0905-0925 - g amarante-mendes -uoft-usp oncology ... ·...
TRANSCRIPT
"Novel role for galec-n-‐1 in
CTL killing" Gustavo P. Amarante-‐Mendes
UofT-‐USP Oncology Conference November 2013
MOLECULAR CELL BIOLOGY LABORATORY DEPARTMENT OF IMMUNOLOGY
INSTITUTE OF BIOMEDICAL SCIENCES UNIVERSITY OF SÃO PAULO
Ins-tute for Inves-ga-on in Immunology Na-onal Ins-tutes for Science and Technology (INCT)
RESEARCH INTERESTS
Molecular Control of Cell Death
Cell Death Signaling in Cancer
Cell Death in the Immune System
Cell Death Signaling in infec-ons
RESEARCH INTERESTS
Molecular Control of Cell Death
Cell Death Signaling in Cancer
Cell Death in the Immune System
Cell Death Signaling in infec-ons
Proteomic analysis of cytotoxic granules
5 8 pI
42
3
6
7
8
10 11
12
14
13
31
18
26 25
28
21
22
16 17
19 20
27
5
29
30
9
23
15
24
1
10%
SD
S-P
AG
E
Histone H2B 40S ribosomal protein S16
Histone H2A Profilin I Myosin light chain alkali (MLC3nm) Cysta-n C
Galec1n-‐1
Cofilin 40s ribosomal protein S10
Ubiqui-n carboxyl-‐terminal hydrolase isozyme L1 (UCH-‐L1) Cyclophilin B
Cyclophilin A superoxide dismutase [Cu-‐Zn]
Peroxiredoxin 1 Adenine phosphoribosyltransferase/APRT
Ferri-n light chain 1
60S ribosomal protein L9 Rab-‐2A
Transla-onally controlled tumor protein (TCTP)
Granzyme C
Granzyme A Granzyme B Granzyme E Granzyme C
Alpha enolase
Granzyme A Granzyme E Granzyme G Granzyme F Granzyme B Purine nucleoside phosphorylase Phosphomannomutase 2 DNAse gamma/DHP2/LS-‐DNAse/DNAse I-‐like 3 Ras-‐related Rab-‐17
Protein dissulfide isomerase A3 precursor (ERp60)
Phosphoglycerate kinase 1 Calneuron 1
Alpha enolase
Pyruvate kinase, M2 isozyme Protein dissulfide isomerase A3 precursor (ERp60) T-‐complex protein 1 (TCP-‐1-‐teta)
Perforin Zinc finger protein 94 (Zfp-‐94)
Heat shock cognate 71kDa protein 78kDa glucose-‐regulated protein precursor (BIP) Heat shock cognate 71kDa protein
NEDD1 protein
Heat shock protein HSP90-‐beta (TSTA)
Perforin
5 8 pI
4 2
3
6 7
8
10 11 12
14
13
31
18
26 25
28
21 22
16 17
19 20
27
5
29 30
9
23
15
24
1
10%
SD
S-P
AG
E
1D and 2D Gel-‐Analysis
Galec-ns • β-‐galactoside binding lec-ns. • Evolu-onarily conserved. • 15 mammalian galec-ns iden-fied.
• Wide -ssue distribu-on.
• Have at least one conserved carbohydrate recogni-on domain (CRD) of approximately 130 amino acids that mediates carbohydrate binding.
• Involved in several biological processes, such as T cell homeostasis, cell signaling, prolifera-on and apoptosis.
Galectin-1 modulates the early stages of T cell activation by limiting the "membrane raft" coalescence and promote incomplete TCR ζ chain phosphorylation allowing only the response of T cells that require partial TCR signaling.
Galectin-1 expression decreases the TCR avidity and regulates the in vivo TCD8+ cells expansion. It is secreted by activated TCD8+ effector cells.
Galec-n-‐1
Experimental Design and Results
Immuniza-on with Ad5βgal or Ad5ASP2
Day 0
i.v. injec-on of syngeneic splenocytes labeled with 1mM
(non target) or 10mM (target) CFSE (1:1 ra-o)
Day 7
Spleen were analyzed by Flow
Cytometry
Day 8
Target cells are pulsed with cognate
pep-de
Med
ium
ICP
Wild Type Gal-‐1 KO non immunized non immunized immunized immunized
Gal-‐1KO mice displayed a higher frequency of an-gen-‐specific T cells compared to WT mice.
TCD8-‐mediated killing
Cellular and Molecular Immunology, 6th Edition
Target cell Apoptosis
Apoptosis
perforin and granzymes release
Granzymes
FAS-FASL interaction
Gal-‐1KO CD8 T cells are as efficient at killing pep-de-‐pulsed FAS-‐deficient target cells
v v v v
ACKNOWLEDGMENTS
Molecular Cell Lab Departamento de Imunologia (ICB-‐USP)
Tiago C. Machado Narciso Vieira Junior
Trinity College Dublin (Ireland)
Seamus J. Mar-n Colin Adrain
Alexander Luthi
Universidade Federal de São Paulo (UNIFESP)
Mariana Dominguez Mauricio Mar-ns Rodrigues
Faculdade de Medicina Ribeirão Preto (USP)
Marcelo Dias Baruffi
Université de Lausanne
Jürg Tschopp
Ins-tut Curie -‐ CNRS (France)
Sebas-an Amigorena