1. What is the mechanism of store-operated Ca 2+ entry?

2. Measuring [Ca 2+ ] ER and I CRAC in single cells Miyawaki et al, 1999

3. The ER sensor for CRAC channel activation has low Ca 2+ affinity Possible candidates: Low-affinity, high-capacity (calreticulin, BiP, etc) Low-affinity, multi-EF hand (CREC family: reticulocalbin, ERC-55, etc)

4. References Smani et al, 2004 Yao et al, 1999 But see: Scott et al, 2003 Bakowski et al, 2003 Ma et al, 2000 But see: Sugawara et al, 1997 Prakriya & Lewis, 2001

5. Smani et al, 2004 A recent “diffusible messenger” model for SOC activation

6. The fingerprint of the CRAC channel is a definitive benchmark for evaluating putative CRAC channel genes

7. Clapham, 2003 The family of mammalian TRP channels TRPC1: Mori et al, 2002 (DT40 knockout, inconsistent phenotype) TRPC3: Philipp et al, 2003 (CRAC mutants, rescues TCR but not TG response) TRPV6: Yue et al, 2001 (pore properties don’t match completely) Cui et al, 2002 (dom negative suppression of Icrac)

9. 5C.C7 B6 thymocytes in B10.BR slice Time compression: 80x, total time 60 min Ca 2+ signals in a positively selecting environment

10. Positive selection Neglect Calcium signals during positive selection are oscillatory

11. Dolmetsch et al, 1998 A reductionist approach to the functions of Ca 2+ dynamics in T cells: The Ca 2+ clamp

12. Dolmetsch et al, 1998 Frequency selectivity of transcriptional activation and gene expression

14. Tomida et al, 2003 vs. [Ca]  interval Ca 2+ oscillations boost signaling efficiency

15. Tomida et al, 2003 Delayed phosphorylation and transport of NFAT underlies the signaling efficiency of Ca 2+ pulses