Mark L. Watson

The mouse fas-ligand gene is mutated in gld mice and is part of a TNF family gene cluster

The gene for the mouse Fas ligand was cloned and its chromosomal position determined. Fasl was tightly linked to gld (no crossovers in 567 meiotic events) on mouse chromosome 1 and closely linked with a novel member of the same TNF family of ligands, the Ox40 ligand (Ox40l, 1 crossover in 567 meiotic events). Southern blot analysis did not reveal any difference between the Fasl gene from gld and wild-type mice and levels of Fasl mRNA transcripts were similar in PMA and ionomycin induced wild-type and coisogenic gld T cells. Sequence analysis of the gld gene indicated a single amino acid change (Phe Leu) in the COOH terminal portion of this type II transmembrane protein, and COS cells transfected with Fasl cDNA from gld mice failed to induce apoptosis of Fas-expressing target cells. Thus, the data demonstrate that the gld phenotype is the result of a point mutation in the Fasl gene and that Fasl is part of a complex of ligands structurally related to TNF mapping within a small region of mouse chromosome 1.

Molecular characterization of murine and human OX40/OX40 ligand systems: Identification of a human OX40 ligand as the HTLV-1-regulated protein gp34

A ligand was cloned for murine OX40, a member of the TNF receptor family, using a T cell lymphoma cDNA library. The ligand (muOX40L) is a type II membrane protein with significant identity to human gp34 (gp34), a protein whose expression on HTLV‐1‐infected human leukemic T cells is regulated by the tax gene. The predicted structures of muOX40L and gp34 are similar to, but more compact than, those of other ligands of the TNF family. Mapping of the muOX40L gene revealed tight linkage to gld, the FasL gene, on chromosome 1. gp34 maps to a homologous region in the human genome, 1q25. cDNAs for human OX40 receptor were cloned by cross‐hybridization with muOX40, and gp34 was found to bind the expressed human receptor. Lymphoid expression of muOX40L was detected on activated T cells, with higher levels found on CD4+ rather than CD8+ cells. The cell‐bound recombinant ligands are biologically active, co‐stimulating T cell proliferation and cytokine production. Strong induction of IL‐4 secretion by muOX40L suggests that this ligand may play a role in regulating immune responses. In addition, the HTLV‐1 regulation of gp34 suggests a possible connection between virally induced pathogenesis and the OX40 system.

A linkage map of mouse Chromosome 1 using an interspecific cross segregating for the gld autoimmunity mutation

An interspecific backross was used to define a high resolution linkage map of mouse Chromosome (Chr) 1 and to analyze the segregation of the generalized lymphoproliferative disease (gld) mutation. Mice homozygous for gld have multiple features of autoimmune disease. Analysis of up to 428 progeny from the backcross [(C3H/HeJ-gld x Mus spretus)F1 x C3H/HeJ-gld] established a map that spans 77.6 cM and includes 56 markers distributed over 34 ordered genetic loci. The gld mutation was mapped to a less than 1 cM segment on distal mouse Chr 1 using 357 gld phenotype-positive backcross mice. A second backcross, between the laboratory strains C57BL/6J and SWR/J, was examined to compare recombination frequency between selected markers on mouse Chr 1. Significant differences in crossover frequency were demonstrated between the interspecific backcross and the inbred laboratory cross for the entire interval studied. Sex difference in meiotic crossover frequency was also significant in the laboratory mouse cross. Two linkage groups known to be conserved between segments of mouse Chr 1 and the long arm of human Chrs 1 and 2 where further defined and a new conserved linkage group was identified that includes markers of distal mouse Chr 1 and human Chr 1, bands q32 to q42.

A 6000 kb segment of chromosome 1 is conserved in human and mouse.

A murine linkage map generated from analyses of 428 meiotic events in an interspecific cross and pulsed field gel electrophoresis allowed examination of the genomic organization of a 6000 kb segment of mouse and human chromosome 1. Analysis of five genes within this syntenic segment of both species revealed striking conservation of gene order, intergenic distance and, to a lesser extent, CpG dinucleotides. In the mouse, meiotic crossover events were not evenly distributed; a hot spot for meiotic recombination was coincident with a CpG‐island. These studies provide a practical approach to aid physical mapping of the human genome and a model for determining the molecular principles that govern meiotic recombination. In addition, these findings demonstrate profound conservation of genomic organization over mammalian evolution.

