Science - USA (2022-05-27)

PILRais a transmembrane protein broadly
expressed on myeloid cells, including macro-
phages, dendritic cells, and neutrophils ( 18 , 19 ).
To evaluate its role in maintaining CD8+Tcell
quiescence, we generated a specific mAb against
mouse PILRa(clone 9B12) that could block
the PILRa–CD8ainteraction (Fig. 3G). Ad-
ministration of 9B12 to B6 WT mice decreased
CD8+Tcellsurvivalinthelymphnodeasearly
as day 1 after antibody treatment, whereas de-

creased CD8+T cell survival was not observed

inthespleenuntilday15(fig.S18AandFig.4A).

Similar results were obtained with thymectom-

ized mice (fig. S18B and Fig. 4B), indicating

that the effect of 9B12 was not dependent on

the thymus. The decrease of CD8+T cells was

not due to mAb-mediated depletion because

PILRais not expressed by T cells ( 19 ). In

contrast to the decrease in CD8+T cells, CD4+

T cell survival appeared to be unaffected by

9B12 treatment (fig. S18C and Fig. 4C). In

addition, blockade of PILRawith 9B12 re-

sulted in increased CD8+T cell activation with

increased expression of CD69 and interferon-g

(IFN-g) (Fig. 4, D and E). CD8+T cells from

b-actin luciferase transgenic mice were pu-

rified and transferred into B6 albino mice to

facilitate real-time imaging analysis in vivo.

Transferred CD8+T cells remained in the lym-

phoid organs, and their total cell number

Zhenget al., Science 376 , 996–1001 (2022) 27 May 2022 2of6

0.032%

0.021%

CFSE

CD45.2

lymph node

0.00

0.02

0.04

0.06

0.08

CFSE

A

C

spleen

memory T cells

(spleen)

naive T cells

(spleen)

spleen

Cd8a+/+

Cd8aloxp/loxp

Cd8a+/+ Cd8aloxp/loxp

CFSE

+CD45.2

+ %

0.15%

0.087%

**

0

5000

10000

15000

CFSE

+CD45.2

+^

cell number

**

CFSE

+CD

45.2

+ %

lymph node

** *

0.00

0.01

0.02

0.03

0.04

*

10000

20000

30000

40000

spleen

CFSE

+CD45.2

+^

cell number

0

Cd8aloxp/loxp

Cd8a+/+

0.00

0.05

0.10

0.15

0.20

0.25

CFSE

+CD45.2

+ %

0

20000

40000

60000

80000 **

spleen

CFSE

+CD45.2

+^

cell number

CD45.2

Cd8a+/+ Cd8aloxp/loxp

Cd8a+/+ Cd8aloxp/loxp

Cd8a+/+ Cd8aloxp/loxp

Cd8a+/+ Cd8aloxp/loxp

Cd8a+/+ Cd8aloxp/loxp

CD45.2

CFSE

0.055%

0.029%

Cd8a+/+

Cd8aloxp/loxp

memory T cells

(lymph node)

0.37%

0.20%

B

CD45.2

CFSE

naive T cells

(lymph node)

D

Cd8a+/+ Cd8aloxp/loxp

****

lymph node

0.0

0.1

0.2

0.3

0.4

CFSE

+CD45.2

+ %

0.5

**

Cd8a+/+ Cd8aloxp/loxp

lymph node

0

10000

20000

30000

40000

50000

CFSE

+CD45.2

+^

cell number

Cd8a+/+

Cd8aloxp/loxp

Fig. 1. Inducible genetic deletion ofCd8adisrupts the homeostasis of
memory and naive CD8+T cells in the periphery.(A andB)MemoryCD44hiCD8+
T cells isolated by fluorescence-activated cell sorting (FACS) fromCre+/+Cd8a+/+
(Cd8a+/+)orCre+/+Cd8aloxp/loxp(Cd8aloxp/loxp) mice were CFSE labeled and
transferred into B6 congenic CD45.1 mice followed by tamoxifen treatment.
The percentage and absolute number of CD45.2+CFSE+cells in the lymph node
(A) and spleen (B) cells were examined 34 days after tamoxifen treatment.
(C andD) FACS-isolated naïve CD44loCD8+T cells fromCre+/+Cd8a+/+(Cd8a+/+)
or Cre+/+Cd8aloxp/loxp(Cd8aloxp/loxp) mice were CFSE labeled and transferred

into B6 congenic CD45.1 mice followed by tamoxifen treatment. The percentage

and absolute number of CD45.2+CFSE+cells in the lymph node (C) and

spleen (D) cells were examined 36 days after tamoxifen treatment. (A to D) (left

panel) Representative flow cytometry plots of CD45.2+CFSE+percentage in

the lymph node or spleen of each group 34 (A and B) and 36 (C and D) days

after tamoxifen treatment; (right panel): bar graphs including all mice from each

group.n = 5 mice per group in (A) to (D). Data shown in (A) to (D) are

representative of at least three independent experiments. Mean ± SD is shown.

*P< 0.05, **P< 0.01, ****P< 0.0001 by unpaired Student’s t test.

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