Date Published: April 10, 2017
Publisher: Public Library of Science
Author(s): Henriette Zimmermann, Ines Subota, Christopher Batram, Susanne Kramer, Christian J. Janzen, Nicola G. Jones, Markus Engstler, Samuel James Black.
For persistent infections of the mammalian host, African trypanosomes limit their population size by quorum sensing of the parasite-excreted stumpy induction factor (SIF), which induces development to the tsetse-infective stumpy stage. We found that besides this cell density-dependent mechanism, there exists a second path to the stumpy stage that is linked to antigenic variation, the main instrument of parasite virulence. The expression of a second variant surface glycoprotein (VSG) leads to transcriptional attenuation of the VSG expression site (ES) and immediate development to tsetse fly infective stumpy parasites. This path is independent of SIF and solely controlled by the transcriptional status of the ES. In pleomorphic trypanosomes varying degrees of ES-attenuation result in phenotypic plasticity. While full ES-attenuation causes irreversible stumpy development, milder attenuation may open a time window for rescuing an unsuccessful antigenic switch, a scenario that so far has not been considered as important for parasite survival.
Pathogenic bacteria and protozoan parasites often employ a coat of surface molecules to protect themselves from host immune attack. These surface coats are sometimes variable and hence, not only act as a physical shield but have evolved as an efficient camouflage strategy. The surface-exposed proteins are mostly members of large families and are subject to antigenic variation, i.e. they are sporadically exchanged. This allows the persistence of the pathogens in the host, as well as reinfection. The genetic mechanisms underlying antigenic variation differ greatly, ranging from transcriptional changes in Plasmodium to duplicative events for example in Borrelia or Neisseria . An extensively studied model for antigenic variation is the protozoan parasite Trypanosoma brucei and the phenomenon was, in fact, first described in trypanosomes [2,3]. The surface coat of trypanosomes consists of millions of identical copies of a variant surface glycoprotein (VSG) [4,5]. The highly immunogenic VSGs cause a rapid host immune response, which is thought to lead to an almost complete elimination of the parasite population. Only parasites that have switched to the expression of an immunologically distinct VSG survive. Thus, at any given time just one VSG out of a repertoire of several hundreds of VSG genes is expressed and dominates the cell surface of the pathogen [6,7]. At all times the parasite has to maintain the shielding function of the coat and hence, the concentration of VSGs on the cell surface. This is not a straightforward task as the VSG coat is continuously endocytosed and recycled with unprecedented kinetics . Consequently, VSGs are constantly produced in large quantities. Uniquely, this high level expression of VSG is driven by RNA-polymerase I .
Little is known about the control of in situ VSG switching. Basically, there are two possibilities: either the old ES is shut-down and then a new ES is activated, or a new ES is transcriptionally activated before the old one is switched off. Support for the first possibility comes from tagging two ESs with selectable markers [53,54]. In the presence of the drugs rapid switching between the tagged ESs occurred. This suggested that one silent ES lingers in a pre-active state and, thus, is immediately activated once the active ES is silenced. However, another study reported that the inducible block of ES transcription caused growth inhibition and subsequent probing of several silent ESs . This suggested that the silencing of the active ES does not cause an immediate antigenic switch. In addition, depletion of VSG mRNA results in a rapid precytokinesis arrest, which suggests that an inactivation of the ES without the simultaneous activation of a new one would be lethal . Therefore, in a previous study, we tested the possibility that a new VSG is activated, before the old one is silenced. This was achieved by inducible overexpression of a second VSG . Surprisingly, the trypanosomes responded with attenuation of the active ES and growth retardation. It is important to note that these cells never stopped growth, i.e. they never arrested in the cell cycle, but rather lingered in a prolonged G1-phase. Interestingly, growth retardation was accompanied by signs of developmental competence. This raised the question whether ectopic VSG overexpression or ES-attenuation could lead to stumpy development. This possibility, however, remained unexplored, as the monomorphic trypanosome strains routinely used in the laboratory are developmentally deficient. Only more natural, pleomorphic parasites are suitable for analyses of trypanosome differentiation [35,36], but large scale cultivation and genetic manipulation are very difficult.