phosphate-6Mana(1-2)Mana(1-6)Mana(1-4)Glc
Na(1-6)-phosphatidylinositol. After its incorporation, this structure is modified by the attachment of several galactose residues to the mannose
next to the glucosamine [113]. The most closely
investigated of the VSG variants is MITat 1.4 of
T. brucei: the complete amino acid sequence,
details of the secondary structure, the glycolipid
anchor and the structure of the Asn-bound carbohydrate are all known [201].
T. equiperdum possesses at least 100 and T. brucei more than 1000 closely linked VSG genes,
which together make up about 10 % of the DNA
content of 8.6 . 10 7 bp. As the chromosomes of
the trypanosomes never appear in a condensed
form in the cell cycle, the ploidy level cannot be
directly determined; however, genetic observations suggest that T. brucei is diploid. Nevertheless, there is only one copy of each VSG gene in
the genome; the evolution of the VSG genes is
apparently so rapid that homologous positions on
the chromosomes are always occupied by different genes. The mechanisms for maintaining the
VSG repertoire are mainly those of gene duplication and conversion [139].
The VSG genes are expressed only when they
are at one of the expression sites at the ends of
the chromosomes (telomeres). The VSG genes
lying internally on the chromosome can be activated by the transposition of the silent basic copy
(BC) to one of the telomeric expression sites
(duplicative transposition) to give an "expressionlinked copy" (ELC). The transposed DNA segment begins 1-2 kb in front of the gene and ends
just behind it. Some ELCs are hybrids of different
BCs [155]. T. brucei has about 200 chromosomes
whose telomeres carry VSG genes; only one of
the expression sites is active at anyone time, but
the selection mechanism is unknown. Silent VSG
genes located close to expression sites can be activated in various ways; the inactive gene can be
transposed duplicatively to an expression site or
can exchange places with an active gene. ELCs
that become inactive are not retransposed to
internal chromosomes sites but are eliminated
[102, 139]. The activation ofVSGs after infection
follows a certain course whose regulation is not
known but whose biological advantage is obvious. The repeated activation of anyone gene
would undermine the successful competition with
the host immune system. In the tsetse fly, one particular set of 10-20 VSG genes is activated. The
VSGs produced during the early states of infection in vertebrates arise from genes which are
already located at the telomeres [102].
6.9.4 Surface Proteins of Other Protozoa
241
The transcription unit of a VSG gene has a
length of 60 kb and is polycistronic. In front of
the VSG genes at the expression site (ES) are
more than seven ES-associated genes (ESAGs)
together with a more than 10-kb-Iong segment of
70-bp repeats. The importance of the ESAGs is
unclear; one was found to code for a membranebound protein of unknown function [2, 139].
Quite surprisingly, the transcription of the VSG
genes, and of the procyclic acidic repetitive protein (PARP) genes, described below, from insect
trypanosomes, is not sensitive to a-amanitin. All
other protein-coding trypanosome genes are transcribed by a normal, amanitin-sensitive polymerase II. In this case, there is either an amanitininsensitive form in addition to the normal polymerase II, or the normal polymerase II can be
made insensitive by means of some additional
factor [59, 157]. As for all mRNAs of the Kinetoplastida, the VSG mRNAs contain a 5'-terminal
mini-exon of 35 nt that is introduced by transsplicing (p. 39). The primary translation product
(pre-VSG) has not only a relatively long, 25- to
33-bp N-terminal signal sequence but also a Cterminal extension of 17-23 non-polar amino
acids. Immediately after translation, but whilst
still in the endoplasmic reticulum, this is
exchanged for the phospho glycolipid which
anchors the VSG to the cell membrane. A phospholipase C can release the VSG, in which case
sn-l,2-dimyristylglycerol remains behind in the
membrane. This enzyme, which may playa role in
the exchange ofVSG variants, has been isolated;
it has a molecular weight of 39 kDa and is specific
for VSGs [20]. The procyclic stages of T. brucei
from the intermediate host Glossina sp. or from
cultures at 27°C have no VSGs on their surface
but instead have PARPs. This is a family of invariable proteins composed mainly of Glu-Pro
repeats with a single N-glycosylation site [33].
6.9.4 Surface Proteins of Other Protozoa
The South American T. cruzi, the agent causing
Chagas' disease, has no VSG system. In this case,
both the epimastigotes from the insect host and
the trypomastigotes from mammalian blood have
different glycoproteins on their surfaces, but
these do not show the variability typical of VSGs
[192]. The penetration into cells by parasitic
forms in vertebrate blood is inhibited by antibodies against the surface protein. The gene for this
protein has been sequenced: the coding sequence
contains the nonapeptide DKKESGDSE in five
Na(1-6)-phosphatidylinositol. After its incorporation, this structure is modified by the attachment of several galactose residues to the mannose
next to the glucosamine [113]. The most closely
investigated of the VSG variants is MITat 1.4 of
T. brucei: the complete amino acid sequence,
details of the secondary structure, the glycolipid
anchor and the structure of the Asn-bound carbohydrate are all known [201].
