Genetic structure of the mating-type locus of Chlamydomonas reinhardtii.

Portions of the cloned mating-type (MT) loci (mt(+) and mt(-)) of Chlamydomonas reinhardtii, defined as the approximately 1-Mb domains of linkage group VI that are under recombinational suppression, were subjected to Northern analysis to elucidate their coding capacity. The four central rearranged segments of the loci were found to contain both housekeeping genes (expressed during several life-cycle stages) and mating-related genes, while the sequences unique to mt(+) or mt(-) carried genes expressed only in the gametic or zygotic phases of the life cycle. One of these genes, Mtd1, is a candidate participant in gametic cell fusion; two others, Mta1 and Ezy2, are candidate participants in the uniparental inheritance of chloroplast DNA. The identified housekeeping genes include Pdk, encoding pyruvate dehydrogenase kinase, and GdcH, encoding glycine decarboxylase complex subunit H. Unusual genetic configurations include three genes whose sequences overlap, one gene that has inserted into the coding region of another, several genes that have been inactivated by rearrangements in the region, and genes that have undergone tandem duplication. This report extends our original conclusion that the MT locus has incurred high levels of mutational change.     

A novel RNA helicase gene tightly linked to the Triplo-lethal locus of Drosophila.

The Triplo-lethal (Tpl) locus of Drosophila is the only known locus which is lethal when present in three copies rather than the normal two. After recovering a hybrid-dysgenesis-induced mutation of Tpl we used a rapid combination of transposon tagging, chromosome microdissection and PCR to clone the P element that had transposed into the Tpl region. That P element is located within the gene for a new and unique member of the RNA helicase family. This new helicase differs from all others known by having glycine-rich repeats at both the amino and carboxyl termini. Curiously, genetic analysis shows that the P element inserted into this gene is not responsible for the Tpl mutant phenotype. We present possible explanations for these findings.     

Lyb-7, a new B cell alloantigen controlled by genes linked to the IgCH locus.

Anti-Lyb-5.1 serum contains antibodies against two different B cell surface antigenic determinants, Lyb-5.1 and Lyb-7.1, which are defined in cytotoxicity and functional tests, respectively. The antibody against Lyb-7.1 is identified by its ability to specifically inhibit in vitro primary antibody responses to TNP-Ficoll. Anti-Lyb-5.1 serum can be made monospecific for anti-Lyb-7.1 activity by absorption with spleen cells from AL/N mice which have been typed as Lyb-5.1+, Lyb-7.1-. Lyb-5.1 and Lyb-7.1 are each under control of one or one set of closely linked genes. The loci specifying Lyb-5.1 and Lby-7.1 are not linked to each other nor to M1s and H-2 loci. However, the gene controlling the expression of Lyb-7.1 is linked to the genes coding for the constant region of immunoglobulin heavy chains.     

The mating-type locus B alpha 1 of Schizophyllum commune contains a pheromone receptor gene and putative pheromone genes.

Analysis of the multispecific B alpha mating-type locus of Schizophyllum commune provided evidence that pheromones and pheromone receptors govern recognition of self versus non-self and sexual development in this homobasidiomycetous fungus. Four subclones of an 8.2 kb genomic fragment carrying B alpha 1 specificity induced B-regulated sexual morphogenesis when introduced into a strain with one of the eight compatible B alpha specificities that are known to exist in nature. One of these clones, which activated all other B alpha specificities, contains a gene termed bar1. The predicted protein product of bar1, as well as that of bar2, a homologous gene isolated from a B alpha 2 strain, has significant homology to known fungal pheromone receptor proteins in the rhodopsin-like superfamily of G protein-linked receptors. The other three active B alpha 1 clones were subcloned further to identify the minimal active element in each clone. Every active subclone contains a putative pheromone gene ending in a signal for possible isoprenylation. A message of approximately 600 bp was observed for one of these genes, bap1(1). This paper presents the first evidence for a system of multiple pheromones and pheromone receptors as a basis for multispecific mating types in a fungus.     

Mutations in a novel gene,TMIE, are associated with hearing loss linked to the DFNB6 locus

We have identified five different homozygous recessive mutations in a novel gene, TMIE (transmembrane inner ear expressed gene), in affected members of consanguineous families segregating severe-to-profound prelingual deafness, consistent with linkage to DFNB6. The mutations include an insertion, a deletion, and three missense mutations, and they indicate that loss of function of TMIE causes hearing loss in humans. TMIE encodes a protein with 156 amino acids and exhibits no significant nucleotide or deduced amino acid sequence similarity to any other gene.     