CD1 defines conserved linkage group border between human chromosomes 1 and mouse chromosomes 1 and 3

The mouse chromosome to which Ly-38 (CD1) genes map has not been reported. Consequently, the present studies were undertaken to examine the genomic organization of CD1 on human chromosome 1 as well as to identify the Ly-38 (CD1) locus in the mouse

PHYSICAL AND GENETIC LINKAGE OF THE GENES ENCODING LY-9 AND CD48 ON MOUSE AND HUMAN CHROMOSOMES 1

By virtue of sequence similarity, the genes encoding CD2, CD48, CD58, and Ly-9 have been assigned to a distinct subset within the immunoglobulin superfamily. Previous gene mapping studies in human and mouse have suggested that CD2, CD48, and CD58 arose by gene duplication. Here we show the gene encoding Ly-9 to be located adjacent to CD48 and the Na,K-ATPase α2 subunit gene on human and mouse chromosome 1. The proximity in human and mouse genomes of the genes encoding CD2, CD58, and the Na,K-ATPase α1 subunit, and of the Ly-9, CD48, and the Na,K-ATPase α2 subunit genes may be explained by the occurence of two, successive duplication events during vertebrate evolution, and suggest that Ly-9 may also participate in adhesion reactions between T lymphocytes and accessory cells by homophilic interaction.

A linkage map of mouse chromosome 19: definition of comparative mapping relationships with human chromosomes 10 and 11 including the MEN1 locus

A linkage map of mouse Chromosome (Chr) 19 was constructed using an interspecific cross and markers defined by restriction fragment length variants. The map includes 20 markers, 9 of which had not been mapped previously in the mouse. The data further defined the relationship between genes on mouse Chr 19 and those on the long arm of human Chr 10 and the pericentric region of the long arm of human Chr 11. The comparative mapping analysis suggests that the proximal segment of mouse Chr 19 may contain the MEN1 locus and that the current study has identified additional genes that may be useful for positional cloning of this putative tumor suppressor gene.

Rapid and efficient construction of yeast artificial chromosome contigs in the mouse genome with interspersed repetitive sequence PCR (IRS-PCR): Generation of a 5-cM, >5 megabase contig on mouse Chromosome 1

We have developed a new technique for the generation of YAC contigs in the mouse genome that is based on the ability to detect overlapping clones by hybridization of shared IRS-PCR products. As a demonstration of the technique, a 5-cM, >5 megabase contig was developed on the distal half of mouse Chromosome (Chr) 1, spanning the region from Lamb2 to At3. The contig covers roughly 5% of the genetic distance of the chromosome and is comprised of more than 80 clones; 71 probes were assigned physical order on the chromosome, of which 59 were new markers generated in this study. Eight of the new probes were shown to be polymorphic between C3H/HeJ-gld and M. spretus. Three probes were mapped on a [(C3H/HeJ-gld x M. spretus) x C3H/HeJ-gld] interspecific backcross to integrate the physical map with a high-resolution genetic map of the region.

Long-range restriction site mapping of a syntenic segment conserved between human chromosome 1 and mouse chromosome 3

A linkage map determined from segregation analysis of 338 meiotic events in an interspecific mouse cross was utilized to help investigate genomic organization of a linkage group conserved between human chromosome 1p and mouse chromosome 3. Using pulsed-field gel electrophoresis, the genes encoding the lymphocyte adhesion molecule human CD2/murine Ly-37, the alpha 1-subunit of Na, K-ATPase, the beta-subunit of thyrotropin, the beta-subunit of nerve growth factor, and muscle adenylate deaminase were similarly positioned on long-range restriction maps in both species. These studies indicate that the development of detailed genetic maps using interspecific Mus crosses facilitates rapid analysis of murine genomic organization and may enable physical mapping of syntenic regions within the human genome. Moreover, the data suggest profound conservation of genomic organization during mammalian evolution.

Physical linkage of genes encoding the lymphocyte adhesion molecules CD2 and its ligand LFA-3.

The present study was undertaken to determine the physical proximity of LFA3 and CD2 on human chromosome 1. Physical mapping studies of CD2 and LFA3 were undertaken using pulsed field electrophoresis