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Proviral copies of the mouse leukemia viruses behave as classical Mendelian loci and as mobile genetic elements. As mobile elements they can reinsert in somatic and germ cells; these insertions may lead to cancer or heritable mutations. The mobility of these proviral elements results in great diversity among mice in the number and chromosomal position of proviral genes. We discuss factors which may influenced the rate of acquisition of novel proviruses in the germ line.

The gene coding for the ethanol-inducible, developmentally regulated P450IIE1 was isolated from an lambda EMBL 3 rat genomic library and completely sequenced. The gene spanned 10,373 base pairs and contained nine exons. Upstream and downstream DNA of 1530 and 825 base pairs, respectively, was also sequenced, and the transcription start site was identified by both S1 mapping and primer extension. A typical TATA box was found just upstream of the start site; however, no CCAAT box was apparent. Other repetitive sequences were identified including a partial R.dre.1 sequence upstream of the gene and a long stretch of 160 alternating purines in the second intron. This latter repetitive element is found in many mammalian genes. By use of a panel of mouse-hamster somatic cell hybrids, the P450IIE1 gene was localized to mouse chromosome 7. The hepatic P450IIE1 gene is transcriptionally activated within 1 day after birth and reaches a maximal level of expression at 6 days of age. Using restriction endonuclease sites generated from the gene sequence data and the cytosine methylation-sensitive enzymes HhaI and HpaII, we found that this transcriptional activation during early development is coincident with specific demethylation only at the 5' end of the P450IIE1 gene. Interestingly, other cytosine residues in the middle of the gene became demethylated as rats aged from 1 to 10 weeks, at which time no changes in gene expression occur.

HoMuLV is an NB-tropic wild mouse leukemia virus (MuLV) with ecotropic host range which induces lymphoma, erythroleukemia, and myelogenous leukemia in NIH Swiss mice. Although HoMuLV uses the same cell surface receptor as other ecotropic MuLVs, hybridization studies suggested that the HoMuLV envelope glycoprotein differs from that of other ecotropic MuLVs. We have now molecularly cloned HoMuLV and sequenced its LTR, gag , and env regions. HoMuLV differs markedly from other MuLVs in the LTR U3 region, in the SU protein of env , and in p12 gag . U3 contains a single copy of a sequence analogous to the direct repeat found in other LTRs, and this region includes several previously defined protein binding sites. The predicted amino acid sequence for the coding regions of env and gag reveal that p12 and the SU protein show less than 59 and 65% sequence identity, respectively, with those of other MuLVs. A 0.6-kb segment of the 5′ region of the HoMuLV env was used as a hybridization probe to examine inbred and wild mouse genomic DNAs for proviral sequences. HoMuLV env sequences were not present in the germline of any of the inbred strains or wild mice examined including the Mus hortulanus mice which harbored infectious virus. Therefore, HoMuLV represents an evolutionarily related, but distinct, subgroup of ecotropic MuLV which is not genetically transmitted in its natural host.

The gene coding for the M r 26000 γ chain of the human CD3 (T3) antigen/T-cell antigen receptor complex was mapped to chromosome band 11q23 by using a cDNA clone (pJ6T3 γ-2), by in situ hybridization to metaphase chromosomes and by Southern blot analysis of a panel of human-rodent somatic cell hybrids. The mouse homolog, here termed Cdg-3, was mapped to chromosome 9 using the mouse γ cDNA clone pB10.AT3 γ-1 and a panel of mouse-hamster somatic cell hybrids. Similar locations for the CD3 δ genes have been described previously. Thus, the corporate results indicate that the CD3 γ and δ genes have remained together since they duplicated about 200 million years ago.

The mammalian protooncogene homologue of the avian v-ets sequence from the E26 retrovirus consists of two sequentially distinct domains located on different chromosomes. Using somatic cell hybrid panels, we have mapped the mammalian homologue of the 5' v-ets-domain to chromosome 11 (ETS1) in man, to chromosome 9 (Ets-1) in mouse, and to chromosome D1 (ETS1) in the domestic cat. The mammalian homologue of the 3' v-ets domain was similarly mapped to human chromosome 21 (ETS2), to mouse chromosome 16 (Ets-2), and to feline chromosome C2 (ETS2). Both protooncogenes fell in syntenic groups of homologous linked loci that were conserved among the three species. The occurrence of two distinct functional protooncogenes and their conservation of linkage positions in the three mammalian orders indicate that these two genes have been separate since before the evolutionary divergence of mammals.