A sex recognition glycoprotein is encoded by the plus mating-type gene fus1 of Chlamydomonas reinhardtii.

Sexual fusion between plus and minus gametes of the unicellular green alga Chlamydomonas reinhardtii entails adhesion between plus-specific and minus-specific "fringe" proteins displayed on the plasma membrane of gametic mating structures. We report the identification of the gene (fus1) encoding the plus fringe glycoprotein, which resides in a unique domain of the mating-type plus (mt+) locus, and which was identified by transposon insertions in three fusion-defective mutant strains. Transformation with fus1+ restores fringe and fusion competence to these mutants and to the pseudo-plus mutant imp11 mt-, defective in minus differentiation. The fus1 gene is remarkable in lacking the codon bias found in all other nuclear genes of C. reinhardtii.     

Identification of a gene linked to the Brassica S (self-incompatibility) locus by differential display

Self-incompatibility in Brassica is controlled by a complex locus, the S locus, that includes several expressed genes. Two S locus genes, SLG and SRK, are expressed in the stigma and have been implicated in self-pollen recognition. The male component of this recognition system is also predicted to be encoded by a gene at the S locus but this gene has not been identified to date. In this study, we have used differential display to screen for polymorphic, S-locus-linked genes that are expressed in anthers. This approach has allowed the identification of a gene, named S5J, which was shown to segregate completely with the S locus. We discuss the possible role of this gene in the self-incompatibility response and evaluate the utility of differential display for the identification of genes at specific genetic loci.     

A third gene locus for tuberous sclerosis is closely linked to the phenylalanine hydroxylase gene locus

Summary Following the observation of a patient suffering from tuberous sclerosis (TSC) with a de novo reciprocal translocation t(3;12)(p26.3;q23.3), we have undertaken a linkage study in 15 TSC families using polymorphic DNA markers neighbouring the chromosome breakpoints. Significant lod scores have been obtained for markers D12S7 (z _max=2.34, =0.14) and PAH (phenylalanine hydroxylase) (z _max=4.34, =0.0). In multipoint linkage analysis, the peak lod score was 4.56 at the PAH gene locus. These data suggest the existence of a third gene locus for TSC (TSC3) on chromosome 12q22-24.1. The regions that have been found to be linked to TSC in different families map to the positions of three enzymes, phenylalanine hydroxylase (12q22-24), tyrosinase (11q14-22), and dopamine-beta-hydroxylase (9q34), all of which are involved in the conversion of phenylalanine to catecholamine neurotransmitters or melanin. Disorders of these biochemical pathways might be involved in the pathogenesis of TSC.     

The organization of genes tightly linked to the Ha locus in Aegilops tauschii, the D-genome donor to wheat.

The grain hardness locus, Ha, is located at the distal end of the short arm of chromosome 5D in wheat. Three polypeptides, puroindoline-a, puroindoline-b, and grain softness protein (GSP-1), have been identified as components of friabilin, a biochemical marker for grain softness, and the genes for these polypeptides are known to be tightly linked to the Ha locus. However, this region of the chromosome 5D has not been well characterized and the physical distance between the markers is not known. Separate lambda clones containing the puroindoline-a gene and the puroindoline-b gene have been isolated from an Aegilops tauschii (the donor of the D genome to wheat) genomic lambda library and investigated. Considerable variation appears to exist in the organization of the region upstream of the gene for puroindoline-b among species closely related to wheat. Using in situ hybridization the genes for puroindoline-a, -b, and GSP-1 were demonstrated to be physically located at the tip of the short arm of chromosome 5 of A. tauschii. Four overlapping clones were isolated from a large-insert BAC library constructed from A. tauschii and of these one contained genes for all of puroindoline-a, puroindoline-b, and GSP-1. The gene for puroindoline-a is located between the other two genes at a distance no greater than approximately 30 kb from either gene. The BAC clone containing all three known genes was used to screen a cDNA library constructed from hexaploid wheat and cDNAs that could encode novel polypeptides were isolated.     

A new cytotoxic lymphocyte-defined antigen coded by a gene closely linked to the H-3 locus

F1 complementation results indicate that a new gene, putatively controlling a minor histocompatibility antigen, is closely linked to the minor histocompatibility gene, H-3, in the fifth linkage group of chromosome 2 of the mouse. This gene controls a product that was capable of inducing as well as acting as a target for cytotoxic lymphocytes (CTL). The lytic activity of CTL developed in B10.LP-H-3D mice specific for the product of the new gene of B10 was restricted to target cells possessing H-2Db antigens. This contrasts to the H-2Kb-restricted activity of H-3.1 specific CTL.     

