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Fig. 1. Life cycle and summary of the major findings of proteome analysis in T. cruzi. T. cruzi
trypomastigotes circulate in the blood of infected hosts, including humans, but must enter host cells
(oftentimes muscle cells) and convert to amastigote forms to replicate. Triatomine bug vectors
become infected by ingesting trypomastigotes during the course of a blood meal on infected
mammalian hosts. Conversion of the trypomastigotes into epimastigotes, replication of these
epimastigotes, and their eventual transformation into metacyclic trypomastigotes, occurs in the
insect gut. Metacyclic trypomastigotes initiate new infection in mammals when infected insects
are ingested or by deposition of parasites in the feces, usually during a blood meal.
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hypothetical,[ validating these as bona fide
genes in T. cruzi. More than half of these hypothetical genes have orthologs in the Leishmania major and/or T. brucei genomes.
T. cruzi trypomastigotes circulate in the
blood, where they are exposed to host immune
effector molecules, including specific antibodies.
Unlike the related African trypanosomes, T.
cruzi trypomastigotes do not undergo antigenic
variation but instead express on their surface
multiple members of several large families of
molecules; the best characterized of these are
the mucin and trans-sialidase (ts) families (13).
Thirty of the 50 top-scoring proteins detected
exclusively in trypomastigotes are ts family
members. Likewise, the amastigote and metacyclic stages appear to express subsets of ts
molecules unique to each stage, whereas no ts
expression was detected in the epimastigote
proteome (Fig. 2 and table S2). Trans-sialidase
enzymatic activity is reportedly present in only
a small subset of the 91000 ts proteins encoded
in the T. cruzi genome, and it has been linked to
the presence of Tyr342 in the catalytic Nterminal region and SAPA repeats in the C
terminus (13). Among the 223 ts proteins
detected in the proteome are the products of
all 15 genes predicted to encode enzyme-active
ts. The production of a large number of nonenzymatic ts family members coincident with
these ts enzymes may deflect immune responses
away from the enzymatically active targets or
may provide a pool of altered peptides that
could antagonize T cell responses (14).
In addition to the ts and mucin families,
the T. cruzi genome contains several other
high-copy multigene families (Table 2 and
supporting online text). We detected expression of several mucin-associated surface proteins (MASPs), a gene family first discovered
as part of the sequencing and annotation effort
(10), predominantly in the trypomastigote
proteome. Like proteins from the other multigene families in T. cruzi, many MASP family
members have predicted signal sequences
and glycosylphosphatidylinositol anchor addition sites and thus are likely to be surface
expressed. Nine MASP gene family proteins
were identified in our analysis, each by only a
single peptide match. This result suggests either
that MASPs are not as abundantly expressed as
the trans-sialidase proteins or that, like the
mucins, MASPs have extensive posttranslational modifications that complicate their detection by shotgun proteomics. However, detection
of the MASPs in the relatively undersampled
trypomastigote stage suggests that they are not
minor constituents of the T. cruzi proteome.
The transition from trypomastigote to
amastigote can be stimulated extracellularly
by simulating the low pH environment of the
phagosomal/lysosomal compartment that T.
cruzi initially encounters upon cell entry (15),
making early time points in the transformation
process to the amastigote stage amenable to
474
transcriptome and proteome analysis. The results from this proteome analysis of amastigotes are in agreement (with one exception)
with the restricted data set generated by comparison of trypomastigotes and early-stage
amastigotes using DNA microarray analysis
(3) (table S4), further supporting the quality of
this analysis. In addition to the expression of a
distinct subset of trans-sialidasefamily genes,
many of which are related to the amastigote
surface protein 2 molecule previously reported
to be preferentially expressed in amastigotes
(16) (Fig. 2), the transition of trypomastigotes
to amastigotes also appears to be accompanied
by a dramatic shift from carbohydrate- to lipiddependent energy metabolism (table S3). This
is demonstrated by the virtual absence of glucose transporters and the detection of enzymes
that oxidize fatty acids to give acetylcoenzyme
A. Enzymes of the citric acid cycle, which
oxidize acetyl coenzyme A to carbon dioxide
and water, are also abundant in amastigotes.