We derived an amphotropic murine leukemia virus (MuLV) type-specific probe for use in Southern blot hybridizations with cloned and genomic DNAs. A 133-base-pair RsaI-RsaI fragment from the 5' env region of the amphotropic viral isolate 4070A was subcloned into M13mp18 and radiolabeled in vitro. The probe detected the proviral DNAs in mink cells infected with seven different amphotropic MuLV isolates. The probe did not cross hybridize with the DNAs of molecular clones of ecotropic, mink cell focus-forming, or xenotropic MuLVs; nor did it anneal to the proviral DNAs of four xenotropic or six mink cell focus-forming viral isolates grown in mink cells. DNAs of 12 inbred laboratory mouse strains and more than 15 different wild mouse species and subspecies were examined for the presence of endogenous amphotropic env-related fragments. Amphotropic env-related sequences were found only in the DNAs of wild mice trapped in southern California in an area previously shown to harbor mice producing infectious amphotropic virus. Restriction enzyme analyses of DNAs from these mice showed that amphotropic sequences were not present as germ line copies but were the result of congenital or horizontal infection or both in this population. The DNAs of 11 various mammalian and avian species, including both natural predators of mice and squabs from the farms with virus-positive mice, lacked amphotropic envelope-related sequences.

cDNA clones encoding proteolipid protein (PLP) were isolated from a mouse brain library and sequenced. We describe two transcripts arising from the PLP locus by alternative splicing: the major one encodes the 277-amino acid PLP protein and the minor one corresponds to the DM-20 protein, a PLP-like protein of 20,000 Mr that shares both amino and carboxyl regions with PLP. These two transcripts lack approximately 70 bases in PLP mRNA from the dysmyelinating jimpy mutant. The deletion spans amino acids 208-232; however, this region is present in the jimpy PLP-encoding gene. We propose that the jimpy mutant suffers a point mutation or the deletion of a few bases in the PLP gene that alters the normal splicing pattern and generates partially deleted PLP transcripts.

A single mendelian gene was identified for induction of the endogenous xenotropic murine leukemia virus in five mouse strains (C57BL/10, C57L, C57BR, AKR, and BALB/c). This locus, designated Bxv-1, mapped to the same site on chromosome 1 in all strains: Id-1-Pep-3-[Bxv-1-Lp]. Thus, inducibility loci for xenotropic virus are more limited in number and chromosomal distribution than ecotropic inducibility loci. Virus expression in mice with Bxv-1 was induced by treatment of fibroblasts with 5-iododeoxyuridine or by exposure of spleen cells to a B cell mitogen, bacterial lipopolysaccharide. An analysis of the hamster X mouse somatic cell hybrids indicated that chromosome 1, alone, was sufficient for virus induction.

We used hybridization probes that react specifically with xenotropic and mink cell focus-forming virus envelope sequences to characterize the nonecotropic proviruses of BALB/c and NFS/N mice. Analysis of somatic cell hybrids with different BALB/c chromosomes showed that the 9 xenotropic and more than 20 MCF virus-related proviral sequences in this mouse were present on more than nine BALB/c chromosomes. Multiple copies were found on chromosomes 1, 4, 7, 12, and probably 11, and the copies found on a single chromosome were not identical by restriction enzyme mapping. We also identified and characterized the proviral sequences that give rise to infectious xenotropic virus in both BALB/c and NFS/N mice. BALB/c contains the major locus for induction of infectious virus in inbred mice, Bxv-1, which is on chromosome 1. We showed that this locus contains a single xenotropic provirus on an 18-kilobase HindIII fragment. Restriction enzyme analysis of a hybrid cell DNA that contains only the Bxv-1 xenotropic provirus showed that the Bxv-1 provirus contains restriction enzyme sites characteristic of the infectious virus induced from BALB/c fibroblasts. The Bxv-1 provirus and its flanking sequences also contain the same restriction sites as the provirus thought to contribute U3 long terminal repeat sequences to leukemogenic (class I) AKR MCF viruses. Analysis of cell hybrids made with the nonvirus-inducible strain NFS/N showed that the single xenotropic virus env gene of NFS mice, here termed Nfxv-1, is not on chromosome 1. Unlike that of Bxv-1, the restriction map of Nfxv-1 does not resemble that of any known infectious xenotropic virus including xenotropic viruses isolated from NFS mice. These data suggest that Bxv-1, but not Nfxv-1, is a full-length xenotropic provirus that can be transcribed directly to produce infectious virus.

A rat cDNA clone encoding a portion of phosphate-activated glutaminase was used to identify DNA restriction fragment length polymorphisms (RFLPs) in sets of somatic cell hybrids and between wild-derived and inbred strains of mice. Segregation of rat and mouse chromosomes among somatic cell hybrids indicated assignment to rat chromosome 9 and mouse chromosome 1. Analysis of chromosome 1 alleles for several genes in an interspecific cross between Mus spretus and C3H/HeJ-gld/gld mice indicates that glutaminase can be positioned within 5.5 +/- 2.0 cM proximal to Ctla-4. Similarly, human-hamster somatic cell hybrids were examined for RFLPs, and four human EcoRI restriction fragments were found to hybridize with the rat glutaminase probe. Two of these restriction fragments cosegregated and mapped to human chromosome 2 in a region that is syntenic with mouse chromosome 1 and rat chromosome 9.

