The left and right hemispheres of the human cerebral cortex are functionally and anatomically distinct. Although recent work has identified genes whose expression underlies the rostral-caudal segmentation of the nervous system, no gene (s) that contributes to the lateralization of the cerebral hemispheres has been identified. Since the development of cerebral asymmetry (laterality) underlies language and other uniquely human cognitive abilities, and its disruption has been implicated in a number of pathologic conditions, the identification of such genes is of great importance to the clinical and basic neurosciences. The experiments outlined in this proposal seek to identify and characterize genes that are asymmetrically expressed in the developing human left and right cerebral hemispheres. Since no such gene has been previously identified, this work has the potential to open an entirely new avenue of neurobiological research. The preliminary results are encouraging, 2 candidate asymmetrically expressed genes have been identified thusfar. Representational Difference Analysis (RDA) has been used to subtract the right and left posterior perisylvian regions from each other during fetal development, proximal to the emergence of gross cerebral asymmetry. Analysis of approximately 1600 RDA subtraction products using cDNA microarray technology and a 2-color fluorescent detection system has resulted in the identification of 198 candidate clones for asymmetrical expression. Partial sequencing identified a subset of genes for further characterization, some of which share intriguing homologies with genes involved in visceral asymmetry or nervous system pattern formation. Further secondary screening using RT-PCR or Northern blotting has confirmed the differential expression of 2of these genes: one that is enriched on the right and the other that is enriched on the left. Many of clones identified in this initial screen appear to represent novel genes, about 25 percent of which have no EST (expressed sequence tag) matches in the nucleic acid and protein databases. This and other data suggest that the utility of further microarray screening of this RDA is far from saturated. Therefore, cDNA microarrays will be used to evaluate an additional 7680 RDA subtraction clones for asymmetric expression, followed by a second stage screening using the step-wise approach that we have used to identify the 2 differentially expressed clones mentioned above. Once these clones are identified, full-length cDNAs will be obtained, and the developmental expression pattern of several of these genes will be studied in the nervous system and in non- nervous system tissues, using a combination of techniques including Northern blotting, in situ hybridization and immunohistochemistry. In addition, the evolutionary conservation of asymmetrically expressed genes in non-human species will be investigated. The availability of molecular markers correlated with the development of cerebral asymmetry will greatly serve the study of the development and evolution of cognitive functions such as language, and disorders that disrupt them.
