Gene_locus Report for: mouse-ndr1

Mice affected by a spontaneous mutation which arose within our colony exhibited a neuromuscular phenotype involving tremor and characteristic stretching of the rear limbs. The mutant, named stretcher, was used to breed a backcross cohort for genetic mapping studies. The gene responsible for the mutant phenotype was mapped to a small region on mouse chromosome 15, with a LOD score above 20. Candidate genes within the region included the Ndrg1 gene. Examination of this gene in the mutant mouse strain revealed that exons 10 to 14 had been deleted. Mutations in the human orthologue are known to result in Charcot-Marie-Tooth disease type 4D (CMT4D) a severe early-onset disorder involving Schwann cell dysfunction and extensive demyelination. The stretcher mutant mouse is more severely affected than mice in which the Ndrg1 gene had been knocked out by homologous recombination. Our results demonstrate that the Ndrg1 (str) mutation provides a new model for CMT4D, and demonstrate that exons 10 to 14 of Ndrg1 encode amino acids crucial to the appropriate function of Ndrg1 in the central nervous system.

To identify genes regulated by N-myc, subtraction of whole embryo cDNA was carried out between wild type and N-myc-deficient mutant mice. Six cDNA clones were isolated as representing genes expressed higher in the mutant embryos and two as those expressed lower. One of them, Ndr1, coding for 43 kDa cytoplasmic protein was studied in detail. The Ndr1 gene was augmented 20-fold in the mutant embryos at 10.5 days post coitus which is indicative of repression by N-myc. An inverse relationship actually existed between the expression of N-myc and Ndr1 in various developing tissues of the wild type embryos. In the early stage of differentiation of these tissues when N-myc expression was high Ndr1 expression was low or undetectable, and later when N-myc activity diminished Ndr1 expression was augmented concomitantly with the occurrence of terminal differentiation. To establish the direct link between N-myc activity and the Ndr1 regulation, the Ndr1 gene was cloned and analyzed. The Ndr1 promoter activity was down-regulated by N-myc, and more strongly by the combination of N-myc and Max in the cotransfection assay. This repressive effect was mediated by the promoter region within 52 base pairs from the transcription start site but direct binding of N-myc:Max to the promoter sequence was not demonstrated, which is analogous to the cases recently reported for transcriptional repression by c-myc. c-myc also repressed Ndr1 promoter activity similarly to N-myc. The effect of N-myc:Max was sensitive to Trichostatin A, indicating involvement of histone deacetylase activity in repression of the Ndr1 promoter. The strategy we adopted in identifying target genes of a transcription factor should prove widely applicable when mutant animals are available.

By using mRNA polymerase chain reaction differential display technique (DDPCR), we have identified one early responsive cDNA fragment, TDD5, from a 5alpha-reductase-deficient T cell hybridoma. The DDPCR profiles of TDD5 suggest that its expression can be repressed by testosterone (T) within 2 hr. More importantly, both DDPCR and Northern blot analysis further demonstrated that the expression of TDD5 was differentially repressed by T and dihydrotestosterone (DHT) at the mRNA level. To our knowledge, this is the first androgen target gene to show a preference in response to T over DHT in cell culture. TDD5 is expressed in several tissues with particular abundance in kidney. Full-length TDD5 cDNA (2,916 bp) encodes a protein with a calculated molecular weight of 42,000. Finally, our animal studies further confirm that TDD5 mRNA levels can be repressed to the basal level 8 hr after DHT administration. The isolation and characterization of the early-responsive androgen target gene TDD5 and the fact that TDD5 mRNA level can be differentially regulated by T and DHT may provide a useful tool to study the molecular mechanism of androgen preference on target gene regulation.

