Control of VSG gene expression sites

doi:10.1016/S0166-6851(01)00243-2

Molecular and Biochemical Parasitology

Volume 114, Issue 1, 25 April 2001, Pages 17-27

https://doi.org/10.1016/S0166-6851(01)00243-2 Get rights and content

Abstract

Trypanosoma brucei survives in mammals by antigenic variation of its surface coat consisting of variant surface glycoprotein (VSG). Trypanosomes change coat mainly by replacing the transcribed VSG genes in an active telomeric expression site by a different VSG gene. There are about 20 different expression sites and trypanosomes can also change coat by switching the site that is active. This review summarizes recent work on the mechanism of site switching and on the way inactive expression sites are kept silent.

Introduction

Trypanosoma brucei is covered by a dense surface coat consisting of a single glycoprotein, the Variant Surface Glycoprotein (VSG) [1]. VSGs are translated from mRNAs transcribed from VSG genes located in telomeric expression sites. There are two types of expression sites (ESs), schematically depicted in Fig. 1, metacyclic ESs (M-ESs) and bloodstream form ESs (B-ESs). The M-ESs are used in metacyclic trypanosomes in the salivary gland of the tsetse fly and in the first few days after the tsetse fly introduces the trypanosomes with its saliva in a mammalian host. Then the B-ESs take over and they are used throughout a chronic infection.

Trypanosomes can change the VSG gene transcribed in a B-ES in two ways.

By replacing the VSG gene in an active expression site by a different one. This is quantitatively by far the most important source of VSG antigenic variation and it requires a DNA rearrangement.
By switching expression sites. This involves switching off the active site and activating an earlier silent site. As there are only about 20 B-ESs per trypanosome nucleus, the switching of ESs makes a very modest contribution to antigenic variation of the VSG coat. It is, therefore, likely that multiple B-ESs exist to allow expression of different sets of the ESAGs present in each ES [2], [3]. (This point will be discussed by Pays et al. in an upcoming review.)

How trypanosomes switch between expression sites and how they keep silent sites silent (allelic exclusion) is the subject of this review.

Section snippets

Control of metacyclic expression sites

In preparation for their entry into the mammalian host, trypanosomes that have reached the salivary gland of the tsetse fly acquire a VSG coat. The VSG mRNAs used for this coat are transcribed from metacyclic expression sites, M-ESs, shown in Fig. 1. M-ESs probably arose from B-ESs in evolution [4]. They lack functional ESAGs and the promoter is directly upstream of the VSG gene. The VSG genes in these sites are rarely replaced and antigenic variation in the metacyclic phase of the life cycle

Control of bloodstream expression sites: old facts and misinterpretations

Two types of models have been considered to explain control of bloodstream expression sites, cross-talk and non-cross talk models. In the cross-talk models, the activation of a new ES-A and the inactivation of the old one B are coupled events. In the non-cross-talk model, the two events are unrelated and switching from A to B is helped by selection. If site A is switched off before B is switched on, the trypanosome becomes coatless and dies; if site B is switched on without switching off A, the

Control of bloodstream expression sites; recent developments

Two recent developments have changed thinking on expression site control. First, it has become clear that there is cross-talk between ESs, and that activation of a new ES is coupled to inactivation of the earlier active site. Secondly, it has been shown that silent sites are not completely silent and that control of expression sites is at least in part at the level of transcription attenuation. The data are complex and not entirely unambiguous, and we shall present them here in some detail.

The B-ES promoter

The minimal B-ES promoter still maximally active in transient assays is remarkably small, only about 70 bp [42], [43]. The promoter sequences required for regulated expression in vivo have been analyzed by promoter remodeling in intact trypanosomes [30], [44], [45]. Rudenko et al. [30] showed that the ES core promoter itself is dispensable and that a B-ES in which this promoter is replaced by a ribosomal DNA promoter can be maximally activated and also silenced and reactivated at a normal rate.

