Characterization of candidate disease genes from human chromosomes 11g13 and 22q

Sammanfattning: Two regions of the human genome, 11q13 and 22q, have been shown to harbor numerous disease-related genes, among them tumor suppressor genes (TSGs). The aim of this project was to construct transcription maps within these chromosomal segments, and thus provide a basis for further analysis of some of the genes for their involvement in disease-related processes. The main strategy used in identification of candidate genes was large-scale genomic sequencing combined with "in silico" cloning. In total 27 genes were cloned, including human paralogs and orthologs in other organisms. The following diseases have been assigned to 11q13: Bardet-Biedl syndrome; spino-cerebellar ataxia type 5; familial paraganglioma type 2; insulin dependent diabetes mellitus; the translocation t(11;17) in B-cell non-Hodgkin's lymphoma; and multiple endocrine neo-plasia type 1 (MEN 1). Our aim was to identify genes in the vicinity of PYGM and FAU loci using a comparative genomic sequencing approach. We sequenced a PAC around the PYGM locus from human 11q13 and a cosmid from Fugu rubripes containing the Pygm gene. The region surrounding Pygm locus in F. Rubripes was not syntenic with the region neighboring the human PYGM gene. We established a transcriptional map around the human PYGM gene which includes: i) two genes previously localized to 11q13, PYGM and a zinc-finger protein (ZFM1) gene; ii) germinal center kinase (GCK) gene; iii) a novel human CDC25-like (HCDC25L) gene; iv) a dystrophia myotonica protein kinase-like (DMPKL) gene; and v) a novel ubiquitously expressed gene of unknown function (germinal center kinase neighboring gene, GCKNG). The GCKNG gene was also cloned by other groups and confirmed to be the MEN1 tumor suppressor gene. Deletions on 22q have been described in multiple tumors, which implies that this chromosome harbors TSGs. Previous deletion mapping studies in meningiomas have defined four candidate regions on 22q. From one of these the neurofibromatosis type 2 gene (NF2) was cloned and shown to be inactivated in a subset of meningiomas. However, further studies indicate that additional 22q-located gene(s) may be important in development of these and other tumors. We analyzed six genes from meningioma-deleted regions on 22q. These were: clathrin heavy chain 2 gene (CLH-22); beta-prime adaptin gene (BAM22); human homologue of C. elegans NIPSNAP gene 1 (NIPSNAP1); synaptogyrin 1 gene (SYNGR1a, SYNGR1b, SYNGR1c); human homolog of chicken TOM1 gene (TOM1); and SWI/SNF-related, matrix-associated, actin-dependent regulator of chromatin, subfamily B, member 1 (SMARCB1a, SMARCB1b). For the majority of these human genes, human paralogs and mouse orthologs were also investigated. This resulted in the characterization of a novel family of genes called synaptogyrins (SYNGR1-4), as well as NIPSNAP2 and TOM1L genes in humans. The human SMARCB1 gene is a tumor suppressor gene located on 22q11.2 and it is inactivated in malignant rhabdoid tumors. We performed mutational analysis of the SMARCB1 gene as well as CLH22, BAM22, and TOM1 genes in meningiomas and schwannomas with negative results. The closest paralogs of three genes (CLH-22, BAM22, and SYNGR1) studied in this thesis are located on human chromosome 17. Furthermore, the available literature and searches of databases show that 13 pairs of paralogs are located on chromosome 17 and 22. We propose that this is a resuit of an ancient chromosomal duplication event.