1. Cytotoxic T cell Helper T cell Tc TH T cell receptor Class I MHC
protein
T cell receptor
Fragments
of foreign TH
Class II
Helper T cell
Cytotoxic T cell
infected
target cell
Antigen
presenting cell

2. Helper T cells:
• Express CD4 molecule on the cell surface.
• Recognize antigen peptides bound to class II MHC molecules (exogenous or ingested antigens).
• Provide help to B cells, macrophages and other T cell effector responses though the production of cytokines
Cytotoxic T cells:
• Express CD8 molecule on the cell surface.
• Recognize antigen peptides bound to class I MHC molecules (endogenously produced antigen such as viral antigen).
• Kill infected target cells.

3. IL-2 Gene NFAT Z Construct lac Z TK promoter Hygromycin
Resistance gene
NF-AT
IL-2
TATA
-70 +1
+47
NFAT Z Construct
NFIL-2D
NFIL-2C
NFIL-2B
NFIL-2A
IL-2 Gene
-286
-257
-256
-242
-208
-188
-158
-145
-93
-66

4. Detection of ß-Galactosidase:
Upon addition of its chromogenic substrate, chlorophenol red galactoside (CPRG), ß-galactosidase hydrolyzes CPRG, turning it from a yellow color to purple at alkaline pH.
The color changs thus provide a convenient measure of gene transcription activities during T cell activation.

5. IL-2 Receptor Ca IP3 Ca2+ NF-ATc IL-2 DAG PKC NF-ATn Ras, Rac, PKC
T cell
receptor
activation
IP3
Ca2+
NF-ATc
Phosphoinositol
pathway
IL-2
DAG
PKC
NF-ATn
Ras, Rac, PKC
NF-ATn
NF-ATc
PO3 Ca 2+
Calcineurin
NF-AT
kinase
CsA/Cph
CRAC
IL-2
Receptor
IP3R

6. Con A EGTA Ionomycin thapsigargin Ca CsA/Cph PMA IL-2 Receptor Ca 2+
CRAC
Ionomycin Ca 2+
thapsigargin
IP3R
Ras, Rac, PKC
CsA/Cph
PMA
NF-ATc
PO3
Calcineurin
NF-AT
kinase
NF-ATn
NF-ATc
IL-2

7. This week’s experiments
1. Mammalian cell culture:
Goal: learn how to count cells using a hemacytometer
Learn to grow mammalian cell lines
2. Signal transduction lab:
Examine the effects of various inhibitors and activators on
IL-2 expression.

8. How to count cells using a hemacytometer?
Cell unit per box: 1 x 104 cells/ml
Blue cells : 3 x 104/ml