Dimitrina D. Pravtcheva

Interferons as gene activators: A cluster of six interferon-activatable genes is linked to the erythroid α-spectrin locus on murine chromosome 1

Several interferon-activatable murine genes were mapped to murine chromosomes by hybridizing cDNA probes to Southern blots of genomic DNA samples from a panel of mouse-hamster somatic cell hybrid lines. The 12 gene is located on chromosome 12 and it specifies a 3.6-kb mRNA. The 204 gene (specifying a 2.5-kb mRNA), and three genes of the 203 gene family (hybridizing to five mRNAs of sizes between 2 and 4.5 kb), together with the 202 gene (specifying a 2-kb mRNA) are located on murine chromosome 1. By restriction fragment length polymorphism analysis of DNA samples prepared from a panel of recombinant inbred mouse lines (C57BL/6J D DBA/2J) and from 85 [C3H/HeJ-gld/gld x Mus spretus) F1 X C3H/HeJ-gld/gld] backcross mice we established a close linkage of the 202, 203, and 204 genes to the erythroid alpha-spectrin gene (Spna-1) on distal murine chromosome 1. Cosmids containing the 202, 203, and 204 genes were isolated from a library derived from AKR mouse DNA. Southern blot analysis of such cosmids revealed: (a) hybridization of a partial 203 cDNA to three genes of the 203 gene family; (b) cross-hybridization of the 202 and 204 genes with one another and with a third gene (designated as 201 gene), and (c) a close linkage of genes of the 203 family with the 201, 202, and 204 genes. These results indicate the existence of a cluster of at least six closely linked, interferon-activatable genes on distal murine chromosome 1 in the vicinity of the Spna-1 locus and also of the Minor lymphocyte stimulating locus (Mlsa).

Localization of the cryptdin locus on mouse chromosome 8

Cryptdin is a defensin-related peptide, and its mRNA accumulates to high abundance in epithelial cells of intestinal crypts beginning in the second week of postnatal development. The cryptdin (Defcr) locus was assigned to mouse chromosome 8 by Southern blotting of DNAs from mouse/hamster somatic hybrid cell lines. Analysis of somatic hybrid DNAs for mouse-specific restriction fragments showed zero discordance and perfect concordance with chromosome 8. The Defcr locus was localized on chromosome 8 by analysis of DNAs from recombinant inbred (RI) strains of mice after identification of three potential Defcr alleles based on restriction fragment length polymorphisms (RFLPs) in inbred strains. The strain distribution patterns of the Defcr locus were compared with those of chromosome 8 markers in five panels of RI strains. Analysis of cosegregation of Defcr with xenotropic proviral locus Xmv-26 and additional loci confirmed the chromosomal assignment and showed that Defcr is on proximal chromosome 8 within approximately 6 (1.3 to 21.3) cM of Xmv-26. The mouse Defcr locus and the human defensin gene(s) located on chromosome 8p23 appear to map to homologous regions.

Mapping of haploid expressed genes: genes for both mouse protamines are located on chromosome 16.

Mouse spermatozoa contain two protamines with different amino acid sequences. By hybridizing Southern blots of a series of mouse-hamster somatic cell hybrids containing subsets of mouse chromosomes and a complete set of hamster chromosomes with32P-labeled cDNAs for each mouse protamine, we assign the two mouse protamine genes to chromosome 16. This report presents the first evidence for chromosomal linkage of two sperm-specific, haploid regulated gene products.

Delayed Onset of Igf2-Induced Mammary Tumors in Igf2r Transgenic Mice

The insulin-like growth factor-II (IGF-II) receptor (IGF2R) regulates the level or activity of numerous proteins, including factors that control growth and differentiation. Frequent loss or inactivation of this receptor in a diverse group of tumors indicates that it may act as a tumor suppressor, but it is not known which functions of this receptor are selected against in the tumors. Lysosomal targeting and degradation of the growth-promoting IGF-II has been proposed as a mechanism for the tumor suppressor effects of IGF2R. As a genetic test of this hypothesis in vivo, we have produced Igf2r transgenic mice that ubiquitously express the transgene and have crossed these mice with mice that develop mammary tumors as a consequence of Igf2 overexpression. Our findings indicate that the presence of the Igf2r transgene delays mammary tumor onset and decreases tumor multiplicity in Igf2 transgenic mice. These findings are relevant to human tumors and preneoplastic conditions accompanied by altered IGF2 expression.

