiestrasse 1, CH-3012 Bern, Switzerland.
Sebastian
S
Knüsel
Isabel
I
Roditi
D000818:Animals; D018507:Gene Expression Regulation, Developmental; D006801:Humans; D008018:Life Cycle Stages; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D015800:Protozoan Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014346:Trypanosoma brucei brucei / Q000737:chemistry / Q000235:genetics / Q000254:growth & development* / Q000378:metabolism; D014353:Trypanosomiasis, African / Q000469:parasitology*
C406751:GPEET protein, Trypanosoma brucei; D008562:Membrane Glycoproteins; D015800:Protozoan Proteins; D012333:RNA, Messenger
3′ UTR; 5-ethynyl-2′-deoxyuridine; ActD; Differentiation; EdU; GRE; PF; Procyclin; RNA stability; SBPW; Trypanosoma brucei; U; UTR; Uridylation; VSG; actinomycin D; glycerol responsive element; half-life; procyclic form; saponin-based permeabilisation and wash reagent; t(1/2); untranslated region; uridine; variant surface glycoprotein