A cDNA encoding human class III (χ, ADH5) alcohol dehydrogenase was isolated, sequenced and used to comparatively map this unusual ADH. In their coding sequences, the three major ADH classes were approximately equisimilar, class II and III ADHs sharing the highest sequence identity (67%). A class III-like ADH was mapped to mouse chromosome 3, site of the ADH gene complex, and synteny of ADH5 with four other ADH loci on human chromosome 4 was confirmed. The nearly full-length 1613 nucleotide cDNA contained 433 nucleotides of 3′ nontranslated sequence and two possible initiation sites for translation. A protein of 374 amino acid residues could be synthesized using the potential initiation codon at nucleotide 59. However, use of the likely initiation codon at nucleotide 5 would produce a protein of 392 residues with 19 additional N-terminal residues as compared to the known protein sequence. The derived protein sequence also differs at residue 166, where Tyr is found. This difference, due to a single base substitution, could result from cloning artifact, polymorphism, or two expressed class III ADH genes.

Congenic mouse strains NIH,Akv-1 and NIH,Akv-2 carry the two high ecotropic virus-inducing loci of the AKR mouse on the NIH Swiss genetic background. Progeny tests of animals in three separate congenic families show that both Avk-1 and Akv-2 are stably transmitted as classical mendelian loci in these mice. However, during the process of inbreeding, additional chromosomal viral loci were detected in several NIH.Akv-1 sublines. These loci appeared only in the progeny of virus-positive females. They segregate with mendelian ratios, are unlinked to markers on chromsome 7 near Akv-1, and are phenotypically expressed as high-virus-inducing loci. The generation of new loci for viurs induction, no doubt resulting from the rare germ-line reintegration of the endogenous ectropic provirus, represents a unique form of gene duplication and rearrangement.

Several strains of laboratory and wild-derived mice from Japan carry the dominant allele at the Fv-4 locus (Fv-4r) that is responsible for resistance to infection by exogenous ecotropic murine leukemia virus. We have used blot hybridization with a probe specific for the ecotropic viral envelope to show that a unique envelope-reactive sequence is present in the Japanese mouse Mus musculus molossinus and in four independently derived partially congeneic strains carrying Fv-4r. Analysis of 31 backcross mice shows complete concordance between inheritance of this fragment and resistance to Friend virus complex-induced erythroblastosis. Inheritance of this sequence also suppresses spontaneous expression of the endogenous ecotropic viruses carried by M. m. molossinus. Restriction enzyme analysis shows that the Fv-4r-associated sequence is different from the full-length ecotropic proviruses of laboratory mice. Infectious virus cannot be induced from mice carrying only the Fv-4r-associated sequence. Examination of other wild-derived mice resistant to Friend virus complex shows that Mus cervicolor cervicolor also contains the Fv-4r sequence. Our data indicate that a unique or incomplete provirus containing ecotropic envelope-related sequences is responsible for Fv-4-mediated resistance to both exogenous and endogenous ecotropic virus in various Asian mice.

The dioxin-inducible P-450 gene family in the C57BL 6N mouse comprises two genes, P 1 -450 and P 3 -450 . Restriction endonuclease-digested genomic DNA was probed with P1-450 and P3-450 full-length cDNA clones in an attempt to find species-specific fragment length differences between mouse and hamster cell lines and any restriction fragment length polymorphism among four inbred mouse strains. With this Southern blot hybridization technique, Pst I fragments were used to distinguish between the mouse and hamster P 1 -450 P 3 -450 genes, and Pvu II fragments were used to distinguish P 3 -450 differences between the AKR J and C57L J inbred strains. Analysis of nineteen mouse x hamster somatic cell hybrid lines and sixteen AKXL ( AKR J × C57L J ) recombinant inbred lines showed that the P 1 -450 P 3 -450 genes are located near the Mpi-1 locus, between the Thy-1 and Pk-3 loci, in the middle portion of mouse chromosome 9.

