Sahota, Virender Kumar; (1998) Molecular genetic studies of human and mouse cell adhesion genes: The desmosomal cadherin locus on chromosome 18. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

The desmosome cell junction has two types of associated transmembrane glycoproteins belonging to the cadherin superfamily of calcium dependent cell adhesion molecules, the desmogleins (DSG) and the desmocollins (DSC). Each is present as three isoforms encoded by separate genes. The expression patterns of the type 1 and type 3 isoforms are restricted to specific layers within the epidermis and the position of the genes on the chromosome appears to be related to the spatial order of expression during morphogenesis. DSG1 and DSG3 are the autoantigens involved in the blistering skin diseases pemphigus foliaceous and pemphigus vulgaris respectively. The inherited skin disease striated palmoplantar keratoderma has been mapped to the same region of chromosome 18, and since the DSC and DSG genes are expressed in the epidermis, they are good candidates for this disease. The six genes are tightly clustered at 18q12.1 and previous work in our lab has shown that they are all present on three overlapping Yeast Artificial Chromosome (YAC) clones spanning no more than 700 kb. The locus consists of two gene clusters, one contains the DSG genes, the other the DSC genes, with transcription proceeding outwards from the interlocus region. This close linkage may perhaps indicate that the expression of these genes is co-ordinately controlled by common regulatory components. I have screened cosmid and P1-derived Artificial Chromosome (PAC) libraries which are considered more stable and less prone to rearrangements than YACs. STS markers have been generated using vectorette PCR which were then mapped to the PAC and cosmid clones thereby constructing a complete bacterial clone contig. In addition, some of the PAC clones were sequenced directly using automated sequencers. This mapping has confirmed the order of the genes and has also increased the resolution of the map to provide a sequence ready contig, enabling us to pursue an intensive study of the control of expression of the whole locus. The DSC gene is the earliest desmocollin gene to be expressed in the developing mouse embryo, being present from the 16-cell stage. In order to study the function, I have also generated a targeting construct with the aim of ablating the function of the DSC2 gene by homologous recombination in embryonic stem (ES) cells. The vector has been tested and the neomycin gene is functioning correctly. Several clones have already been picked and screening is in progress. The order of the desmosomal cadherin genes in mouse have not yet been determined, although it is expected that the order in mouse will reflect the order in humans. I have screened a mouse YAC library by PCR and isolated a clone that contains the DSC2 gene.

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