Mice affected by a spontaneous mutation which arose within our colony exhibited a neuromuscular phenotype involving tremor and characteristic stretching of the rear limbs. The mutant, named stretcher, was used to breed a backcross cohort for genetic mapping studies. The gene responsible for the mutant phenotype was mapped to a small region on mouse chromosome 15, with a LOD score above 20. Candidate genes within the region included the Ndrg1 gene. Examination of this gene in the mutant mouse strain revealed that exons 10 to 14 had been deleted. Mutations in the human orthologue are known to result in Charcot-Marie-Tooth disease type 4D (CMT4D) a severe early-onset disorder involving Schwann cell dysfunction and extensive demyelination. The stretcher mutant mouse is more severely affected than mice in which the Ndrg1 gene had been knocked out by homologous recombination. Our results demonstrate that the Ndrg1 (str) mutation provides a new model for CMT4D, and demonstrate that exons 10 to 14 of Ndrg1 encode amino acids crucial to the appropriate function of Ndrg1 in the central nervous system.

Although most tissues in an organism are genetically identical, the biochemistry of each is optimized to fulfill its unique physiological roles, with important consequences for human health and disease. Each tissue's unique physiology requires tightly regulated gene and protein expression coordinated by specialized, phosphorylation-dependent intracellular signaling. To better understand the role of phosphorylation in maintenance of physiological differences among tissues, we performed proteomic and phosphoproteomic characterizations of nine mouse tissues. We identified 12,039 proteins, including 6296 phosphoproteins harboring nearly 36,000 phosphorylation sites. Comparing protein abundances and phosphorylation levels revealed specialized, interconnected phosphorylation networks within each tissue while suggesting that many proteins are regulated by phosphorylation independently of their expression. Our data suggest that the "typical" phosphoprotein is widely expressed yet displays variable, often tissue-specific phosphorylation that tunes protein activity to the specific needs of each tissue. We offer this dataset as an online resource for the biological research community.

The mouse (Mus musculus) is the premier animal model for understanding human disease and development. Here we show that a comprehensive understanding of mouse biology is only possible with the availability of a finished, high-quality genome assembly. The finished clone-based assembly of the mouse strain C57BL/6J reported here has over 175,000 fewer gaps and over 139 Mb more of novel sequence, compared with the earlier MGSCv3 draft genome assembly. In a comprehensive analysis of this revised genome sequence, we are now able to define 20,210 protein-coding genes, over a thousand more than predicted in the human genome (19,042 genes). In addition, we identified 439 long, non-protein-coding RNAs with evidence for transcribed orthologs in human. We analyzed the complex and repetitive landscape of 267 Mb of sequence that was missing or misassembled in the previously published assembly, and we provide insights into the reasons for its resistance to sequencing and assembly by whole-genome shotgun approaches. Duplicated regions within newly assembled sequence tend to be of more recent ancestry than duplicates in the published draft, correcting our initial understanding of recent evolution on the mouse lineage. These duplicates appear to be largely composed of sequence regions containing transposable elements and duplicated protein-coding genes; of these, some may be fixed in the mouse population, but at least 40% of segmentally duplicated sequences are copy number variable even among laboratory mouse strains. Mouse lineage-specific regions contain 3,767 genes drawn mainly from rapidly-changing gene families associated with reproductive functions. The finished mouse genome assembly, therefore, greatly improves our understanding of rodent-specific biology and allows the delineation of ancestral biological functions that are shared with human from derived functions that are not.

NDRG1 is an intracellular protein that is induced under a number of stress and pathological conditions, and it is thought to be associated with cell growth and differentiation. Recently, human NDRG1 was identified as a gene responsible for hereditary motor and sensory neuropathy-Lom (classified as Charcot-Marie-Tooth disease type 4D), which is characterized by early-onset peripheral neuropathy, leading to severe disability in adulthood. In this study, we generated mice lacking Ndrg1 to analyze its function and elucidate the pathogenesis of Charcot-Marie-Tooth disease type 4D. Histological analysis showed that the sciatic nerve of Ndrg1-deficient mice degenerated with demyelination at about 5 weeks of age. However, myelination of Schwann cells in the sciatic nerve was normal for 2 weeks after birth. Ndrg1-deficient mice showed muscle weakness, especially in the hind limbs, but complicated motor skills were retained. In wild-type mice, NDRG1 was abundantly expressed in the cytoplasm of Schwann cells rather than the myelin sheath. These results indicate that NDRG1 deficiency leads to Schwann cell dysfunction, suggesting that NDRG1 is essential for maintenance of the myelin sheaths in peripheral nerves. These mice will be used for future analyses of the mechanisms of myelin maintenance.

