ICI 2016_Abstract Book

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International Congress of Immunology 2016 Abstract Book
RLTPR comprises concatenated leucine-rich repeats (LRR) and
mice harboring a mutation - called Basilic - in one of those
repeats mimicked fully a CD28 deficiency.
The lymphoid lineage-specific actin-uncapping protein RLTPR
is essential for costimulation via CD28 and the development of
regulatory T cells. RLTPR colocalizes with CD28 at the immune
synapse and couples CD28 to the Carma1 cytosolic effectors.
However, the precise molecular function of RLTPR in CD28
costimulation and role in humans remains elusive. We have
used quantitative mass spectrometry and
T cells from mice in which a tag for affinity purification was
knocked in the Rltpr gene to determine the composition and
dynamics of the signaling complex that formed around RLTPR
following T cell stimulation. We demonstrated that a physical
association existed between RLTPR and upstream (CD28) and
downstream (CARMA1) components of the CD28 pathway.
Proteins not associated before with the CD28 pathway were
also identified in the RLTPR interactome. By developing
RLTPR deficient mice, we showed that the Basilic mutation
corresponded to a null mutation, thereby establishing the
functional importance of the LRR domain. Using Crispr-Cas9-
based gene editing of Jurkat T-cell line, we demonstrated that
RLTPR is also essential in human for costimulation via CD28. Our
findings underpin the similar role exerted by RLTPR in human
and mouse T cells and point in a new direction regarding its
mode of action during CD28 costimulation.
3906
“Microsynapse” composed of focal adhesion molecules
surrounding TCR microcluster is essential for T cell
activation
Saito, T.1,2, Hashimoto-Tane, A.1
1RIKEN Center for Integrative Medical Sciences, Laboratory for Cell
Signaling, Yokohama, Japan, 2Osaka University, WPI Immunology
Frontier Research Center, Suita, Japan
Immunological synapse (IS) formed at the interface between
T cells and antigen-presenting cells represent hallmark of
initiation of acquired immunity. T cell activation is initiated
at T cell receptor (TCR) microclusters (MCs), in which TCR and
signaling molecules assemble at the interface prior to IS
formation. We found that each TCR-MC was transiently bordered
by a ring structuremade of integrin and focal adhesionmolecule
in early phase of activation, which is similar structure to mature
immunological synapse (IS) in micro-scale and we named
“Microsynapse”. The micro adhesion-ring in the microsynapse is
composed of LFA-1, focal adhesion molecules such as paxillin
and Pyk2, and myosin II (MyoII), and is supported by F-actin
core and MyoII activity through LFA-1 outside-in signals. The
formation of microsynapse and the adhesion-ring was transient
and especially sustained upon weak TCR stimulation to recruit
LAT and SLP76. Perturbation of the microsynapse induced
impairment of TCR-MC development and resulted in impaired
cellular signaling and T cell activation and functions. Thus, the
microsynapse composed of the core TCR-MC and surrounding
micro adhesion-ring is a critical structure for initial T cell
activation through integrin outside-in signals.
629
TRAF adaptors limit IL-6 receptor signaling through an
unexpected binding to the signaling transducer receptor
gp130
So, T.1, Nagashima, H.1, Okuyama, Y.1, Hayahi, T.1, Asao, A.1,
Kawabe, T.1, Yamaki, S.1, Nakano, H.2, Croft, M.3, Ishii, N.1
1 Tohoku University Graduate School of Medicine, Microbiology and
Immunology, Sendai, Japan, 2Toho University School of Medicine,
Biochemistry, Tokyo, Japan, 3La Jolla Institute for Allergy and
Immunology, Immune Regulation, La Jolla, United States
There is growing evidence that TNF receptor-associated factors
(TRAFs) are recruited to unconventional cytokine receptors and
control their key signaling pathways in lymphocytes. We have
found that TRAF5 expressed in CD4+ T cells constitutively binds
to a cytoplasmic region of the signal transducing receptor gp130
through its C-terminal TRAF domain and inhibits the recruitment
and activation of STAT3 mediated by IL-6. This function of
TRAF5 significantly impacts IL-6-driven Th17 cell differentiation.
However, it is not clear how TRAF5 inhibits the gp130-STAT3
signaling axis and whether other TRAF family molecules
contribute to this process. We found that amino acid residues
774-798 in gp130, which contain TRAF-binding consensus
motifs, were critical not only for association with TRAF5 but also
with TRAF2. gp130 expressed on naïve CD4+ T cells gradually
decreased after T cell activation and its expression reached a
minimum level at 48 h. Whereas TRAF5 rapidly disappeared from
activated naïve T cells within 4 h, TRAF2 was stably expressed in
primed T cells during the course of Th17 development. These
results demonstrate that both TRAF2 and TRAF5 inhibit the
early signaling activity of the IL-6 receptor complex in naïve
CD4+ T cells and that TRAF2 works as a negative regulator even
in the later stage of Th17 development. Accordingly, shRNA-
mediated Traf2-knockdown in differentiating Traf5-/- CD4 + T cells
strongly promoted IL-6-driven Th17 differentiation. We propose
a novel signaling mechanism of Th17-lineage commitment that
is spatiotemporally regulated by the adaptor proteins TRAF2
and TRAF5.
3633
A structured pathway for T cell receptor signal transmission
through the membrane
Krshnan, L.1,2, Park, S.3, Im, W. 3, Call, M.1,2, Call, M.1,2
1 Walter & Eliza Hall Institute of Medical Research, Structural
Biology, Parkville, Australia, 2The University of Melbourne, Parkville,
Australia, 3University of Kansas, Department of Molecular
Biosciences and Center for Computational Biology, Lawrence,
United States
The octameric T cell receptor (TCR)-CD3 complex signals the
recognition of antigenic peptide:MHC ligands on target cells
by initiating a cascade of intracellular biochemical events
beginning with phosphorylation of immunoreceptor tyrosine-
based activation motifs (ITAMs) on CD3 and ζ cytoplasmic
tails by the src-family kinase lck. How ligand-sensing is
communicated across the cell membrane is an important
unresolved problem in T cell biology, and a lack of structural
information on the intact receptor complex stands as a major