• 한국발생생물학회:학술대회논문집
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• 한국발생생물학회 2001년도 발생공학 국제심포지움 및 학술대회 발표자료집
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• pp.14-17
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• 2001

Positional clonging (map-based cloning) of mutations or genetic variations has been served as an invaluable tool to understand in-vivo functions of genes and to identify molecular components underlying phenotypes of interest. Mice homozygous for the cerebellar deficient folia (cdf) mutation are ataxic, with cerebellar hypoplasia and abnormal lobulation of the cerebellum. In the cdf mutant cerebellum approximately 40% of Purkinje cells are ectopically located within the white matter and the inner granule cell layer (IGL). To identify the cdf gene, a high-resolution genetic map for the cdf-gene-encompassing region was constructed using 1997 F2 mice generated from C3H/HeSnJ-cdf/cdf and CAST/Ei intercross. The cdf gene showed complete linkage disequilibrium with three tightly linked markers D6Mit208, D6Mit359, and D6Mit225. A contig using YAC, BAC, and P1 clones was constructed for the cdf critical region to identify the gene. A deletion in the cdf critical region on chromosome 6 that removes approximately 150 kb of DNA selection. cdf mutant mice with the transgenic copy of the identified gene restored the brain abnormalities of the mutant mice. The positional cloning of cdf gene provides a good example showing the identification of a gene could lead to finding a new component of important molecular pathways.

• 한국동물번식학회:학술대회논문집
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• 한국동물번식학회 2001년도 발생공학 국제심포지움 및 학술대회 발표자료집
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• pp.14-17
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• 2001

Positional cloning (map-based cloning) of mutations or genetic variations has been served as an invaluable tool to understand in-vivo functions of genes and to identify molecular components underlying phenotypes of interest. Mice homozygous for the cerebellar deficient folia (cdf) mutation are ataxic, with cerebellar hypoplasia and abnormal lobulation of the cerebellum. In the cdf mutant cerebellum approximately 40% of Purkinje cells are ectopically located within the white matter and the inner granule cell layer (IGL). To identify the cdf gene, a high-resolution genetic map for the cdf-gene-encompassing region was constructed using 1997 F2 mice generated from C3H/HeSnJ-cdf/cdf and CAST/Ei intercross. The cdf gene showed complete linkage disequilibrium with three tightly linked markers D6Mit208, D6Mit359, and D6Mit225. A contig using YAC, BAC, and P1 clones was constructed for the cdf critical region to identify the gene. A deletion in the cdf critical region on chromosome 6 that removes approximately 150kb of DNA was identified. A gene associated with this deletion was identified using cDNA selection. cdf mutant mice with the transgenic copy of the identified gene restored the brain abnormalities of the mutant mice. The positional cloning of cdf gene provides a good example showing the identification of a gene could lead to finding a new component of important molecular pathways.

• Animal cells and systems
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• 제1권4호
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• pp.589-594
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• 1997

The neuronal voltage-sensitive calcium channels (VSCCs) are multisubunit complexes consisting of $\alpha_1,\;\alpha_2-\delta$ and $\beta$ subunits. Heterologous expression and biochemical studies have shown that the activity of VSCCs is regulated by their $\beta$ subunits in a $\beta$ subunit isoform-specific manner. To elucidate the $\beta$ subunit identity of the P/Q-type calcium channel encoded by an $\alpha_{1A}$ subunit, which is exclusively expressed in the Purkinje and granule cell of the cerebellum, we have examined the spatial and temporal expression patterns of $\beta$ subunits and compared them with those of $\alpha_{1A}$ subunit in the developing rat cerebellum. Reverse transcriptase- polymerase chain reaction (RT-PCR) and Northern blot analysis have shown that $\beta_4$ subunit mRNA was prominently expressed in the cerebellum and much more abundant than any other distinct $\beta$ subunits. RNase protection assay has further demonstrated that the expression of $\alpha_{1A}$ and $\beta_4$ subunits increased during cerebellar development, while the amount of $\beta_2$ and $\beta_3$ mRNAs did not significantly change. In addition, a $\beta_4$ transcript was present in cultured cerebellar granule cells, but not in astrocyte cells, and the level of $\beta_4$ mRNA expression increased gradually in vitro seen as in vivo. Based on the spatial and temporal expression patterns of $\beta_4$ subunit, we conclude that $\beta_4$ may predominantly associate, but probably not exclusively, with the $\alpha_{1A}$ subunit in rat cerebellar granule cells.

• Molecules and Cells
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• 제26권2호
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• pp.186-192
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• 2008

NELL2, a neural tissue-enriched protein, is produced in the embryo, and postembryonically in the mammalian brain, with a broad distribution. Although its synthesis is required for neuronal differentiation in chicks, not much is known about its function in the adult mammalian brain. We investigated the distribution of NELL2 in various regions of the adult rat brain to study its potential functions in brain physiology. Consistent with previous reports, NELL2-immunoreactivity (ir) was found in the cytoplasm of neurons, but not in glial fibrillary acidic protein (GFAP)-positive glial cells. The highest levels of NELL2 were detected in the hippocampus and the cerebellum. Interestingly, in the cerebellar cortex NELL2 was observed only in the GABAergic Purkinje cells not in the excitatory granular cells. In contrast, it was found mainly in the hippocampal dentate gyrus and pyramidal cell layer that contains mainly glutamatergic neurons. In the dentate gyrus, NELL2 was not detected in the GFAP-positive neural precursor cells, but was generally present in mature neurons of the subgranular zone, suggesting a role in this region restricted to mature neurons.

• Experimental and Molecular Medicine
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• 제50권11호
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• pp.8.1-8.14
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• 2018

We previously demonstrated that the direct transplantation of human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs) into the dentate gyrus ameliorated the neurological symptoms of Niemann-Pick type C1 (NPC1)-mutant mice. However, the clinical presentation of NPC1-mutant mice was not fully understood with a molecular mechanism. Here, we found 14,15-epoxyeicosatrienoic acid (14,15-EET), a cytochrome P450 (CYP) metabolite, from hUCB-MSCs and the cerebella of NPC1-mutant mice and investigated the functional consequence of this metabolite. Our screening of the CYP2J family indicated a dysregulation in the CYP system in a cerebellar-specific manner. Moreover, in Purkinje cells, CYP2J6 showed an elevated expression level compared to that of astrocytes, granule cells, and microglia. In this regard, we found that one CYP metabolite, 14,15-EET, acts as a key mediator in ameliorating cholesterol accumulation. In confirming this hypothesis, 14,15-EET treatment reduced the accumulation of cholesterol in human NPC1 patient-derived fibroblasts in vitro by suppressing cholesterol synthesis and ameliorating the impaired autophagic flux. We show that the reduced activity within the CYP system in the cerebellum could cause the neurological symptoms of NPC1 patients, as 14,15-EET treatment significantly rescued cholesterol accumulation and impaired autophagy. We also provide evidence that the intranasal administration of hUCB-MSCs is a highly promising alternative to traumatic surgical transplantation for NPC1 patients.