Differentiation-related gene-1 (Drg-1) has been identified as a gene whose expression is increased in several processes related to differentiation, but its function is currently unknown. In this report, we show that Drg-1 was expressed in keratinocytes, this expression being rapidly increased as a result of induction by 12-O-tetradecanoylphorbol-13-acetate (TPA) or the presence of an activating form of Ha-ras. Induction by TPA occurred both in cultured cell lines and primary keratinocytes as well as in mouse skin after a single TPA application. Overexpression of Drg-1 was also observed in TPA-induced hyperplastic skin. In agreement, mouse skin papillomas and carcinomas also overexpressed Drg-1. In addition, Drg-1 was induced when keratinocytes were forced to differentiate by calcium switch or serum starvation. Analysis of the expression of Drg-1 during the keratinocyte cell cycle demonstrated relatively high levels of Drg-1 mRNA in G(0), which increased in early G(1) and decreased afterwards in late G(1)/S. In situ analysis showed an accumulation of Drg-1 in the suprabasal layers of the skin, as well as in the more differentiated areas of mouse skin papillomas. These results suggest that, in addition to being upregulated during keratinocyte differentiation, the Drg-1 gene might have a complex function in skin tumorigenesis.

To identify genes regulated by N-myc, subtraction of whole embryo cDNA was carried out between wild type and N-myc-deficient mutant mice. Six cDNA clones were isolated as representing genes expressed higher in the mutant embryos and two as those expressed lower. One of them, Ndr1, coding for 43 kDa cytoplasmic protein was studied in detail. The Ndr1 gene was augmented 20-fold in the mutant embryos at 10.5 days post coitus which is indicative of repression by N-myc. An inverse relationship actually existed between the expression of N-myc and Ndr1 in various developing tissues of the wild type embryos. In the early stage of differentiation of these tissues when N-myc expression was high Ndr1 expression was low or undetectable, and later when N-myc activity diminished Ndr1 expression was augmented concomitantly with the occurrence of terminal differentiation. To establish the direct link between N-myc activity and the Ndr1 regulation, the Ndr1 gene was cloned and analyzed. The Ndr1 promoter activity was down-regulated by N-myc, and more strongly by the combination of N-myc and Max in the cotransfection assay. This repressive effect was mediated by the promoter region within 52 base pairs from the transcription start site but direct binding of N-myc:Max to the promoter sequence was not demonstrated, which is analogous to the cases recently reported for transcriptional repression by c-myc. c-myc also repressed Ndr1 promoter activity similarly to N-myc. The effect of N-myc:Max was sensitive to Trichostatin A, indicating involvement of histone deacetylase activity in repression of the Ndr1 promoter. The strategy we adopted in identifying target genes of a transcription factor should prove widely applicable when mutant animals are available.

By using mRNA polymerase chain reaction differential display technique (DDPCR), we have identified one early responsive cDNA fragment, TDD5, from a 5alpha-reductase-deficient T cell hybridoma. The DDPCR profiles of TDD5 suggest that its expression can be repressed by testosterone (T) within 2 hr. More importantly, both DDPCR and Northern blot analysis further demonstrated that the expression of TDD5 was differentially repressed by T and dihydrotestosterone (DHT) at the mRNA level. To our knowledge, this is the first androgen target gene to show a preference in response to T over DHT in cell culture. TDD5 is expressed in several tissues with particular abundance in kidney. Full-length TDD5 cDNA (2,916 bp) encodes a protein with a calculated molecular weight of 42,000. Finally, our animal studies further confirm that TDD5 mRNA levels can be repressed to the basal level 8 hr after DHT administration. The isolation and characterization of the early-responsive androgen target gene TDD5 and the fact that TDD5 mRNA level can be differentially regulated by T and DHT may provide a useful tool to study the molecular mechanism of androgen preference on target gene regulation.