Chromatin structure of active and inactive B-ESs

The chromatin structure of active and inactive B-ESs has been analyzed by exposure to nucleases in isolated nuclei or permeabilized trypanosomes. The VSG gene in an active B-ES is slightly more sensitive to DNase I than the same gene in an inactive ES (Pays et al., 1981)

Much larger differences in sensitivity were observed when the VSG genes were exposed to single-strand DNA specific endonucleases [46]. The presence of single-stranded DNA regions within VSG gene chromatin was interpreted as an

Silencing of expression sites

Early studies on silencing of B-ESs suggested that shut-off is complete. No mRNA corresponding to VSG genes of silent ESs was detected and this has been confirmed by recent experiments using more sensitive techniques, such as RT-PCR [18], [47]. These early studies appeared to be substantiated by the analysis of silenced ESs tagged with drug resistance genes or a luciferase gene inserted just downstream of the ES promoter. Silencing of the ES reduced the transcript level of these genes more than

How does it work?

We do not know yet how the control of trypanosome ESs works, but it may be useful to briefly summarize the main elements that should find a place in any future model. The requirements for maximal transcription of a B-ES and for the ability to switch are minimal, only a 70 bp core promoter will do as long as this is in the context of an ES and sufficiently distant from the upstream 50-bp repeats. Even the 70-bp core sequence is not essential, as it can be replaced by a ribosomal DNA promoter

Acknowledgements

We thank Dr G. Rudenko (Wellcome Trust Centre for the Epidemiology of Infectious Disease, Department of Zoology, University of Oxford, UK) and Dr R. Mussmann for helpful comments on the manuscript. The experimental work on trypanosomes in our lab is supported in part by grants from The Netherlands Foundation for Chemical Research (SON), with financial support of The Netherlands Organization for Scientific Research (NWO). S. Ulbert. is supported by a grant from The Boehringer Ingelheim Fonds.

References (56)

J.D. Barry et al.
VSG gene control and infectivity strategy of metacyclic stage Trypanosoma brucei
Mol. Biochem. Parasitol.
(1998)
Y.L. Nagoshi et al.
The putative promoter for a metacyclic VSG gene in African trypanosomes
Mol. Biochem. Parasitol.
(1995)
M. McAndrew et al.
Testing promoter activity in the trypanosome genome: isolation of a metacyclic-type VSG promoter, and unexpected insights into RNA polymerase II transcription
Exp. Parasitol.
(1998)
K.S. Kim et al.
Co-duplication of a variant surface glycoprotein gene and its promoter to an expression site in African trypanosome
J. Biol. Chem.
(1997)
S.V. Graham et al.
A structural and transcription pattern for variant surface glycoprotein gene expression sites used in metacyclic stage Trypanosoma brucei
Mol. Biochem. Parasitol.
(1999)
C.M. Alarcon et al.
Leaky transcription of variant surface glycoprotein gene expression sites in bloodstream African trypanosomes
J. Biol. Chem.
(1999)
A.W. Cornelissen et al.
Two simultaneously active VSG gene transcription units in a single Trypanosoma brucei variant
Cell
(1985)
D. Horn et al.
Analysis of Trypanosoma brucei vsg expression site switching in vitro
Mol. Biochem. Parasitol.
(1997)
G.A.M. Cross et al.
Regulation of vsg expression site transciption and switching in Trypanosoma brucei
Mol. Biochem. Parasitol.
(1998)
P.J. Myler et al.
Changes in telomere length associated with antigenic variation in Trypanosoma brucei
Mol. Biochem. Parasitol.
(1988)

G.A.M. Cross

Identification, purification and properties of clone-specific glycoprotein antigens constituting the surface coat of Trypanosoma brucei

Parasitology

(1975)

P. Borst et al.

The expression sites for variant surface glycoproteins of Trypanosoma brucei

W. Bitter et al.

How Trypanosoma brucei may cope with the diversity of transferrins in its mammalian hosts

Nature

(1998)

Cited by (103)