Comparative mapping in man and mouse has revealed frequent conservation of chromosomal segments, offering a potential approach to human disease genes via their murine homologs. Using DNA markers near the Huntington disease gene on the short arm of chromosome 4, we defined a conserved linkage group on mouse chromosome 5. Linkage analyses using recombinant inbred strains, a standard outcross, and an interspecific backcross were used to assign homologs for five human loci, D4S43, D4S62, QDPR, D4S76, and D4S80, to chromosome 5 and to determine their relationships with previously mapped markers for this autosome. The relative order of the conserved loci was preserved in a linkage group that spanned 13% recombination in the interspecific backcross analysis. The most proximal of the conserved markers on the mouse map, D4S43h, showed no recombination with Emv-1, an endogenous ecotropic virus, in 84 outcross progeny and 19 recombinant inbred strains. Hx, a dominant mutation that causes deformities in limb development, maps approximately 2 cM proximal to Emv-1. Since the human D4S43 locus is less than 1 cM proximal to HD near the telomere of chromosome 4, the murine counterpart of the HD gene might lie between Hx and Emv-1 or D4S43h. Cloning of the region between these markers could generate new probes for conserved human sequences in the vicinity of the HD gene or possibly candidates for the murine counterpart of this human disease locus.

A phage clone which contained a functional U1a1 snRNA gene was isolated from a mouse genomic library. A single copy fragment was isolated from the 3' flanking region of the U1a1 gene and used as a hybridization probe for Southern blotted DNAs from recombinant inbred strains of mice, mouse-hamster hybrid cells, and the offspring from backcrosses between BALB/c mice and mice which were heterozygous for the Rex (Re) marker. The results of these experiments prove that the U1a1 gene is located on chromosome 11 near the Delb-1 and Re loci.

SummaryThe gene coding for human p68, a membrane-associated calcium-binding protein, has been assigned to chromosome 5, using a cDNA clone to probe genomic DNA from rodent-human somatic cell hybrids by Southern hybridisation. The gene was localised, by in situ hybridisation, to 5q32–34. The murine gene was assigned to chromosome 11, using a murine cDNA clone to probe genomic DNA from rodent-rodent somatic cell hybrids.

Debrisoquine 4-hydroxylase (P450db1) was purified from rat liver microsomes. Polyclonal antibody was produced and, in conjunction with immunoblots, was used to identify and purify a second immunorelated P450 (P450db2) that does not have debrisoquine hydroxylating activity. The cDNA clones to db1 and db2 were isolated from a lambda gt11 expression library, sequenced, and found to share 78% nucleotide and 73% deduced amino acid similarities. These similarities are evenly dispersed along the sequence except for a region of 190 nucleotides with 99% similarity near the carboxyl terminus of the protein-coding region; this similarity is probably the remnant of a gene conversion event. Both proteins share between 38% and 43% amino acid similarity with P450a, P450b, P450e, P450f, P450PB1, and P450j; these data indicate that P450db1 and P450db2 are members of a separate subfamily within the P450II gene family. Southern blot analysis and preliminary genomic cloning suggest that at least four genes exist in the subfamily, although the present evidence suggests that only db1 and db2 are expressed in rat liver. With the use of 19 mouse X hamster somatic cell hybrids, the db1 and db2 genes were localized to mouse chromosome 15 (P450-2D locus). A polymorphism has been described for debrisoquine metabolism in the DA rat strain, adult females having markedly decreased debrisoquine 4-hydroxylase activity. Our immunoblot analysis and mRNA analysis suggest that debrisoquine 4-hydroxylase deficiency in the female DA rat is not due to a decrease in db1 protein or mRNA. The db1 and db2 proteins are differentially regulated: during development db2 is present at birth while db1 is absent, and db1 increases by 1 week of age; in addition, db1 is slightly induced by phenobarbital, 3-methyl-cholanthrene, and dexamethasone whereas db2 is marginally increased by these latter two agents. These results demonstrate that debrisoquine 4-hydroxylase is a member of a new constitutively expressed P450II sub-family containing two or more genes in the rat and establish that the debrisoquine polymorphism in the DA rat is probably due to a structurally altered db1 protein.

A cDNA was isolated from rat brain by low stringency hybridization with the rat mu opioid receptor cDNA. Sequence analysis of this clone indicated that it contains an open reading frame capable of encoding a 367 amino acid protein. The deduced amino acid sequence of this protein shows high degrees of homology to all three opioid receptors, mu, kappa, and delta, suggesting that it is a member of the opioid receptor gene family. RNA blot analysis detected high level expression of the receptor mRNA in the brain. Southern blot analysis suggests that it is a single-copy gene, and mapping studies localized the gene on mouse chromosome 2. Despite the high sequence homologies between this protein and the other opioid receptors, expression studies of this clone in COS-7 cells did not show binding to [3H]diprenorphine, a ligand that binds to the other three opioid receptors. Furthermore, co-expression of this receptor with a G protein-activated potassium channel in Xenopus oocytes did not show functional coupling upon stimulation with mu, kappa and delta agonists. Given the similar degrees of high homology to the mu, kappa and delta opioid receptors and the lack of apparent affinity for their ligands, this receptor does not appear to belong to any of the three known classes of opioid receptors. Rather, it represents a novel member of the opioid receptor gene family, not identified from previous pharmacological studies.

