Cell Host & Microbe

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production of polyamines is normally

restrictive to S. aureus growth, given the

sensitivity of most strains to these mole-

cules (Joshi et al., 2011). However,

USA300 strains contain ACME-encoded

speG, which confers resistance to poly-

amine toxicity in vitro (Joshi et al., 2011).

Here, the authors extend this finding using

in vivo models to demonstrate that speG

mutants in USA300 display increased

sensitivity to polyamines during the

postinflammatory phase of skin and soft

tissue infection. Further, their data indi-

cate that bacterial burden in late-stage

abscesses is largely dependent on

SpeG, a novel finding that links ACME to

persistence during skin and soft tissue

infection.

Thurlow et al. continue their investiga-

tion into the role of ACME in skin infections

by addressing an apparent paradox

between their findings and that of earlier

studies, which were unable to uncover

a role for ACME in skin and soft tissue

infections (Montgomery et al., 2009). The

authors demonstrate that if speG is

deleted, bacterial burden is significantly

reduced and clearance occurs more

quickly in vivo. However, it was previously

observed that an ACME mutant (lacking

speG and arc) is equally as virulent as an

ACME-containing strain (Montgomery

et al., 2009). Thus, it is somewhat perplex-

ing that an ACMEmutant is not also atten-

uated in vivo. Through the use of murine

infection models, the authors rationalize

these findings by demonstrating that

S. aureus Arc diverts L-arginine away

from NO$ production by the host, thereby

fostering polyamine synthesis, a result of

the increased production of L-ornithine

by the bacterium (Figure 1). The presence

of Arc appears to enhance host polyamine

production and requires SpeG to detoxify

the system. Indeed, deletion of arc in

a speG mutant increases the virulence of

the strain to WT levels. Thus, the fitness

advantage conferred upon USA300 via

ACME requires both Arc and SpeG.

All together, the work presented in this

new study both describes a novel role

for ACME as an important virulence deter-

minant in USA300 skin and soft tissue

infection and provides a unique rationale

for how this strain may have rapidly out-

competed other S. aureus clones for

dominance over the human skin niche.

Furthermore, it highlights the remarkable

adaptability of S. aureus in the face of

host immunity, one that extends beyond

traditional mechanisms of pathogenesis

(toxins, immune-modulatory molecules,

etc.). It is worth mentioning that these

findings do not discount the role of major

virulence factors in S. aureus skin and soft

tissue infections; rather, they add to the

Cell Host & Microbe

ever-evolving armament of S. aureus

survival mechanisms.

REFERENCES

Diep, B.A., Stone, G.G., Basuino, L., Graber, C.J.,Miller, A., des Etages, S.A., Jones, A., Palazzolo-Ballance, A.M., Perdreau-Remington, F., Sensa-baugh, G.F., et al. (2008). J. Infect. Dis. 197,1523–1530.

Foster, T.J. (2005). Nat. Rev. Microbiol. 3,948–958.

Joshi, G.S., Spontak, J.S., Klapper, D.G., and Ri-chardson, A.R. (2011). Mol. Microbiol. 82, 9–20.

Li, M., Diep, B.A., Villaruz, A.E., Braughton, K.R.,Jiang, X., DeLeo, F.R., Chambers, H.F., Lu, Y.,and Otto, M. (2009). Proc. Natl. Acad. Sci. USA106, 5883–5888.

Miko, B.A., Uhlemann, A.C., Gelman, A., Lee, C.J.,Hafer, C.A., Sullivan, S.B., Shi, Q., Miller, M., Zenil-man, J., and Lowy, F.D. (2012). Microbes Infect.14, 1040–1043.

Miller, L.S., and Cho, J.S. (2011). Nat. Rev. Immu-nol. 11, 505–518.

Montgomery, C.P., Boyle-Vavra, S., Adem, P.V.,Lee, J.C., Husain, A.N., Clasen, J., and Daum,R.S. (2008). J. Infect. Dis. 198, 561–570.

Montgomery, C.P., Boyle-Vavra, S., and Daum,R.S. (2009). Infect. Immun. 77, 2650–2656.

Richardson, A.R., Libby, S.J., and Fang, F.C.(2008). Science 319, 1672–1676.

Thurlow, L.R., Joshi, G.S., Clark, J., Spontak, J.,Neely, C.J., Maile, R., and Richardson, A.R.(2013). Cell Host Microbe 13, this issue, 100–107.

How Trypanosoma cruzi Feastsupon Its Mammalian Host

Nicola S. Carter1 and Buddy Ullman1,*1Department of Biochemistry and Molecular Biology, Oregon Health & Science University, 3181 SW Sam Jackson Park Road, Portland,OR 97239-3098, USA*Correspondence: ullmanb@ohsu.eduhttp://dx.doi.org/10.1016/j.chom.2013.01.003

Trypanosoma cruzi has a complex relationship with its mammalian host in which parasite and host metabolicnetworks are intertwined. A genome-wide functional screen of T. cruzi infection in HeLa cells (Caradonnaet al., 2013) divulges host metabolic functions and signaling pathways that impact intracellular parasitereplication and reveals potential targets for therapeutic exploitation.

