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6

International Congress of Immunology 2016

Abstract Book

between the remaining

Irf8

-/-

progenitors and their wild-type

counterparts. Nevertheless, the enhancer landscapes of

Irf8

-/-

progenitors were significantly distinct from those of wild-type

cells, with the enhancer landscapes of

Irf8

-/-

GMPs, MDPs, and

cMoPs all remaining similar to that of wild-type GMPs. IRF8

was required particularly for the establishment of enhancers

common in monocytes and DCs. These results illustrate that

enhancer establishment and gene expression are dynamically

and hierarchically regulated by transcription factors during

mononuclear phagocyte development.

1506

Batf2/Irf1 induces inflammatory responses in classically

activated macrophages, lipopolysaccharides, and

mycobacterial infection

Roy, S.

1,2

, Guler, R.

3,4

, Parihar, S.P.

3,4

, Schmeier, S.

5

, Kaczkowski, B.

1,2

,

Nishimura, H.

1,2

, Shin, J.W.

1,2

, Negishi, Y.

1,2

, Ozturk, M.

3,4

, Hurdayal,

R.

3,4

, Kubosaki, A.

1

, Kimura, Y.

1

, de Hoon, M.J.L.

1,2

, Hayashizaki, Y.

2,6

,

Brombacher, F.

3,4

, Suzuki, H.

1,2

1

RIKEN Center for Life Science Technologies, Yokohama, Japan,

2

Riken Omics Science Center, Yokohama, Japan,

3

University of Cape

Town/Institute of Infectious Disease and Molecular Medicine (IDM),

Division of Immunology, Cape Town, South Africa,

4

International

Centre for Genetic Engineering and Biotechnology (ICGEB), Cape

Town Component, Cape Town, South Africa,

5

Massey University,

Institute of Natural and Mathematical Sciences, North Shore

City, New Zealand,

6

Riken Preventive Medicine and Diagnosis

Innovation Program (PMI), Yokohama, Japan

Basic leucine zipper transcription factor Batf2 is poorly

described, whereas Batf and Batf3 have been shown to play

essential roles in dendritic cell, T cell, and B cell development

and regulation. Batf2 was drastically induced in IFN-γ-activated

classical macrophages (M1) compared with unstimulated or IL-

4-activated alternative macrophages (M2). Batf2 knockdown

experiments

from

IFN-γ-activated

macrophages

and

subsequent expression profiling demonstrated important roles

for regulation of immune responses, inducing inflammatory

and host-protective genes Tnf, Ccl5, and Nos2.

Mycobacterium

tuberculosis

(Beijing strain HN878)-infected macrophages

further induced Batf2 and augmented host-protective Batf2-

dependent genes, particularly in M1, whose mechanism was

suggested to be mediated through both TLR2 and TLR4 by

LPS and heat-killed HN878 (HKTB) stimulation experiments.

Irf1 binding motif was enriched in the promoters of Batf2-

regulated genes. Coimmunoprecipitation study demonstrated

Batf2 association with Irf1. Furthermore, Irf1 knockdown

showed downregulation of IFN-γ- or LPS/HKTB-activated host-

protective genes Tnf, Ccl5, Il12b, and Nos2. Batf2 deficiency

in mice resulted in resistance to

M. tuberculosis

infection with

reduced immunopathology. Conclusively, Batf2 is an activation

marker gene for M1 involved in gene regulation of IFN-γ-

activated classical macrophages, as well as LPS/HKTB-induced

macrophage stimulation, possibly by Batf2/Irf1 gene induction.

Taken together, these results underline the role of Batf2/Irf1 in

inducing inflammatory responses in

M. tuberculosis

infection.

2879

The effects of transgenic expression of a T-box molecule,

eomesodermin, in naive T cells on their activation and

exhaustion status

Eshima, K., Misawa, K., Noma, H., Iwabuchi, K.

Kitasato University School of Medicine, Department of

Immunology, Sagamihara, Japan

A T-box family transcription molecule, Eomesodermin (Eomes),

controls effector function of CD8

+

CTLs. Eomes is expressed

in effector or memory CD8

+

T-cells, but is also found in

“exhausted” cells, although its precise roles there remain to be

fully delineated. In order to determine the relevance of Eomes

in T-cell activation statuses, we developed Eomes-transgenic

mouse using human CD2 promoter/enhancer. In Eomes-Tg

mouse, the number of peripheral T-cells was smaller than

that in wild type mice. This was accompanied with decreased

expression of CD127 and Bcl-2 in Tg mature T-cells, which is

frequently observed in exhausted T-cells. It was also noticed

that higher proportion of peripheral CD8

+

T-cells in Eomes-Tg

mice expressed some of exhausted T-cells markers, such as,

PD-1, LAG-3, or TIM-3, suggesting that part of exhausted T-cell

phenotypes may result from higher expression of Eomes. As

for other activation markers, expression of CD44, CD122, Ly-6C

was also up-regulated on Eomes-Tg CD8

+

T-cells. By utilizing

Tg-TCR, it was suggested that optimal expression of many of

these markers, except for CD122, still requires TCR-stimulation.

Transgenic expression of Eomes alone may not be adequate

for provision of functional activity to CD8

+

T-cells, either, since

CD44

lo+

CD8

+

T-cells from Eomes-Tg did not produce IFN-γ or

exert cytotoxicity until antigenically primed. Collectively, these

results imply that Eomes induction alone may be insufficient for

proper CTL differentiation and that Eomes expression without

TCR stimulation might induce exhausted phenotypes.

2692

Deciphering the importance of H3K27me3 demethylation

during early CD8

+

T cell responses against influenza A virus

Li, J., Olshansky, M., Turner, S.

University of Melbourne, Department of Microbiology &

Immunology, Melbourne, Australia

The acquisition and maintenance of cytotoxic CD8

+

T cell

(CTL) function in response to influenza virus infection is partly

regulated by co-ordinated changes in chromatin structure,

histone modifications and binding of transcription factors.

Developmental or lineage-defining genes in stemcells and CD4

+

T cells are bivalently marked by the repressive trimethylated

lysine 27 (H3K27me3) and activating trimethylated lysine 4

(H3K4me3) on the H3 histone. In naïve CD8

+

T cells, we have

also shown that a number of bivalent genes rapidly resolve to

a transcriptionally permissive state when differentiated into

virus-specific effector and memory T cells (Russ

et al

., 2014).

To understand how the early removal of the repressive mark,

H3K27me3, defines immediate transcriptional responses of

bivalent genes, naive CD8

+

T cells were stimulated with the

SIINFEKL peptide for 3, 5 and 24hrs. Whole-genome H3K27me3

chromatin-immunoprecipitation-sequencing (ChIP-seq) and