Infection and Immunity
2022, Clinical Immunology
Human infections are caused by bacteria, fungi, parasites, and viruses. While the human body defends against infections consistently via innate and adaptive immune responses, the face of infection in human disease is changing globally. For instance, the immunocompromised population is on the rise, mainly due to extensive chemotherapies, immunosuppressive treatment for transplantation, and emerging infections such as infection with Human immunodeficiency virus (HIV). In this light, the development of effective immunological diagnostic tests and immune-mediating treatments may be vital. The present chapter provides an updated review on definition and epidemiology, molecular mechanisms, diagnosis, and treatment of infectious diseases, as well as highlights areas of further research in the field.
Extracellular vesicles in parasitic disease
2019, Exosomes: A Clinical Compendium
Pathogens shed extracellular vesicles (EVs) in the host microenvironment. EVs released by the pathogens and those produced by the host have been characterized and isolated from diverse organisms and these EVs have been shown to play an important role in intracellular communication between pathogens and hosts Virus, bacteria, fungi, protozoa and helminth release EVs continuously into extracellular environment. These EVs measure between 20 and 200 nm in diameter and contain proteins, glycoconjugates, lipids, nucleic acid (RNAs, non-transcribed RNAs and microRNAs). They can be detected in various body fluids (serum, plasma, saliva, urine, breast milk, etc.). EVs modulate the communication and interaction with the host, also delivering virulence factors and antigens. Indeed, EVs isolated from parasites act either as messengers priming host cells for specific interactions, promoting invasion by the pathogen, or activating host innate and acquired immunity. Many studies have shown that EVs released from parasites are able to affect host cells. For example, this interaction changes the composition of exosomes released by macrophages. Pathogens can also stimulate the release of host EVs containing proteins or other components of the parasite as a mechanism for transporting contents of these microorganisms. Moreover, some pathogens have as mechanism of adaptation to the host, the release of EVs, which facilitates their dissemination or escape of the immune system. In this chapter, we intend to present updated information regarding the role of EVs released by some major Protozoan parasites.
Universal primers suitable to assess population dynamics reveal apparent mutually exclusive transcription of the Babesia bovis ves1α gene
2009, Molecular and Biochemical Parasitology
Babesia bovis is an intraerythrocytic hemoparasite of widespread distribution, which adversely affects livestock production in many regions of the world. This parasite establishes persistent infections of long duration, at least in part through rapid antigenic variation of the VESA1 protein on the infected-erythrocyte surface. To understand the dynamics of in vivo antigenic variation among the parasite population it is necessary to have sensitive and broadly applicable tools enabling monitoring of variation events in parasite antigen genes. To address this need for B. bovis, “universal” primers for the polymerase chain reaction have been designed for the ves1α gene, spanning from exon 2 to near the 3′ end of cysteine–lysine-rich domain (CKRD) sequences in exon 3. These primers robustly amplified this segment, with minimal bias, from essentially the entire repertoire of full-length ves1α sequences in the B. bovis Mexico isolate genome, and are equivalently present in other isolates. On purified genomic DNA, this primer set can achieve a sensitivity of 10 genome equivalents or less. When applied to the amplification of cDNA derived from the B. bovis C9.1 clonal line evidence consistent with mutually exclusive transcription of the ves1α gene was obtained, concomitant with detection of numerous mutational events among members of the parasite population. These characteristics of the primers will facilitate the application of polymerase chain reaction-based methodologies to the study of B. bovis population and antigenic switching dynamics.
African Trypanosomiasis
2009, Vaccines for Biodefense and Emerging and Neglected Diseases
Bioinformatic insights to the ESAG5 and GRESAG5 gene families in kinetoplastid parasites
2008, Molecular and Biochemical Parasitology
Trypanosoma brucei, the causative agent of African sleeping sickness, evades the immune response by expressing a coat of variant surface glycoprotein (VSG). VSG is expressed from a single telomeric expression site (ES), along with a number of expression site associated genes (ESAGs). Thus far, the function of most ESAGs is unknown. One ES contains the serum resistance associated gene (SRA), which confers resistance to trypanosome lytic factor in T. b. rhodesiense. Only three other ESAGs – 5, 6 and 7 – are present in this ES. ESAGs 6 and 7 encode a heterodimeric transferrin receptor, but the function of ESAG5 has not been identified. We present here a bioinformatic analysis of ESAG5 and distinguish between T. brucei-specific ESAGs and Genes Related to ESAG5 (GRESAGs), which occur outside of ESs in chromosomal-internal contexts. Further, a genome-wide survey of these genes across kinetoplastids identifies a family of GRESAG5s in a number of species. Analysis of phylogenetic relationships indicates that this family may have evolved from a single ancestral copy. Predicted properties of (GR)ESAG5 proteins indicate a glycosylated protein containing either a signal peptide or transmembrane domain. Further analysis indicates a possible relationship to the lipid transfer/lipopolysaccharide-binding family which includes the bactericidal/permeability increasing (BPI) protein. Together, these results provide insights into the structure and evolution of an important extended gene family, and present a number of testable hypotheses which will aid in elucidating the function of ESAG5.
SnSAG5 is an alternative surface antigen of Sarcocystis neurona strains that is mutually exclusive to SnSAG1
2008, Veterinary Parasitology
Sarcocystis neurona is an obligate intracellular parasite that causes equine protozoal myeloencephalitis (EPM). Previous work has identified a gene family of paralogous surface antigens in S. neurona called SnSAGs. These surface proteins are immunogenic in their host animals, and are therefore candidate molecules for development of diagnostics and vaccines. However, SnSAG diversity exists in strains of S. neurona, including the absence of the major surface antigen gene SnSAG1. Instead, sequence for an alternative SnSAG has been revealed in two of the SnSAG1-deficient strains. Herein, we present data characterizing this new surface protein, which we have designated SnSAG5. The results indicated that the protein encoded by the SnSAG5 sequence is indeed a surface-associated molecule that has characteristics consistent with the other SAGs identified in S. neurona and related parasites. Importantly, Western blot analyses of a collection of S. neurona strains demonstrated that 6 of 13 parasite isolates express SnSAG5 as a dominant surface protein instead of SnSAG1. Conversely, SnSAG5 was not detected in SnSAG1-positive strains. One strain, which was isolated from the brain of a sea otter, did not express either SnSAG1 or SnSAG5. Genetic analysis with SnSAG5-specific primers confirmed the presence of the SnSAG5 gene in Western blot-positive strains, while also suggesting the presence of a novel SnSAG sequence in the SnSAG1-deficient, SnSAG5-deficient otter isolate. The findings provide further indication of S. neurona strain diversity, which has implications for diagnostic testing and development of vaccines against EPM as well as the population biology of Sarcocystis cycling in the opossum definitive host.

