1. Functions of TBX1: an update Antonio Baldini & Elizabeth Illingworth Telethon Institute of Genetics and Medicine & Institute of Genetics and Biophysics Napoli, Italy

2. Goals of Basic Research Molecular diagnostics Identification of the disease genes Fnctional studies Develop the basis for future treatments (conventional or gene therapy)

3. 22 del Diagnosis of del22q11 FISH

4. Gene AGene B A BDEC

5. Del22q11.2 T10 PRODH DGCR6 IDDSTK22AGSCL RANBP1 SLC25A1ARVCFCOMTTRXR2 GNB1L TBX1 GPIB  CDCREL1 CLD5 CDC45 UFD1L NLVCFES2EL HTF9C ZNF74 HIRA CRKL LZTR1 CLTCL 3Mb (90%) Cen 1.5Mb (10%) HCF2

6. Genes and clinical findings in multigene deletion syndromes Deleted region Genes a b c def g h Clinical findings ?

7. Mouse models and human genetic diseases “Chromosome engineering” Generation of the first animal model of del22q11.2 (Lindsay-Illingworth, 1999) Df1 mouse -Interrupted aortic arch -Small Thymus -Small parathyroids -Neurobehavioral abnormalities

8. Identification of the critical gene in the mouse Df1 Es2elUfd1lHiraComtIdd Tbx1Cdcrel1 del22q11 Lindsay-Illingworth et al., 2001 Jerome et al., 2001 Merscher et al., 2001

9. Mutations of TBX1 found in VCFS patients without deletion Yagi et al., 2003 Paylor et al., 2006 Zweier et al., 2007

10. TBX1 mutations T-box F157YG319S 1250delC 1320-1342del TBX1 NLS1 H194Q L411P

11. G ene Functions Definition of developmental processes that require Tbx1. Definition of the critical time(s) when Tbx1 should carry out its function.

12. The developmental process (heart) Progenitor cells Specialized, differntiated cells (e.g. cardiac muscle cells) expansiondifferentiation Tbx1 + -

13. ProliferationDifferentiation Tbx1 Cardiac Progenitors (SHF)

14. How does it work (I) Tbx1 Target gene

15. How does it work (I) Tbx1 Target gene Activation of “target genes”

16. How does it work (II) Tbx1 X Y

17. How does it work (II) Tbx1 X Y Protein-protein interactions leading to functional modification of the proteins involved

18. Tbx1 Target gene Tbx1 X A B (e.g. Fgf8) (e.g. Smad1)

19. Tbx1-M Target gene Tbx1 X A B (e.g. Fgf8) (e.g. Smad1)

20. Tbx1 Target gene Tbx1-M X A B (e.g. Fgf8) (e.g. Smad1) Mutation in a VCFS patient

21. Tbx1?

22. How to re-establis the balance (in the mouse)? Increase Tbx1 dosage –Excess of Tbx1 is deleterious (published examples) Increase the dosage of target genes –Many genes involved (e.g. Fgf8). Intervene on developmental processes –Inhibit genes that enhance differentiation or that reduce proliferation. Reduce the sensitivity of tissues to Tbx1 dosage –?–?

23. 7 8 9 1011 Mouse Heart defects Cleft palate 12 16 20 24 2833 Humans 41 Critical times Gestation days Psychiatric disease ?

24. Future goals Exactly define the (molecular) damage from the loss of Tbx1. If and when there are irreversible damages. Identify mechanisms to “boost” the one remaining copy of Tbx1, or its target genes.

25. Funding National Institutes of Health (NIH), USA Telethon Italia European Community Italian Ministry of University and Research

26. Acknowledgments COLLABORATORS Elizabeth Lindsay-Illingworth Peter J Scambler Robert Schwartz (Baylor and IBT/TexasA&M) Houston Huansheng Xu Zhen Zhang Francesca Vitelli Li Chen Hedda Leeming Tom Huynh Gabriella Lania Gabriella Fulcoli Cinzia Caprio Luna Pane Rosa Serrentino Diego Circolo Sara Cioffi