Science - USA (2021-11-05)

RESEARCH ARTICLE SUMMARY

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IMMUNOLOGY

Small proline-rich protein 2A is a gut bactericidal

protein deployed during helminth infection

Zehan Hu†, Chenlu Zhang†, Luis Sifuentes-Dominguez, Christina M. Zarek, Daniel C. Propheter,
Zheng Kuang, Yuhao Wang, Mihir Pendse, Kelly A. Ruhn, Brian Hassell, Cassie L. Behrendt,
Bo Zhang, Prithvi Raj, Tamia A. Harris-Tryon, Tiffany A. Reese, Lora V. Hooper*

INTRODUCTION:The mammalian intestinal tract
is colonized with bacteria, fungi, viruses, and
parasites, which present a diverse array of im-
munological challenges. A number of antimi-
crobial mechanisms have evolved to cope with
these challenges. A key defense mechanism
is the production of antimicrobial proteins
(AMPs), which rapidly inactivate microorgan-
isms by targeting their cell wall or cell mem-
brane structures. However, we still have a
limited understanding of the repertoire of gut
AMPs and how these AMPs work cooperatively
to regulate intestinal microbial communities
and protect against infection in varied immu-
nological settings.

RATIONALE:Prior studies have shown that the
gene encoding small proline-rich protein 2A
(SPRR2A) is highly up-regulated in the mouse
small intestine after colonization with com-
mensal bacteria. To elucidate the function of
SPRR2A in the intestine, we studied its regu-
lation, mechanism of action, impact on the
microbiota, and ability to limit pathogenic
bacterial infection. When mice are infected
with intestinal helminths (parasitic worms),
Sprr2aexpression is further elevated above
the levels induced by the commensal micro-
biota. Helminth infections drastically alter in-

testinal immunological responses and cause

epithelial damage and increased tissue inva-

sion by intestinal bacteria. We therefore also

addressed the molecular basis for SPRR2A

up-regulation and its physiological role during

helminth infection in our study.

RESULTS:We report that SPRR2A is an intes-

tinal bactericidal protein that is not phyloge-

netically related to any previously discovered

mammalian AMP. Experiments in mice re-

vealed that SPRR2A was induced by the in-

testinal microbiota in specialized secretory

cell lineages (Paneth cells and goblet cells) of

the intestinal epithelium. SPRR2A was also

expressed in the human intestine, where it se-

lectively localized to goblet cells. Transcripts

encoding SPRR2A were detected in large in-

testinal biopsies and stool from inflammatory

bowel disease patients but not from those of

healthy controls.

We conducted in vitro biochemical studies

to identify the biological activity of SPRR2A.

Purified recombinant SPRR2A selectively killed

Gram-positive bacteria in vitro, whereas Gram-

negative bacteria were resistant to killing by

SPRR2A. The SPRR2A bactericidal mechanism

involved binding to negatively charged lipids

and disrupting bacterial membranes.

We generatedSprr2a-deficient mice and

found that overall bacterial loads remained the

same. However, the absence of SPRR2A led to

the expansion of Gram-positive bacteria in the

small intestinal lumen and mucus layer, con-

sistent with the selective bactericidal activity of

SPRR2A against Gram-positive bacteria in vitro.

Sprr2a-deficient mice also showed an increased

susceptibility to infection byListeria mono-

cytogenes, a Gram-positive intestinal pathogen.

Intestinal helminth infections can cause

damage to the intestinal epithelium, which

promotes bacterial breach of the intestinal

barrier and highlights the need for AMP ex-

pression during infection with these parasitic

worms. Although other AMPs [such as regen-

erating family member 3 gamma (REG3G)

and lysozyme] showed reduced expression

during infection of mice with the intestinal

helminthHeligmosomoides polygyrus, SPRR2A

expression was induced above the levels elicited

by bacterial colonization. Helminth infection

elicits the production of type 2 cytokines, such

as interleukin (IL)–4 and IL-13. SPRR2A dif-

fered from other intestinal AMPs in that it was

further induced by type 2 cytokines during

infection. Accordingly,Sprr2ainduction during

helminth infection required STAT6, a tran-

scription factor that acts downstream of IL-4

and IL-13 to induce type 2 immunity. Finally,

SPRR2A was necessary to prevent intestinal

bacteria, mainly the Gram-positive subset,

from invading the intestinal barrier during

H. polygyrusinfection.

CONCLUSION:SPRR2A is an intestinal antibac-

terial protein that is unrelated to any previously

discovered mammalian AMP. SPRR2A contrib-

utes to intestinal innate immunity by shaping

the composition of the gut microbiota and by

limiting pathogenic infection by Gram-positive

bacteria. A distinctive aspect of SPRR2A func-

tion is its essential role during helminth infec-

tion of the intestine. Although such infection

lowers the expression of a number of intestinal

AMPs, SPRR2A expression is selectively in-

creased by the type 2 cytokines that are produced

duringhelminthinfection.SPRR2Adefendsthe

intestinal barrier against bacterial invasion and

dissemination that is provoked by helminth

infection. This study highlights how different

AMPs may be produced in distinctive immu-

nological settings and deepens our understand-

ing of why a diverse repertoire of epithelial

AMPs has evolved for host immune defense.

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RESEARCH

710 5 NOVEMBER 2021•VOL 374 ISSUE 6568 science.orgSCIENCE

The list of author affiliations is available in the full article online.

*Corresponding author: Email: [email protected]

(Z.H.); [email protected] (T.A.R.);

[email protected] (L.V.H.)

These authors contributed equally to this work.

Cite this article as Z. Huet al.,Science 374 , eabe6723

(2021). DOI: 10.1126/science.abe6723

READ THE FULL ARTICLE AT

https://doi.org/10.1126/science.abe6723

Helminths

(parasitic worms)

Gut

microbiota

Type 2 cytokines

(IL-4, IL-13)

SPRR2A

Sprr2a

Intestine Epithelium

Helminth-induced

bacterial invasion

SPRR2A protects the intestinal barrier during helminth infection.SPRR2A is an antimicrobial protein
produced by intestinal epithelial cells. During infection with the parasitic helminthH.polygyrus, SPRR2A
expression is boosted by the type 2 cytokines IL-4 and IL-13. SPRR2A kills bacteria by damaging their
membranes and thus limits helminth-induced invasion of intestinal tissue by the gut microbiota.