Amastigotes are likely to be dependent on gluconeogenesis for the synthesis of glycoproteins and glycoinositolphospholipids (GIPLs),
Table 1. Protein group and protein identifications for each developmental stage.
Protein groups
(proteins)
29
21
44
335
27
65
146
24
43
47
53
12
187
92
43
1168
(49)
(41)
(161)
(838)
(84)
(110)
(538)
(50)
(125)
(122)
(93)
(22)
(315)
(162)
(74)
(2784)
Amastigote
X
X
X
X
X
X
X
X
Trypomastigote
Metacyclic
trypomastigote
X
X
X
X
X
X
Epimastigote
X
X
X
X
X
X
X
X
X
691 (1871)
582 (1486)
X
X
X
X
X
X
X
X
X
X
732 (1861)
969 (2339)
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212
67
61
49
36
223
399
29
30
9
0
155
505
348
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amastigotes (table S3). We also extend the data
on the selective expression in amastigotes and
epimastigotes of several ABC transporters
(7164.t00003, 8319.t00008) that are hypothesized to have a role in cargo selection and/or
vesicular transport in trypanosomes (17). A putative lectin (6865.t00003) with homology to
ERGIC (ER Golgi intermediate compartment),
a protein involved in cargo selection in coat
protein complex II vesicles, is also detected in
trypomastigotes and amastigotes but not in
metacyclic or epimastigote forms.
In contrast to both T. brucei and L. major,
the T. cruzi genome encodes enzymes capable
of catalyzing the conversion of histidine to glutamate. The first two enzymes in this pathway,
histidine ammonia-lyase (6869.t00022) and
urocanate hydratase (4881.t00011), are abundant in the insect stages but nearly undetectable
in the mammalian stages (only a single
spectrum matching histidine ammonia-lyase in
amastigotes), consistent with the functioning of
this pathway primarily in epimastigotes and
metacyclic trypomastigotes. This expression
pattern is notable, given that histidine is the
dominant free amino acid in both the excreta
and the hemolymph of Rhodnius prolixus
(18, 19), a well-studied vector for T. cruzi. The
abundance of histidine in this and other bloodfeeding insects likely reflects the high histidine
content of hemoglobin (20). Thus, T. cruzi
epimastigotes seem particularly adapted among
the kinetoplastids to take advantage of this
plentiful energy source in the gut of its insect
vector. This is analogous to the use of proline
as an energy source by T. brucei (21).
The transformation of epimastigotes to
metacyclic trypomastigotes is accompanied
by the production of a number of key en-
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21.
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26.
Tau Suppression in a
Neurodegenerative Mouse
Model Improves Memory Function
K. SantaCruz,1* J. Lewis,5* T. Spires,6* J. Paulson,2 L. Kotilinek,2
M. Ingelsson,6 A. Guimaraes,2 M. DeTure,5 M. Ramsden,2
E. McGowan,5 C. Forster,1 M. Yue,5 J. Orne,6 C. Janus,5 A. Mariash,2
M. Kuskowski,7 B. Hyman,6 M. Hutton,5 K. H. Ashe2,3,4,7.
Neurofibrillary tangles (NFTs) are the most common intraneuronal inclusion in
the brains of patients with neurodegenerative diseases and have been
implicated in mediating neuronal death and cognitive deficits. Here, we found
that mice expressing a repressible human tau variant developed progressive
age-related NFTs, neuronal loss, and behavioral impairments. After the
suppression of transgenic tau, memory function recovered, and neuron numbers
stabilized, but to our surprise, NFTs continued to accumulate. Thus, NFTs are
not sufficient to cause cognitive decline or neuronal death in this model of
tauopathy.
Neurofibrillary tangles are composed of filaments of hyperphosphorylated tau (14), a
microtubule-associated protein (5). They correlate well with cognitive deficits (68) and
neuron loss (9), and they have been implicated
in mediating neurodegeneration and dementia
in Alzheimer_s disease (AD) (1013) and other
tauopathies (14). Transgenic mice expressing
human tau have consistently demonstrated
neurological deficits and neuron loss appearing
with NFTs (1519). The association between
NFTs, neuron loss, and brain dysfunction in
1
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