• Molecules and Cells
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• v.47 no.1
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• pp.100005.1-100005.15
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• 2024

Amyotrophic lateral sclerosis is a devastating neurodegenerative disease with a complex genetic basis, presenting both in familial and sporadic forms. The hexanucleotide (G₄C₂) repeat expansion in the C9orf72 gene, which triggers distinct pathogenic mechanisms, has been identified as a major contributor to familial and sporadic Amyotrophic lateral sclerosis cases. Animal models have proven pivotal in understanding these mechanisms; however, discrepancies between models due to variable transgene sequence, expression levels, and toxicity profiles complicate the translation of findings. Herein, we provide a systematic comparison of 7 publicly available Drosophila transgenes modeling the G₄C₂ expansion under uniform conditions, evaluating variations in their toxicity profiles. Further, we tested 3 previously characterized disease-modifying drugs in selected lines to uncover discrepancies among the tested strains. Our study not only deepens our understanding of the C9orf72 G₄C₂ mutations but also presents a framework for comparing constructs with minute structural differences. This work may be used to inform experimental designs to better model disease mechanisms and help guide the development of targeted interventions for neurodegenerative diseases, thus bridging the gap between model-based research and therapeutic application.

• Molecules and Cells
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• v.43 no.9
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• pp.821-830
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• 2020

Altered dendritic morphology is frequently observed in various neurological disorders including amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), but the cellular and molecular basis underlying these pathogenic dendritic abnormalities remains largely unclear. In this study, we investigated dendritic morphological defects caused by dipeptide repeat protein (DPR) toxicity associated with G4C2 expansion mutation of C9orf72 (the leading genetic cause of ALS and FTD) in Drosophila neurons and characterized the underlying pathogenic mechanisms. Among the five DPRs produced by repeat-associated non-ATG translation of G4C2 repeats, we found that arginine-rich DPRs (PR and GR) led to the most significant reduction in dendritic branches and plasma membrane (PM) supply in Class IV dendritic arborization (C4 da) neurons. Furthermore, expression of PR and GR reduced the number of Golgi outposts (GOPs) in dendrites. In Drosophila brains, expression of PR, but not GR, led to a significant reduction in the mRNA level of CrebA, a transcription factor regulating the formation of GOPs. Overexpressing CrebA in PR-expressing C4 da neurons mitigated PM supply defects and restored the number of GOPs, but the number of dendritic branches remained unchanged, suggesting that other molecules besides CrebA may be involved in dendritic branching. Taken together, our results provide valuable insight into the understanding of dendritic pathology associated with C9-ALS/FTD.

• The Plant Pathology Journal
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• v.15 no.5
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• pp.270-279
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• 1999

cDNAs complementary to the 3'-terminal regions of two potyvirus genomes were cloned and sequenced. The clone G7 contains one open reading frame (ORF) of 1,338 nucleotides and a 3' untranslated region (3'-UTR) of 403 nucleotides at the 3'-end excluding the 3'end poly(A) tail. The putative viral coat protein (CP) shows 55%-92% amino acid sequence homology to those of Allium potyviruses. The genome size of the virus was analyzed to be about 9.0 kb by Northern blot analysis. Five cDNA clones were screened out using GPV2 oligonucleotide as a probe. One of these clones, DEA72, which has a longest cDNA insert, contains one ORF of 1,459 nucleotides and a 3'-UTR of 590 nucleotides at the 3'-end excluding the 3'-end poly(A) tail. The putative viral CP shows 57%-88% amino acid sequence homologies to those of Allium potyviruses. The genome size of the virus was analyzed to be about 9.6 kb by Northern blot analysis. The results of immunoblot and Northern blot analyses suggest that almost all of the tested garlic plants showing mosaic or streak symptoms are infected with DEA72-potyvirus in variable degrees but rarely infected with G7-potyvirus in variable degrees but rarely infected with DEA72-potyvirus in variable degrees but rarely infected with G7-potyvirus. Immunoelectron microscopy using anti-DEA72 CP antibody shows that this potyvirus is about 750 nm long and flexuous rod shaped.

• Korean Journal of Veterinary Service
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• v.27 no.3
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• pp.273-280
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• 2004

The studies were performed for the PRRS antigen and antibody detection from breeding farms, artificial insemination(AI) center and growing farms in Jeonbuk province. 1. Specific PRRS primers were successfully amplified ORF6 617bp and ORF7 448 bp on agarose gel. 2. RT-PCR method has been establish by commercial kit and the thermal cycler program consisted of 30 cycles: $95^{\circ}C$ for 30 sec, $45^{\circ}C$ for 30 sec, and $72^{\circ}C$ for 45 sec. 3. The results of PRRS antibody test by ELISA method in AI centers were $6.6\%,\;53.3\%$ and breeding farms $65\%,\;65\%\;and\;38.7\%$, respectively. The serological positive of the antibody in gilt higher than sow. 4. The sero-positive of the PRRS antibody showed average $21\%$ in domestic farms, $56.2\%$ in breeding farms, and $29.9\%$ in AI center.