Chagas’ disease is a devastating, perni-

cious, and often fatal disease of the

cardiovascular system for which the

hemoflagellate protozoan parasite, Try-

panosoma cruzi, is the etiologic agent.

The neurological system and digestive

tract can also be impacted by T. cruzi

infection. Chagas’ disease is endemic to

all 22 countries of South and Central

13, January 16, 2013 ª2013 Elsevier Inc. 5

Cell Host & Microbe

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America and causes the greatest disease

burden of any parasitic disease in

the western hemisphere. Approximately

8–11 million people are affected, with

�10,000 deaths per year and an addi-

tional >25,000,000 people at risk for

infection (http://www.cdc.gov/parasites/

chagas/epi.html). Autochthonous infec-

tion in the United States has also

been documented, and the Centers

for Disease Control now estimate that

�300,000 individuals in the United

States are currently seropositive for the

parasite (Bern and Montgomery, 2009).

Transmitted to humans by insects known

as ‘‘kissing bugs,’’ insects that are

endemic to the southern half of the

United States, the parasite—which can

also be acquired by blood transfusion,

food contamination, organ transplanta-

tion, transplacental passage, birthing, or

nursing—exists in two forms in the human

host: the nondividing flagellated trypo-

mastigote stage that is primarily found in

the bloodstream, and the aflagellated

amastigote stage that replicates within a

variety of host cells (http://www.cdc.gov/

parasites/chagas/biology.html). A major

goal for treating Chagas’ disease is to

stem disease progression; however, con-

ventional Chagas’ remedies are based

upon antitrypanosomal therapeutics, and

the few drugs that are available for treat-

ment are compromised by toxicity and

demonstrate limited efficacy once chronic

cardiomyopathy develops. An alternative

approach for therapeutic development—

one adopted by several laboratories,

including that of Caradonna et al. (2013),

whose research is featured here—has

focused instead on delineating hostmeta-

bolic pathways that are vital for disease

progression and parasite persistence.

Previous investigations evaluating host

factors perturbed by T. cruzi invasion

have primarily focused on profiling the

host transcriptional response after

T. cruzi infection of cardiomyocytes and

fibroblasts, as well as a variety of other

cells, using classical DNA microarray

technology. These studies, which have

contributed tremendously to our knowl-

edge about the invasion processes

utilized by T. cruzi trypomastigotes, as

well as provided links between host dys-

regulation and disease progression,

have shed little light on the host metabolic

processes important for sustaining intra-

cellular T. cruzi infections that are requi-

6 Cell Host & Microbe 13, January 16, 2013 ª

site for the long-term persistence of

this parasite. In this issue of Cell Host &

Microbe, Caradonna et al. describe

a particularly sophisticated set of studies

founded upon an unbiased, genome-

scale, forward genetic screen to identify

host genes and proteins, as well as

their participatory metabolic partners,

that impact the intracellular growth of

T. cruzi. The primary screen conducted

in HeLa cells consisted of a genome-

wide knockdown of 18,263 host mRNAs

and their corresponding proteins using

siRNA technology and offers a powerful,

all-encompassing approach for the iden-

tification of both positive and negative

regulators of T. cruzi growth. The sig-

nificance of the data from the primary

screen, which yielded hundreds of

hits (see Table S1 in Caradonna et al.,

2013), was realized by the implemen-

tation of an ingenious secondary screen

to distinguish those host proteins

important for T. cruzi early infection—

i.e., trypomastigote invasion and early

establishment in the host cell (<18 hr

postinfection)—from those important for

later amastigote growth and viability

(18–72 hr postinfection).

While the results from the early infection

data were largely supportive of the known

models for T. cruzi host cell invasion and

parasitism (reviewed in Caradonna and

Burleigh, 2011), they did identify several

additional new factors that are likely

complicit in parasite attachment and

invasion. However, the major thrust of

this elegant composition was the elucida-

tion of host constituents and processes

that impact amastigote growth and

viability. In one of the most wide-ranging

examinations of intracellular host meta-

bolism and its relationship to intracellular

parasite proliferation and viability,

Caradonna et al. have succeeded in

uncovering several unique facets of host

metabolism that offer opportunities for

future drug exploration. One intriguing

discovery was the apparent shift in

host cell energy production away from

the oxidization of glucose—and more

specifically the conversion of pyruvate

to acetyl-CoA—to the b-oxidation of

long chain and very long chain fatty

acids in the infected host cell. Indeed,

the siRNA data, accompanied by addi-

tional experiments with fibroblasts con-

taining a genetic deficiency in the first

mitochondrial step of the b-oxidation

2013 Elsevier Inc.

of long chain and very long chain fatty

acids, suggest that the production of

very long chain fatty acids by the host

cell and their subsequent oxidation is

likely a requisite for parasite viability and

ultimately, perhaps, for parasite persis-

tence. A key question for future studies

will be how the parasite manipulates and

ultimately benefits from these changes in

the host metabolic landscape. Whether

this is a ploy to mobilize and obtain fatty

acid precursors necessary for the syn-

thesis of membrane in the dividing

amastigote population or, alternatively,

to bolster parasite energy production

by providing reducing intermediates (i.e.,

NADH and FADH2) generated during

the oxidation process remains to be

elucidated. Another intrigue concerns

the fate of pyruvate. The siRNA results,

combined with additional experimental

data, indicate that downregulation of

pyruvate metabolism to either acetyl-

CoA or lactate is beneficial for intracellular

amastigote development. While this

might intimate that the accumulation of

pyruvate in the host cell is of benefit to

the parasite, perhaps as an energy

precursor, pyruvate transport into the

amastigote has yet to be confirmed.

