Molecular cloning of the t complex responder genetic locus

Although mouse t haplotypes carry recessive mutations causing male sterility and embryonic lethality, they persist in wild mouse populations via male transmission ratio distortion (TRD). Genetic evidence suggests that at least five t-haplotype-encoded loci combine to cause TRD. One of these loci, called the t complex responder (Tcr), is absolutely required for any deviation from Mendelian segregation to occur. A candidate for the Tcr gene has previously been identified. Evidence that this gene represents Tcr is its localization to the appropriate genomic subregion and testis-specific expression pattern. Here, we report the molecular cloning of the region between recombinant chromosome breakpoints defining the Tcr locus. These results circumscribe Tcr to a 150- to 220-kb region of DNA, including the 22-kb candidate responder gene. This gene and two other homologs were created by large genomic duplications, each involving segments of DNA 10-fold larger than the individual genes.     

Functional analysis of a t complex responder locus transgene in mice

Transmission ratio distortion (TRD) of mouse t haplotypes occurs through the interaction of multiple distorter loci with the t complex responder (Tcr) locus. Males heterozygous for a t haplotype will transmit the t-bearing chromosome to nearly all of their offspring. This process is mediated by the production of functionally inequivalent gametes: wildtype meiotic partners of t spermatozoa are rendered functionally inactive. The Tcr locus, which is required for TRD to occur, is thought to somehow protect its host spermatid from the sperm-inactivating effects of linked distorter genes (Lyon 1984). In previous work, Tcr was mapped to a small genetic interval in t haplotypes, and a candidate gene from this region was isolated (Tcp-10b ^t). In this work, we further localize Tcr to a 40-kb region that contains the 21-kb Tcp-10b ^t gene. A cloned genomic copy of Tcp-10b ^t was used to generate transgenic mice. The transgene was bred into a variety of genetic backgrounds to test for non-Mendelian segregation. Abberrant segregation was observed in some mice carrying either a complete t haplotype or a combination of certain partial t haplotypes. These observations, coupled with those of Snyder and colleagues (in this issue), provide genetic and functional evidence that the Tcp-10b ^t gene is Tcr. However, other genotypes that were predicted to produce distortion did not. The unexpected data from a variety of crosses in this work and those of our colleagues suggest that elements to the TRD system and the Tcr locus remain to be identified.     

Transgenic rescue of the mouse t complex haplolethal locus Thl1

Chromosomal deletions can uncover haploinsufficient or imprinted regions of the genome. Previously, the haploinsufficient locus t haplolethal 1 (Thl1) was identified and localized to a 1.3-Mb region using overlapping deletions around the Sod2 and D17Leh94 loci of the mouse t complex on Chr 17. Germline chimeric mice, produced from embryonic stem (ES) cells containing radiation-induced deletions of the Thl1 locus, never produced viable deletion-bearing progeny when mated to C57BL/6J (B6) females. However, deletion-bearing offspring could be obtained by mating to females of other strains. In this article we describe a transgenic approach to narrow the critical region for Thl1. BAC clones were introduced into a deletion-bearing ES cell line and one was shown to rescue the Thl1 phenotype, reducing the critical region to 140 kb. Analysis of the gene content of this region suggests two strong Thl1 candidates, Pdcd2 and a novel SET domain-containing gene termed Tset1. A more detailed analysis using mice carrying overlapping deletions identified subregions that influence the phenotypic characteristics of Thl1 hemizygotes.     

Recombination between two mouse t haplotypes (t w12 tf) and t Lub-1) : Mapping. of the H-2 complex relative to centromere and tufted (tf) locus

A monoclonal antibody known to recognize the H-2.m3 specificity is shown to react with the class I H-2 product of t ^Lub-1 but not t ^w12 tf mice. This reagent was used to study the segregation of the H-2 complex in the progeny of t ^Lub-1 +/t ^ww12 tf females. The most straightforward interpretation of the results presented here is that these t haplotypes carry an H-2 complex located between the centromere and tufted locus. Possible consequences of such a location with regard to the recombination between t haplotypes and chromosome 17 from laboratory mice are discussed.     