Localization of the murine λ5 gene on chromosome 16

The chromosomal location of the murine lambda 5 gene was analyzed by Southern hybridization using restriction enzyme-digested DNA from a panel of 15 mouse X hamster somatic cell hybrids. Sequences homologous with those of lambda 5 DNA were detected in DNA of 5 hybrids. In all 5 hybrids lambda 5 was contained in restriction fragments of equal sizes, the lengths of which indicated that the germline configuration of lambda 5 with three exons and the restriction sites expected from its genomic structure were present. Southern hybridization with the murine lambda 1 gene as a probe detected the same 5 hybrids as positive. The only mouse chromosome present on all of the positive hybrids, and absent from negative ones, was number 16. We conclude that lambda 5 is situated on the same chromosome as lambda 1, i.e., on the murine chromosome 16.

Isolation and regional localization of the murine homeobox-containing gene Hox-3.3 to mouse chromosome region 15E ☆

A murine homeobox-containing cDNA clone has been isolated from an adult spinal cord library. Using in situ hybridization and somatic cell genetics techniques, the newly isolated homeobox gene has been mapped to mouse chromosome region 15E. Because of its chromosomal location, we called this gene locus Hox-3.3. Nucleotide sequence analysis revealed that the Hox-3.3 gene represents the murine cognate of the human homeobox gene c8. The presumptive organization of the murine Hox-3 homeobox gene cluster is discussed.

Glial fibrillary acid protein, an Astrocytic-specific marker, maps to human chromosome 17

The murine glial fibrillary acid protein (GFAP) gene is located on chromosome 11 in close proximity to the genes encoding transforming protein p53 (Trp53) and myeloperoxidase (Mpo). Both Trp53 and Mpo have been mapped to human chromosome 17, but the chromosomal assignment of human GFAP has not been previously determined. In this report, we have amplified a cDNA fragment encoding a portion of GFAP from human brain and have used this probe to screen a mouse x human somatic cell hybrid panel. The results show that a human-specific GFAP species of approx 3.7 kb maps to one of these lines, TMS5, which contains chromosome 17 as its only human chromosome. On the basis of these data we speculate that there may be evolutionary relatedness between GFAP and other genes that map to both murine chromosome 11 and human chromosome 17.

Mapping of gene encoding mouse placental alkaline phosphatase to chromosome 4

The gene encoding the mouse placental alkaline phosphatase (ALP; orthophosphoric-monoester phosphohydrolase, alkaline optimum, EC 3.1.3.1) is mapped to chromosome 4, based on Southern blot hybridization of the mouse cDNA with DNAs from mouse-Chinese hamster somatic cell hybrids. This assignment is consistent with the genetic analysis of theAkp-2 locus, which is responsible for the genetic variation of alkaline phosphatase enzyme in placenta as well as in liver, kidney, and bone.

Transgene instability in mice injected with an in vitro methylated Igf2 gene.

Foreign DNA injected into mouse embryos integrates into the host chromosomes and is usually transmitted stably to the progeny. Rare cases of transgene instability have been described, and these can help our understanding of the rules that govern the organization and stability of endogenous DNA. We have observed unusual inheritance in three transgenic lines produced with a partially in vitro methylated Igf2 construct. All three founders transmitted to their progeny two different transgene patterns, A and B. Pattern A was inherited in accordance with expectation, whereas pattern B was associated with several abnormal characteristics, including fewer than expected transgenic progeny, evidence for instability and loss from the somatic tissues of some of the progeny, and high incidence of runting and perinatal death that did not appear correlated with transgene retention. The absence of these features in transgenic mice produced with the unmethylated version of the same construct indicated that prior methylation played a role in the unusual behavior of these transgenes. We hypothesize that patterns A and B were formed by transgenes that differed in their methylation, and that pattern B methylation led to instability of the transgene locus. Runting and early lethality in the pattern B sublines may be the result of transgene rearrangements, which result in transgene amplification with adverse effects of increased IGFII dosage, and/or deletions, which may affect endogenous genes required for viability. These findings provide further evidence that DNA methylation plays a role in genome stability and indicate that perturbations in the normal pattern of methylation may have destabilizing effects that extend through several generations.

Association of foreign DNA sequence with male sterility and translocation in a line of transgenic mice

We have analyzed a line of transgenic mice derived from injection of a cloned human interferon cDNA. This line manifests total male sterility of males carrying the human sequence, while male littermates not harboring the foreign DNA are fertile. All females are fertile. Karyotypes of transgenic animals show 2∶12 translocation. The microinjected sequence maps to one of the translocation chromosomes composed of a large portion of chromosome 12 to which has been translocated a segment of chromosome 2. Analysis of the sterile males reveals significant abnormalities of spermatogenesis and faulty chromosome synapsis that involves the translocation chromosomes. These findings show that transfer of foreign DNA into mouse embryos may lead to chromatin breakage and infertility of transgenic animals.