View all citing articles on Scopus

View full text

ReviewControl of VSG gene expression sites

Abstract

Introduction

Section snippets

Control of metacyclic expression sites

Control of bloodstream expression sites: old facts and misinterpretations

Control of bloodstream expression sites; recent developments

The B-ES promoter

Chromatin structure of active and inactive B-ESs

Silencing of expression sites

How does it work?

Acknowledgements

Mol. Biochem. Parasitol.

Mol. Biochem. Parasitol.

Exp. Parasitol.

J. Biol. Chem.

Mol. Biochem. Parasitol.

J. Biol. Chem.

Cell

Mol. Biochem. Parasitol.

Mol. Biochem. Parasitol.

Mol. Biochem. Parasitol.

Cell

Cell

Cell

Cell

Mol. Biochem. Parasitol.

Mol. Biochem. Parasitol.

Mol. Biochem. Parasitol.

Mol. Biochem. Parasitol.

Mol. Biochem. Parasitol.

Mol. Biochem. Parasitol.

J. Mol. Biol.

Mol. Biochem. Parasitol.

J. Biol. Chem.

Trends Genet.

Trends Genet.

Identification, purification and properties of clone-specific glycoprotein antigens constituting the surface coat of Trypanosoma brucei

Parasitology

The expression sites for variant surface glycoproteins of Trypanosoma brucei

How Trypanosoma brucei may cope with the diversity of transferrins in its mammalian hosts

Nature

Review
Control of VSG gene expression sites