Mice carrying the autosomal recessive rd gene experience total degeneration of the photoreceptor cells of the retina by 3 to 4 weeks of life. Biochemical studies of the rd retina have demonstrated a lesion in cyclic guanosine monophosphate (cGMP) metabolism due to depressed rod-specific cGMP-phosphodiesterase (cGMP-PDE) activity. The depressed activity could result from, among other things, a lesion in the cGMP-PDE enzyme itself or in any of a number of proteins in the rod that regulate it. We have used a cDNA clone for the alpha-subunit of bovine rod transducin (T alpha 1) to map the corresponding gene, Gnat-1, to mouse chromosome 9 with a panel of Chinese hamster-mouse somatic cell hybrid DNAs. Transducin, a heterotrimeric G protein, is involved in the stimulation of cGMP-PDE when light hits the rod photoreceptors. Since the primary defect in rd disease occurs in a gene(s) on mouse chromosome 5, our results suggest that Gnat-1 is not the rd gene.

By means of an approach that combined the techniques of somatic cell genetics and Mendelian breeding studies, the inducibility locus, designated Cv, for ecotropic murine leukemia virus in BALB/c mice, was mapped to chromosome 5, 23 units from the locus for phosphoglucomutase-1, with gene order Cv-Pgm-1-Gus. This low-efficiency inducibility locus is therefore not allelic with the chromosome 7 loci previously described for two other mouse strains with high virus inducibility. These studies provide further evidence that endogenous ecotropic viruses represent viral genomes inserted at different chromosomal sites in the various mouse strains.

When mouse L cells are incubated with 32P-labeled recombinant murine immune interferon ( [32P]Mu-IFN-gamma) and subsequently cross-linked with disuccinimidyl suberate, a major complex with an apparent molecular mass of 95,000-125,000 daltons can be visualized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The complex was not formed when the binding was performed in the presence of excess unlabeled Mu-IFN-gamma or when Chinese hamster ovary cells were used. This complex therefore represents the Mu-IFN-gamma receptor (or its interferon-binding subunit). The chromosomal location of the Mu-IFN-gamma receptor (or the binding subunit of the receptor) gene, termed Ifgr, was identified by performing the binding and cross-linking reactions on a series of mouse-hamster somatic cell hybrids with different subsets of mouse chromosomes. The presence of mouse chromosome 10 was shown to be necessary and sufficient for the formation of the cross-linked complex. Thus, the gene coding for the binding subunit of the Mu-IFN-gamma receptor was localized to mouse chromosome 10. The presence of this chromosome in the hybrid cells was not sufficient, however, to confer antiviral resistance to the hybrids when they were treated with Mu-IFN-gamma and challenged with encephalomyocarditis virus.

The virally transduced oncogene v-cbl transforms fibroblasts in vitro and induces early B-cell-lineage lymphomas in vivo. A series of probes derived from a molecular clone of v-cbl were used to map related sequences in the mouse genome. Analyses of Chinese hamster x mouse somatic-cell hybrids showed that two related genes, cbl-1 and cbl-2, were located on chromosomes 6 and 9, respectively. Restriction enzyme studies of DNA from hybrid cells containing either chromosome 6 or 9 suggested that cbl-1 resembles v-cbl and may be a processed gene, whereas cbl-2 has a complex genomic structure. Analyses of Mus domesticus/M. spretus interspecific backcross mice showed that Cbl-1 maps between the immunoglobulin kappa light chain and T-cell receptor beta chain loci and that Cbl-2 is tightly linked to Thy-1.

Publisher Summary The mouse retroviruses are RNA-containing viruses that replicate through a DNA intermediate. This chapter describes the four major classes of endogenous mouse retroviruses—namely, the type C mouse leukemia viruses (MuLVs), the type B mouse mammary tumor viruses (MMTVs), the intracisternal A particles (IAPs), and the virus-like endogenous sequences (VL30s). It also describes the arrangement and stability of these sequences in the mouse germline, the experimental evidence linking specific germline integrations with various polymorphic chromosomal loci, and the association of specific somatically acquired proviruses with tumor induction and progression. The mouse retroviruses are essentially classified into four groups based on differences in virion morphology— MuLVs, MMTVs, IAPs, and VL30s. The virions have type A, B, or C structure as revealed by electron microscopy. MuLVs and MMTVs are replication-competent viruses with type C and type B structures, respectively. IAPs resemble immature type B virions, but are noninfectious and are found only intracellularly. VL30s are endogenous retrovirus-like sequences that are not known to produce any virion structural components, but can be efficiently packaged and transmitted as pseudotypes of type C viruses.