An answer to a complex problem: cloning the mouse t-complex responder

The t-complex is maintained in wild mouse populations by its high transmission (up to 99%) from heterozygous males and provides an example of ``meiotic drive'. Its molecular basis has remained obscure despite long and intensive study. In a major advance, the t-complex responder gene, thought to be the key gene on which several distorters act, has now been cloned. Received: 14 April 2000 / Accepted: 4 May 2000     

Recapitulation of the ovum mutant (Om) phenotype and loss of Om locus polarity in cloned mouse embryos

The ovum mutant (Om) locus in mice affects early interactions between sperm and egg that in turn affect viability of embryos beyond the morula stage. Crosses of DDK females to males of many other inbred strains are 95% lethal around the morula stage, whereas reciprocal crosses are fully viable. Available data indicate that the early lethality is the result of an interaction between a factor in the ooplasm and the paternal genome. In this study, we examined whether this lethal interaction would likewise occur in cloned embryos produced by somatic cell nuclear transfer. We find that the Om effect is recapitulated but that the parental origin effect at the Om locus is no longer evident in cloned embryos.     

The chronic proliferative dermatitis mouse mutation (cpdm): mapping of the mutant gene locus

The chronic proliferative dermatitis (cpdm) mouse mutation was mapped to mouse Chromosome 15 using an intraspecific cross between C57BL/KaLawRij cpdm/cpdm and BALB/cJ mice. A second autosomal recessive mutation that resulted in a phenotype similar to that of cpdm arose spontaneously in a colony of OcB-3/Dem recombinant congenic mice. This new mutation was found to be allelic with cpdm. Therefore, the gene symbol for the allelic mutation is cpdm^Dem. The phenotype of these mutant mice consists of eosinophil-driven severe and progressive inflammatory changes in multiple organs including the skin and lungs.     

Tardy feathering locus (t) located on chromosome 1 in the chicken

The locus for tardy feathering (t), a recessive autosomal rate of feathering mutation, had been previously suggested to be on chromosome 1 in the chicken. Tests using both pea comb (P) and the breakpoint (B) of a chromosome translocation involving chromosome 1 (MN t(Z;l] verify that t lies on the long arm of chromosome 1, and shows linkage of 41.7 +/- 2.2 map units, with P located on the short arm.     

The mouse plasminogen locus maps to the recombination breakpoints of the t Lub2and Tt Orlpartial t haplotypes but is not at the t w73locus

The mouse plasminogen (Plg) locus maps to a region of chromosome (Chr) 17 which is inverted in the t haplotype Chromosomal variant. Here we investigate the genomic organization of the Plg locus in structurally variant forms of Chr 17; wild-type (+), t haplotype (t), and two partial t haplotypes Tt ^Orland t ^Lub2which arose by recombination between + and t chromosomes. Our analysis suggests that the t haplotype chromosomal variant contains extra, inverted copies of the Plg locus, and that a single locus is present in the wild-type variant. Changes in the Plg locus in Tt ^Orland t ^Lub2suggest that they arose by homologous recombination across elements in the Plg locus having the same orientation in the wild-type and t haplotype chromosomes. One hundred ten kb around the wild-type Plg genomic locus have been cloned and the proximal breakpoint of a deletion in the t ^Lub2chromosome has been localized to a fragment 30 kb downstream of the Plg gene. The t ^Lub2deletion has been shown to delete a gene named t ^w73that affects blastocyst implantation, a process probably requiring proteases such as plasminogen. However, the mapping of Plg relative to the t ^Lub2deletion and mRNA analysis of plasminogen in t ^w73heterozygotes suggests that Plg does not lie at the t ^w73locus.     

A haplolethal locus uncovered by deletions in the mouse T complex

Proper levels of gene expression are important for normal mammalian development. Typically, altered gene dosage caused by karyotypic abnormalities results in embryonic lethality or birth defects. Segmental aneuploidy can be compatible with life but often results in contiguous gene syndromes. The ability to manipulate the mouse genome allows the systematic exploration of regions that are affected by alterations in gene dosage. To explore the effects of segmental haploidy in the mouse t complex on chromosome 17, radiation-induced deletion complexes centered at the Sod2 and D17Leh94 loci were generated in embryonic stem (ES) cells. A small interval was identified that, when hemizygous, caused specific embryonic lethal phenotypes (exencephaly and edema) in most fetuses. The penetrance of these phenotypes was background dependent. Additionally, evidence for parent-of-origin effects was observed. This genetic approach should be useful for identifying genes that are imprinted or whose dosage is critical for normal embryonic development.