Identification of the minus-dominance gene ortholog in the mating-type locus of Gonium pectorale

The evolution of anisogamy/oogamy in the colonial Volvocales might have occurred in an ancestral isogamous colonial organism like Gonium pectorale. The unicellular, close relative Chlamydomonas reinhardtii has a mating-type (MT) locus harboring several mating-type-specific genes, including one involved in mating-type determination and another involved in the function of the tubular mating structure in only one of the two isogametes. In this study, as the first step in identifying the G. pectorale MT locus, we isolated from G. pectorale the ortholog of the C. reinhardtii mating-type-determining minus-dominance (CrMID) gene, which is localized only in the MT- locus. 3'- and 5'-RACE RT-PCR using degenerate primers identified a CrMID-orthologous 164-amino-acid coding gene (GpMID) containing a leucine-zipper RWP-RK domain near the C-terminal, as is the case with CrMID. Genomic Southern blot analysis showed that GpMID was coded only in the minus strain of G. pectorale. RT-PCR revealed that GpMID expression increased during nitrogen starvation. Analysis of F1 progeny suggested that GpMID and isopropylmalate dehydratase LEU1S are tightly linked, suggesting that they are harbored in a chromosomal region under recombinational suppression that is comparable to the C. reinhardtii MT locus. However, two other genes present in the C. reinhardtii MT locus are not linked to the G. pectorale LEU1S/MID, suggesting that the gene content of the volvocalean MT loci is not static over time. Inheritance of chloroplast and mitochondria genomes in G. pectorale is uniparental from the plus and minus parents, respectively, as is also the case in C. reinhardtii.     

Detection and estimation of linkage for a co-dominant structural gene locus linked to a gametophytic self-incompatibility locus

Summary The operation of gametophytic self-incompat ibility systems may lead to disturbed segregation ratios for genes at loci linked to the self-incompatibility loci. An exhaustive consideration of the types of crosses, methods of linkage estimation, progeny sizes and controls needed for accurate analysis of disturbed segregation ratios is presented. Examples of the application of these methods are included.     

Molecular identification of human Ia antigens coded for by a gene locus closely linked toHLA-DR locus

Human Ia(-like) specificities controlled by gene loci other thanHLA-DR were searched for at the molecular level in cells of human B-cell-type cell lines which carrytwo established DR specificities. Chevalier cells of DRw3 and 7 and U698M cells of DRw2 and 4 were used. Their Ia molecules were partially purified, radioiodinated and analyzed for Ia specificities by the direct binding and sequential binding assays with a selected panel of human Ia alloantisera. It was possible in both the cell lines to define a third subset of Ia molecules carrying a new specificity in addition to two Ia subsets carrying the established DR specificities. The new specificity was detected by putative anti-DRw4 and anti-DRw7 antisera and was closely associated with DRw4 and DRw7 at population level. It was thus designated provisionally as BR4X7. These results suggest that the BR4X7 specificity is coded for by a separateIa locus closely linked toHLA-DR locus. The determinant(s) responsible for BR4X7 was located on the small subunit of Ia molecules.     

A new cytotoxic lymphocyte-defined antigen coded by a gene closely linked to theH-3 locus

F₁ complementation results indicate that a new gene, putatively controlling a minor histocompatibility antigen, is closely linked to the minor histocompatibility gene,H-3, in the fifth linkage group of chromosome 2 of the mouse. This gene controls a product that was capable of inducing as well as acting as a target for cytotoxic lymphocytes (CTL). The lytic activity of CTL developed in B10.LP-H-3^b mice specific for the product of the new gene of B10 was restricted to target cells possessing H-2D^b antigens. This contrasts to the H-2K^b-restricted activity of H-3.1 specific CTL.     

Alkaline induction of a novel gene locus, alx, in Escherichia coli.

A novel pH-regulated locus inducible over 100-fold in alkaline media was identified in Escherichia coli through screening of 93,000 Mu dI1734 (lacZ Kmr) operon fusions at pH 6.5 and pH 8.5. Four lacZ fusions that showed expression only at the higher pH were mapped at 67.5 min by P1 transduction crosses. The locus was designated alx.