Restriction fragment length variants have been exploited to map genetically Rb-1, the murine homolog of the human retinoblastoma gene. Rb-1 localized to mouse chromosome 14 on the basis of results from analysis of somatic cell hybrids. In an interspecific backcross involving Mus spretus, Rb-1 and the murine homolog of the human esterase D gene (ESD), which we refer to here as Esd, were inseparable. Furthermore, the strain distribution patterns of Rb-1 and Es-10 are the same in 31 of 32 recombinant inbred strains. Close linkage of the chromosome 14 morphological marker hairless (hr) to Rb-1 is also implied. These results localize Rb-1 on the mouse linkage map and provide close genetic markers to follow Rb-1 in somatic as well as in germline genetic experiments. Additionally, the results suggest that Es-10 is the murine homolog of ESD and provide further evidence for linkage conservation during mammalian evolution.

The mouse homologs of the cellular oncogenes c-Ha-ras-1 of Harvey sarcoma virus and c-fes of feline sarcoma virus were both mapped to chromosome 7 by Southern blot analysis of hamster-mouse somatic cell hybrid DNAs.

Spermatid transition protein l (TP1) is a 54 amino acid (aa), highly basic chromosomal protein found in mammals during the brief period when histones are being replaced by protamines in the haploid phase of spermatogenesis. Using a cDNA clone as probe, we have isolated the gene ( Stp-1 ) coding for rat TP1 from a population of recombinant bacteriophage λ. The nucleotide (nt) sequence was established from a point 126 nt upstream from the mRNA cap site to a point about 30 nt downstream from the predicted site of polyadenylation. The gene contains a single intron separating the codon for aa 45 of the mature protein. Comparison of the nucleotide sequences for Stp -1 and the mouse gene coding for protamine P1 suggests a possible evolutionary relationship. Southern blot hybridization to genomic DNA isolated from a panel of mouse-hamster somatic cell hybrids unambiguously mapped Stp -1 to mouse chromosome 1.

We have previously identified a region of genomic DNA which constitutes the site of frequent provirus integration in rat thymomas induced by Moloney murine leukemia virus (Lemay and Jolicoeur, Proc. Natl. Acad. Sci. USA 81:38-42, 1984). This genetic locus is now designated Mis-1 (Moloney integration site). Cellular sequences homologous to Mis-1 are present in mouse DNA. Using a series of hamster-mouse somatic cell hybrids, we mapped the Mis-1 locus to mouse chromosome 15. Frequent chromosome 15 aberrations have been described in mouse thymomas. Mis-1 represents a putative new oncogene which might be involved in the initiation or maintenance or both of these neoplasms.

A novel P450 cDNA was isolated from a rat lung lambda gt11 library by hybridization with the rat P450 IIB1 cDNA probe. The cDNA-deduced amino acid sequence of this clone was 71% and 73% similar to rat IIA1 and IIA2 P450s; it was, therefore, designated IIA3 as the third member of the rat IIA subfamily. IIA3 demonstrates only 55% amino acid similarity with IIB1. Interestingly, this P450 also shared 85% and 94% amino acid similarities with human IIA3 and a mouse testosterone 15 alpha-hydroxylase P450, respectively, indicating that these P450s are orthologous counterparts to rat IIA3. Chromosome mapping, using mouse-hamster somatic cell hybrids, revealed that the IIA3 gene is localized on mouse chromosome 7. The IIA3 mRNA was detected in rat lung, and its level was induced 3-fold by treatment of rats with 3-methylcholanthrene. No IIA3 mRNA was seen in the liver, kidney, or intestine, even after long exposure of Northern blot filters to X-ray film. In contrast, the orthologous mouse and human IIA3 genes are expressed in liver.

The chromosomal localization of the gene for UDP-galactosyltransferase (glycoprotein 4-B-galactosyltransferase, EC 2.4.1.38) has been determined to be on mouse chromosome 4 by the use of mouse × hamster somatic cell hybrids. It has been proposed that galactosyltransferase is associated with the mouse T/t complex which has been localized to mouse chromosome 17. These results show that galactosyltransferase is not encoded within the T/t complex.

Mus musculus subsp. musculus (Czech II) mammary tumor DNA frequently contains an integrated proviral genome of the mouse mammary tumor virus (MMTV) within a specific 0.5-kilobase-pair region of the cellular genome (designated int-3). Viral integration at this site results in activation of expression of an adjacent cellular gene. We mapped int-3 to mouse chromosome 17 by analysis of PstI-restricted cellular DNAs from mouse-hamster somatic cell hybrids. Restriction analysis of cellular DNA from (C3H/OuJ X Czech II) X Czech II backcross mice established the gene order T-H-2-int-3. These results demonstrated that the int-3 locus is distinct from two other common integration regions for mouse mammary tumor virus (designated int-1 and int-2) in mammary tumor DNA and suggest that several cellular genes may be at risk for virally induced activation during mammary tumor development.