T. equiperdum possesses at least 100 and T. brucei more than 1000 closely linked VSG genes,
which together make up about 10 % of the DNA
content of 8.6 . 10 7 bp. As the chromosomes of
the trypanosomes never appear in a condensed
form in the cell cycle, the ploidy level cannot be
directly determined; however, genetic observations suggest that T. brucei is diploid. Nevertheless, there is only one copy of each VSG gene in
the genome; the evolution of the VSG genes is
apparently so rapid that homologous positions on
the chromosomes are always occupied by different genes. The mechanisms for maintaining the
VSG repertoire are mainly those of gene duplication and conversion [139].
The VSG genes are expressed only when they
are at one of the expression sites at the ends of
the chromosomes (telomeres). The VSG genes
lying internally on the chromosome can be activated by the transposition of the silent basic copy
(BC) to one of the telomeric expression sites
(duplicative transposition) to give an "expressionlinked copy" (ELC). The transposed DNA segment begins 1-2 kb in front of the gene and ends
just behind it. Some ELCs are hybrids of different
BCs [155]. T. brucei has about 200 chromosomes
whose telomeres carry VSG genes; only one of
the expression sites is active at anyone time, but
the selection mechanism is unknown. Silent VSG
genes located close to expression sites can be activated in various ways; the inactive gene can be
transposed duplicatively to an expression site or
can exchange places with an active gene. ELCs
that become inactive are not retransposed to
internal chromosomes sites but are eliminated
[102, 139]. The activation ofVSGs after infection
follows a certain course whose regulation is not
known but whose biological advantage is obvious. The repeated activation of anyone gene
would undermine the successful competition with
the host immune system. In the tsetse fly, one particular set of 10-20 VSG genes is activated. The
VSGs produced during the early states of infection in vertebrates arise from genes which are
already located at the telomeres [102].
6.9.4 Surface Proteins of Other Protozoa
241
The transcription unit of a VSG gene has a
length of 60 kb and is polycistronic. In front of
the VSG genes at the expression site (ES) are
more than seven ES-associated genes (ESAGs)
together with a more than 10-kb-Iong segment of
70-bp repeats. The importance of the ESAGs is
unclear; one was found to code for a membranebound protein of unknown function [2, 139].
Quite surprisingly, the transcription of the VSG
genes, and of the procyclic acidic repetitive protein (PARP) genes, described below, from insect
trypanosomes, is not sensitive to a-amanitin. All
other protein-coding trypanosome genes are transcribed by a normal, amanitin-sensitive polymerase II. In this case, there is either an amanitininsensitive form in addition to the normal polymerase II, or the normal polymerase II can be
made insensitive by means of some additional
factor [59, 157]. As for all mRNAs of the Kinetoplastida, the VSG mRNAs contain a 5'-terminal
mini-exon of 35 nt that is introduced by transsplicing (p. 39). The primary translation product
(pre-VSG) has not only a relatively long, 25- to
33-bp N-terminal signal sequence but also a Cterminal extension of 17-23 non-polar amino
acids. Immediately after translation, but whilst
still in the endoplasmic reticulum, this is
exchanged for the phospho glycolipid which
anchors the VSG to the cell membrane. A phospholipase C can release the VSG, in which case
sn-l,2-dimyristylglycerol remains behind in the
membrane. This enzyme, which may playa role in
the exchange ofVSG variants, has been isolated;
it has a molecular weight of 39 kDa and is specific
for VSGs [20]. The procyclic stages of T. brucei
from the intermediate host Glossina sp. or from
cultures at 27°C have no VSGs on their surface
but instead have PARPs. This is a family of invariable proteins composed mainly of Glu-Pro
repeats with a single N-glycosylation site [33].
6.9.4 Surface Proteins of Other Protozoa
The South American T. cruzi, the agent causing
Chagas' disease, has no VSG system. In this case,
both the epimastigotes from the insect host and
the trypomastigotes from mammalian blood have
different glycoproteins on their surfaces, but
these do not show the variability typical of VSGs
[192]. The penetration into cells by parasitic
forms in vertebrate blood is inhibited by antibodies against the surface protein. The gene for this
protein has been sequenced: the coding sequence
contains the nonapeptide DKKESGDSE in five