• Journal of Aquaculture
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• v.20 no.1
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• pp.65-72
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• 2007

Full length complementary DNA encoding apolipoprotein A-I (apoA-I) was isolated and characterized in mud loach (Misgurnus mizolepis). Mud loach apoA-I cDNA encoding 24 bp of 5'-untranslated region (UTR), 762 bp of single open reading frame (ORF) consists of 254 amino acids and 293 bp of 3'-UTR excluding stop codon and poly (A+) tail. Two overlapping polyadenylation signals (AATAAAATAAA) was found 9 bp prior to the poly (A+) tail. Mud loach apoA-I represented considerable homology to those from other teleost species at amino acid level with conserving common features of vertebrate apoA-I. Molecular phylogenetic analysis inferred the phylogenetic hypothesis that was generally in accordance with the previous taxonomic relationship. Apolipoprotein A-I mRNA was detected in various tissues, but the mRNA levels were quite varied depending on tissues based on semi-quantitative RT-PCR. Liver and brain showed the significantly higher levels of apoA-I transcripts than other tissues. mRNA expression of apoA-I was quite low in very early stage of embryonic development, however dramatically enhanced from 8 hours post fertilization. This increased mRNA level was retained consistently up to 14 days post hatching.

• Journal of fish pathology
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• v.23 no.3
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• pp.323-334
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• 2010

Bactericidal/permeability-increasing protein (BPI) and lipopolysaccharide-binding protein (LBP) are important components of the mammalian innate defence system against Gram-negative infections. The BPI/LBP cDNA was identified from the black rockfish ConA/PMA or LPS stimulated leukocyte cDNA library. The full-length BR-BPI/LBP cDNA was 2118 bp long and contained an open reading frame (ORF) of 1422 bp that encoded 473 amino-acid residues. The 5' UTR had a length of 57 bp, and the 3' UTR 639 bp. The molecular weight and theoretical isoelectric point (pI) values were calculated 51.4 kDa and 9.72, respectively. Compared with other known BPI or BPI/LBP peptide sequences, the most conserved regions of the black rockfish BPI/LBP peptide were found to be the BPI1 N-terminal, BPI2 C-terminal domains and a LPS binding domain. Phylogenetic analysis based on the deduced amino acid sequence revealed a homologous relationship between the BPI/LBP sequence of black rockfish and that of other teleosts. The black rockfish BPI/LBP gene was predominantly expressed in the PBLs, head kidney, trunk kidney and spleen. The expression of the black rockfish BPI/LBP molecule was induced in the peripheral blood leukocytes (PBLs) from 1 to 24 h following LPS stimulation, with a peak at 12 h post-stimulation.

• Journal of Microbiology and Biotechnology
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• v.10 no.2
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• pp.238-243
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• 2000

An exoinulinase (${\beta}-D-fructofuranosidase$) gene was cloned by chromosome walking along the upstream region of the endoinulinase gene of Pseudomonas mucidolens isolated from soil. the exoinulinase gene consisted of an ORF of 0,506 bp encoding a polypeptide of 501 amino acids with a deduced molecular weight of 55,000. The exoinulinase produced by the recombinant Escherichia coli $DH5{\alpha}$ strain was also purified to homogeneity as determined by SDS-PAGE and a zymogram. The molecular weight of the purified exoinulinase according to both SDS-PAGE and gel filtration matched the deduced molecular weight of the protein described above, thereby indicating that the native form of the exoinulinase was a monomer. The purified enzyme hydrolyzed activity value of 2.0. Furthermore, no inulo-oligomers were liberated from the inulin substrate in the enzymatic reaction mixtures incubated for 90 min at $55^{\circ}C$. Taken together, these results indicate that the purified ${\beta}-D-fructofuranosidase$ was an exoinulinase. The pH and temperature optima of the exoinulinase were pH 6.0 and $55^{\circ}C$, respectively. the enzymehad no apparent requirement for a cofactor, and its activity was completely inactivated by $Ag^{+},{\;}Hg^{2+},{\;}and{\;}Zn^{2+}$. Kinetic experiments gave $K_m,{\;}V_{max},{\;}and{\;}K_{cat}$ values for inulin of 11.5 mM, 18 nM/s, and $72{\;}s^{-1}$, respectively. the exoinulinase was fairly stable in broad pH conditions (pH 5-9), and at pH 6.0 it showed a residual activity of about 70% after 4 h incubation at $55^{\circ}C$.