A rat thyroid peroxidase cDNA has been isolated from a FRTL-5 thyroid cell library and sequenced. The cDNA is 2776 base pairs long with an open reading frame of 770 amino acids. By comparison to full-length human thyroid peroxidase cDNA and based on its identification of a 3.2 kilobase mRNA in rat thyroid FRTL-5 cell Northern blots, the rat peroxidase cDNA appears to lack 400-500 base pairs at the 5'-end of the mRNA. It exhibits only a 74% nucleotide and 77% amino acid sequence similarity to human thyroid peroxidase cDNA within the total aligned sequences, although the predicted active site regions are highly conserved (greater than 90-100%). The cDNA has been used to map the thyroid peroxidase gene in mice to chromosome 12 and to compare thyroid peroxidase and thyroglobulin gene expression in FRTL-5 rat thyroid cells. Despite the fact TSH action in both cases is duplicated, and presumably mediated, by cAMP, TSH-induced increases in thyroid peroxidase and thyroglobulin mRNA levels differ. Differences exist with respect to hormone concentration and time. The ability of TSH to increase thyroglobulin, but not thyroid peroxidase mRNA levels, requires insulin, 5% serum, or insulin-like growth factor-I. Insulin or insulin-like growth factor-I alone can increase thyroglobulin mRNA levels as well as or better than TSH but have only a small effect on thyroid peroxidase mRNA levels by comparison to TSH. The ability of TSH to increase thyroglobulin gene expression is readily detected in nuclear run-on assays but not the ability of TSH to increase thyroid peroxidase gene expression. Cycloheximide inhibits TSH-increased thyroglobulin but not peroxidase mRNA levels. Finally, methimazole and phorbol 12-myristate 13-acetate show different effects on TSH-induced increases in thyroglobulin and thyroid peroxidase mRNA levels.

The structural gene for the beta-subunit of the mouse erythrocyte spectrin, hereinafter designated as Sp-b, was assigned to the mouse chromosome 12. This assignment was made by Southern analysis of genomic DNA from mouse X Chinese hamster hybrid cells using cloned mouse erythrocyte beta-spectrin cDNA as a probe. In the PstI-digested genomic hamster cell DNA a single band of 2.0 kb was detected, whereas PstI-digested mouse DNA gave a band of 4.2 kb, when probed with the mouse erythroid beta-spectrin cDNA clone. This allowed us to analyze a panel of mouse X Chinese hamster somatic cell hybrids to map this gene to chromosome 12. Interestingly, this assignment is different from that observed for the alpha-subunit of spectrin, which has been mapped to chromosome 1 in mouse. These results serve as a basis for further genetic characterization of the mouse hemolytic anemias.

We have examined EcoRI-restricted cellular DNA from mouse-hamster somatic cell hybrids. Results of this analysis show that the unit II mouse mammary tumor virus proviral genome is located on mouse chromosome 6. Restriction analysis of cellular DNA from (C3H/OuJ X Czech II) X Czech II backcross mice showed a strong linkage between unit II and Igk. The gene order of these markers on chromosome 6 relative to the Raf and Kirsten murine sarcoma virus ras-2 proto-oncogenes was established.

The Gross passage A murine leukemia virus (MuLV) induced T-cell leukemia of clonal (or oligoclonal) origin in inoculated mice. To study the role of the integrated proviruses in these tumor cells, we cloned several newly integrated proviruses (with their flanking cellular sequences) from a single tumor in procaryotic vectors. With each of the five clones obtained, a probe was prepared from the cellular sequences flanking the provirus. With one such probe (SS8), we screened several Gross passage A MuLV-induced SIM.S mouse tumor DNAs and found that, in 11 of 40 tumors, a provirus was integrated into a common region designated Gin-1. A 26-kilobase-pair sequence of Gin-1 was cloned from two lambda libraries, and a restriction map was derived. All proviruses were integrated as a cluster in the same orientation within a 5-kilobase-pair region of Gin-1, and most of them had a recombinant structure of the mink cell focus-forming virus type. The frequency of Gin-1 occupancy by provirus was much lower in thymoma induced by other strains of MuLV in other mouse strains. Using somatic-cell hybrid DNAs, we mapped Gin-1 on mouse chromosome 19. Gin-1 was not homologous to 16 known oncogenes and was distinct from the other common regions for provirus integration previously described. Therefore, Gin-1 appears to represent a new common provirus integration region. The integration of a provirus within Gin-1 might be an important event leading to T-cell transformation, and the Gin-1 region might harbor sequences which are involved in tumor development.