The manipulation of host cell energy

production may not be without con-

sequence, and perturbation of mito-

chondrial function in particular is clearly

associated with programmed cell death

(Kroemer et al., 1995). From the siRNA

screen, a strong relationship between

host cell ATP currency and parasite

growth and viability emerged. In partic-

ular, the downregulation of ATP5B,

a subunit of ATP synthase and presum-

ably a determinant of its activity, nega-

tively impacted parasite growth. In

addition, the opposing effects of siRNA

against AMP-activated kinase (AMPK)

and AKT1, with silencing of the former

boosting parasite intracellular growth

and survival and of the latter being associ-

ated with a reduction in parasitemia,

seems to signify an important connection

between host cell energy balance and

parasite viability. AKT1 is a protein kinase

strongly associated with cell survival, in

part through its repression of proapopto-

tic proteins and activation of TOR kinase

complex 1 (mTORC1), which promotes

protein synthesis and inhibits autophagy.

On the other hand, AMPK, which is

activated in response to rising cellular

Cell Host & Microbe

Previews

levels of AMP, a signal that cellular ATP

levels are decreased, opposes many of

the functions of AKT1 and is itself nega-

tively regulated by AKT1 (Mankouri et al.,

2010). Whether and how the parasite

intervenes in the complex interplay

between these two kinases and their

signaling cascades that mediate cell fate

is unclear but will be of obvious conse-

quence for future studies.

Perhaps the most surprising finding

from this screen, which demonstrates

the power of an unbiased screen such

as this one, was the discovery of the

apparent dependence of the T. cruzi

amastigote on the host for the provision

of preformed pyrimidines, which was

functionally authenticated by the authors

by media supplementation, as well as

the preference for salvageable purine

nucleosides over nucleobases, which

remains to be functionally tested. These

parasites have a full complement of

pyrimidine biosynthesis enzymes located

in a cluster on chromosome 21, as well

as several putative orthologs within their

genome to the leishmanial nucleoside

and nucleobase transporters NT1–NT4

(http://tritrypdb.org/tritrypdb/ and Carter

et al., 2008). Thus, a nutritional depen-

dency on salvageable pyrimidines and

purine nucleosides is quite unexpected.

It is noteworthy that recent studies with

T. cruzi engineered to lack carbamoyl-

phosphate synthetase II (CPSII), the

first enzyme in pyrimidine biosynthesis,

also suggested a functional significance

between this first pyrimidine biosynthetic

step and intracellular parasite develop-

ment (Hashimoto et al., 2012), under-

scoring the significance of pyrimidine

metabolism and the contributions of

both parasite salvage and biosynthesis

pathways for host cell parasitism. Other

nutritional interactions revealed by this

screen between parasite and host, for

example, pteridine metabolism, were

less surprising but confirmed the power

of the general forward genetic approach.

Overall, this investigation has provided

numerous incisive insights into the host

response to T. cruzi infection and persis-

tence and lends itself to the exploita-

tion of previously unpredictable host

Cell Host & Microbe

determinants that could be pharmaco-

logically targeted in a fashion that mini-

mizes the emergence of long-term drug

resistance.

REFERENCES

Bern, C., andMontgomery, S.P. (2009). Clin. Infect.Dis. 49, e52–e54.

Caradonna, K.L., and Burleigh, B.A. (2011). Adv.Parasitol. 76, 33–61.

Caradonna, K.L., Engel, J.C., Jacobi, D., Lee,C.-H., and Burleigh, B.A. (2013). Cell Host Microbe13, this issue, 108–117.

Carter, N.S., Yates, P., Arendt, C.S., Boitz, J.M.,and Ullman, B. (2008). Adv. Exp. Med. Biol. 625,141–154.

Hashimoto, M., Morales, J., Fukai, Y., Suzuki, S.,Takamiya, S., Tsubouchi, A., Inoue, S., Inoue, M.,Kita, K., Harada, S., et al. (2012). Biochem.Biophys. Res. Commun. 417, 1002–1006.

Kroemer, G., Petit, P., Zamzami, N., Vayssiere,J.L., and Mignotte, B. (1995). FASEB J. 9, 1277–1287.

Mankouri, J., Tedbury, P.R., Gretton, S., Hughes,M.E., Griffin, S.D., Dallas, M.L., Green, K.A.,Hardie, D.G., Peers, C., and Harris, M. (2010).Proc. Natl. Acad. Sci. USA 107, 11549–11554.

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