Is it Williams Syndrome ? GTF 2 I implicated in Sociability and GTF 2 IRD 1 in Visual-Spatial Construction Revealed by High Resolution Arrays

Spatial Construction Revealed by High Resolution Arrays L. Dai1, U. Bellugi2, X.-N. Chen1, A.M. Pulst-Korenberg3, A. Järvinen-Pasley2, T. Tirosh-Wagner1, P. S. Eis4, A. L. Reiss5; D. Mills6, A.F. Simon1, Y. Searcy2, J.R. Korenberg1 1Center for Integrated Neurosciences and Human Behavior, The Brain Institute, University of Utah, UT, 84108, USA; 2The Salk Institute for Biological Studies, La Jolla, CA, 92037, USA; 3Yale University, CT, 06511, USA; 4Roche NimbleGen, Inc. WI, 53711, USA; 5School of Medicine, Stanford University, CA, 94305, USA; 6Department of Psychology, Emory University, GA, 30322, USA


Introduction
Williams syndrome (WS) provides a remarkable model for studying human cognition because a hemizygous deletion of merely ~20 genes in chromosome band 7q11.23 is responsible for the unusual array of characteristics present in WS.This unique circumstance enables us to elucidate pathways that lead from genes to behavior; however, the contribution of each gene to the WS phenotype remains unknown.The distinctive pattern of physical, cognitive, and behavioral traits shown in WS is one facet that allows the parsing of the WS profile and so careful comparisons of differences between individuals with WS.Coupled with better methods that sharpen breakpoint analyses of individuals with atypical deletions, we have recently seen the dawn of drawing conclusions about specific genes and the phenotypes for which they are responsible.In this poster, we aim to draw another such conclusion, this time in regards to the connection between GTF2I and WS social behavior

Goal
To understand the neural pathways underlying social behavior in WS, to identify genes responsible for the variability and to evaluate primate models.Studies such as this with atypical deletions help to dissect and reveal distinctive behaviors.These will allow us to infer anatomic and physiologic interconnections and to understand the molecular basis of human cognition and behavior.

II. Psychometric testing
The physical development of 5889 follows typical WS patients for her age; however, her language abilities were in the normal range for age, her visual-spatial performance relatively high and she was not indiscriminantly hypersocial with strange adults, which is extremely atypical of WS individuals.
Consequently subject 5889 is unlike individuals with WS (and putatively typical deletions) who exhibit delay in but later, relative strengths in language combined with weaknesses in aspects of visual spatial processing, as well as a typical idiosyncratic "hypersocial" behavior and increased interest in non-familiar individuals.

III. Southern blotting and PCR
Procedures for DNA isolation and digestion, agarose gel construction, Southern Blotting, probe labeling, hybridization, and autoradiogram development were conducted as described by Korenberg et al. (1989).All probes were isolated as DNA fragments.The blots were hybridized with PCR fragments generated from large scale sequence analyses at 5 kb intervals throughout the cosmid 183E01.Densitometric analyses were performed by using the Alpha Innotech density measurement program (Alpha Innotech Corporation, San Leandro, CA).PCR was preformed using the DNAs extracted from the hybrid cell line with a total of 39 pairs of customer designed primers covering the region in between TFII-IRD1 and TFII-I.

Development of hierarchical high resolution genetic
approaches reveal and validate an infant with WS and a small deletion.

Longitudinal psychometric and clinical analysis reveals visuospatial function above typical WS and social approach
behavior more in the range of normal.

III. Molecular analysis
Breakpoint was determined using high resolution oligonucleotide arrays, confirmation by multicolor fluorescence hybridization with a panel of 45 BACs, somatic cell hybrids, and quantitative gene expression in LB cell lines using qRT-PCR of 12 WS genes.

V. Psychometric Testing
All subjects underwent a battery of neurocognitive measures over a 3-day period.Both standardized and experimental measures were used to assess four neurocognitive domains: general intelligence, language, visuospatial, and face processing.Measures included the Preschool Language Scale, 3rd edition, the Salk Institute Sociability Questionnaire (SISQ), and the ethogram described below.Individual scores from patient 5889 were compared to those of the full deletion WS group (n = 101) by conversion to z-scores with a mean of zero and standard deviation of one.Z-scores of 2.00 or higher were considered to be statistically significant.

Gene expression level analysis
We showed that 14 WS genes have decreased expression in the WS population relative to normal controls (NC) by quantitative RT-PCR of lymphoblast mRNA (Collette, et al., 2007).

GENETICS
Probe: STS3 of General Transcription Factor 2 (GTF2I), with diverse digestion enzyme.All lanes are as indicated.Arrow shows an extra band present in the patient's sample (STS3 break region).A: PstI, B: HindIII, C: PvuII.arrays and oligonucleotide arrays.These are symbolized at the right.On the left is shown the approach for oligonucleotide array generation using maskless sequencing.I. Oligonucleotide Isothermal High Resolution DNA tiling array 1. Isothermal Oligonucleotide array