The inherited retinal degeneration of the rd mouse results in the exclusive loss of one cell type, the photoreceptors. We took advantage of this visual-cell loss to devise a strategy for the isolation of photoreceptor-specific cDNAs based on the use of subtractive and differential hybridizations. The resulting pool of photoreceptor-specific cDNAs was screened for a candidate cDNA for the rd gene, and a putative rd cDNA that maps to mouse chromosome 5, the chromosome to which the rd gene has been assigned, was identified. On Northern blots the candidate rd cDNA hybridizes a 3.3-kilobase RNA species from 9- to 11-day-old developing normal retina and, much more faintly, a 3.6-kb RNA species from age-matched rd retina. The 0.3-kilobase difference in the size of the mRNAs hybridized suggests that a structural alteration in the gene corresponding to the candidate rd cDNA has occurred in the rd mouse. This was further supported by the detection of polymorphisms between rd/rd and +/+ mouse genomic DNA after digestion with restriction endonucleases and probing with the candidate rd cDNA. Expression of mRNAs hybridized by the candidate rd cDNA is detected in normal and diseased retinas at postnatal day 1 but the signal intensity is considerably lower in the rd retina. To our knowledge, this is the earliest molecular defect reported in the rd retina that is observed prior to any phenotypic signs of photoreceptor degeneration.

We have examined EcoRI-restricted cellular DNA from BALB/c mouse-hamster somatic cell hybrids by blot hybridization for the presence of mouse mammary tumor virus-related sequences. Results of this analysis show that mouse mammary tumor virus-related proviral copies are located on chromosomes 16 (16-kilobase-pair fragment) and 12 (10.5- and 7.7-kilobase-pair fragments).

By using alpha-spectrin cDNA clones of murine and human origin and somatic cell hybrids segregating either mouse or human chromosomes, the gene for alpha-spectrin has been mapped to chromosome 1 in both species. This assignment of the mouse alpha-spectrin gene to mouse chromosome 1 by DNA hybridization strengthens the previous identification of the alpha-spectrin locus in mouse with the sph locus, which previously was mapped by linkage analysis to mouse chromosome 1, distal to the Pep-3 locus. By in situ hybridization to human metaphase chromosomes, the human alpha-spectrin gene has been localized to 1q22-1q25; interestingly, the locus for a non-Rh-linked form of elliptocytosis has been provisionally mapped to band 1q2 by family linkage studies.

Chinese hamster X mouse somatic cell hybrids were analyzed by Southern blot hybridization with a probe specific for the cellular c-fms proto-oncogene. Results demonstrate that Fms, the genetic locus containing this sequence, maps to mouse chromosome 18. Mouse Fms is thus not linked to the same set of genes involved in growth regulation that human FMS is linked to.

MUTANT mice homozygous for 'retinal degeneration slow' (rds/rds) are characterized phenotypically by abnormal development of photoreceptor outer segments in the retina, followed by gradual degeneration of photoreceptors1–3. This process of degeneration is complete by one year, with preservation of all other retinal cells4. The biochemical defect that leads to the mutant phenotype is not known. Our strategy for cloning the rds gene was based upon three previously reported observations. First, the rds locus maps to chromosome 175,6. Second, experimental rds/rds ↦+/+ and rds/+ ↦ +/+ tetra-parental mice manifest patchy photoreceptor changes in the retina7,8 suggesting that the wild-type rds locus is expressed within cells of the photoreceptor lineage. Finally, the process of degeneration is specific to photore-ceptors. On the basis of these observations, we predicted that the rds mRNA is encoded by a gene on chromosome 17 and is normally expressed exclusively within photoreceptors in the retina. We here present evidence that this is the case.

The human T lymphocyte antigen CD2 (T11, sheep erythrocyte receptor) is expressed on all peripheral T cells and all but the most immature thymocytes. Experiments with monoclonal antibodies against CD2 suggest that CD2 is the cell surface receptor for a natural ligand involved in T cell proliferation. Clarification of the functional role of CD2 would be facilitated by the identification of CD2 in the mouse. However, antibodies that recognize the murine homologue have not been described. An alternative approach to the identification of the murine homologue was to use cross-species DNA hybridization, employing human CD2 cDNA as a probe. Clones encoding the murine homologue were isolated from a murine T helper cell cDNA library. The murine cDNA sequence encoded a predicted mature polypeptide of 322 amino acids that showed 54% identity with the predicted human sequence. As with the human polypeptide, the cytoplasmic domain was large, and rich in proline and basic residues. CD2 mRNA was expressed in murine thymus and spleen, and in the T cell line EL4. The murine CD2 gene was assigned to chromosome 3 by Southern blot analysis of mouse-hamster somatic cell hybrids. A rabbit antiserum raised against purified human CD2, precipitated from surface-labeled mouse thymocytes a glycoprotein of Mr 55,000-66,000 which decreased to Mr 35,000 on digestion with endo-beta-acetylglucosaminidase F. These sizes are consistent with those predicted for the murine CD2 antigen from the cDNA sequence.