Programs of the Human Genetic Mutant Cell Repository1

AbstractEpidermal growth factor receptor (EGFR) is a key oncogene in lung adenocarcinoma (LUAD). Resistance to EGFR tyrosine kinase inhibitors is a major obstacle for EGFR-mutant LUAD patients. Our gene chip array, quantitative polymerase chain reaction validation, and shRNA-based high-content screening identified the Akt kinase lanthionine synthetase C-like protein 2 (LANCL2) as a pro-proliferative gene in the EGFR-mutant LUAD cell line PC9. Therefore, we investigated whether LANCL2 plays a role in promoting cell proliferation and drug resistance in EGFR-mutant LUAD. In silico clinical correlation analysis using the Cancer Genome Atlas Lung Adenocarcinoma dataset revealed a positive correlation between LANCL2 and EGFR expression and an inverse relationship between LANCL2 gain-of-function and survival in LUAD patients. The EGFR-mutant LUAD cell lines PC9 and HCC827 displayed higher LANCL2 expression than the non-EGFR-mutant cell line A549. In addition, LANCL2 was downregulated following gefitinib+pemetrexed combination therapy in PC9 cells. LANCL2 knockdown reduced proliferation and enhanced apoptosis in PC9, HCC827, and A549 cells in vitro and suppressed murine PC9 xenograft tumor growth in vivo. Notably, LANCL2 overexpression rescued these effects and promoted gefitinib + pemetrexed resistance in PC9 and HCC827 cells. Pathway analysis and co-immunoprecipitation followed by mass spectrometry of differentially-expressed genes in LANCL2 knockdown cells revealed enrichment of several cancer signaling pathways. In addition, Filamin A and glutathione S-transferase Mu 3 were identified as two novel protein interactors of LANCL2. In conclusion, LANCL2 promotes tumorigenic proliferation, suppresses apoptosis, and promotes gefitinib+pemetrexed resistance in EGFR-mutant LUAD cells. Based on the positive association between LANCL2, EGFR, and downstream Akt signaling, LANCL2 may be a promising new therapeutic target for EGFR-mutant LUAD.

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A mammalian mutant cell lacking detectable lanosterol 14 alpha-methyl demethylase activity.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(19)57293-3 ◽

1988 ◽

Vol 263 (3) ◽

pp. 1248-1254

Author(s):

H W Chen ◽

D A Leonard ◽

R T Fischer ◽

J M Trzaskos

Keyword(s):

Mutant Cell ◽

Demethylase Activity

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A Mutation in Paramecium tetraurelia Reveals Functional and Structural Features of Developmentally Excised DNA Elements

Genetics ◽

10.1093/genetics/148.1.139 ◽

1998 ◽

Vol 148 (1) ◽

pp. 139-149 ◽

Cited By ~ 4

Author(s):

Kimberly M Mayer ◽

Kazuyuki Mikami ◽

James D Forney

Keyword(s):

Mutant Cell ◽

Consensus Sequence ◽

Surface Protein ◽

Structural Features ◽

Inverted Terminal Repeat ◽

Paramecium Tetraurelia ◽

Coding Region ◽

Conserved Sequence ◽

Variable Surface ◽

Dna Elements

Abstract The excision of internal eliminated sequences (IESs) from the germline micronuclear DNA occurs during the differentiation of a new macronuclear genome in ciliated protozoa. In Paramecium, IESs are generally short (28–882 bp), AT rich DNA elements that show few conserved sequence features with the exception of an inverted-terminal-repeat consensus sequence that has similarity to the ends of mariner/Tc1 transposons (Klobutcher and Herrick 1995). We have isolated and analyzed a mutant cell line that cannot excise a 370-bp IESs (IES2591) from the coding region of the 51A variable surface protein gene. A single micronuclear C to T transition within the consensus sequence prevents excision. The inability to excise IES2591 has revealed a 28-bp IES inside the larger IES, suggesting that reiterative integration of these elements can occur. Together, the consensus sequence mutation and the evidence for reiterative integration support the theory that Paramecium IESs evolved from transposable elements. Unlike a previously studied Paramecium IES, the presence of this IES in the macronucleus does not completely inhibit excision of its wild-type micronuclear copy through multiple sexual generations.

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Selection of a mutant cell line based on differential expression of glycosphingolipid, utilizing anti-lactosylceramide antibody and complement.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(18)53983-1 ◽

1993 ◽

Vol 268 (3) ◽

pp. 2211-2216

Author(s):

T. Tsuruoka ◽

T. Tsuji ◽

H. Nojiri ◽

E.H. Holmes ◽

S. Hakomori

Keyword(s):

Cell Line ◽

Differential Expression ◽

Mutant Cell ◽

Mutant Cell Line ◽

Selection Of

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DDRE-03. IDH1-MUTANT GBM CELLS ARE HIGHLY SENSITIVE TO COMBINATION OF KDM6A/B AND HDAC INHIBITORS

Neuro-Oncology Advances ◽

10.1093/noajnl/vdab024.025 ◽

2020 ◽

Vol 3 (Supplement_1) ◽

pp. i6-i7

Author(s):

Alişan Kayabölen ◽

Gizem Nur Sahin ◽

Fidan Seker ◽

Ahmet Cingöz ◽

Bekir Isik ◽

...

Keyword(s):

Cell Cycle ◽

Cell Cycle Arrest ◽

Cell Lines ◽

Mutant Cell ◽

Glioma Cells ◽

Epigenetic Therapy ◽

Low Grade ◽

Orthotopic Model ◽

Cycle Arrest ◽

Mutant Cells

Abstract Mutations in IDH1 and IDH2 genes are common in low grade gliomas and secondary GBM and are known to cause a distinct epigenetic landscape in these tumors. To interrogate the epigenetic vulnerabilities of IDH-mutant gliomas, we performed a chemical screen with inhibitors of chromatin modifiers and identified 5-azacytidine, Chaetocin, GSK-J4 and Belinostat as potent agents against primary IDH1-mutant cell lines. Testing the combinatorial efficacy of these agents, we demonstrated GSK-J4 and Belinostat combination as a very effective treatment for the IDH1-mutant glioma cells. Engineering established cell lines to ectopically express IDH1R132H, we showed that IDH1R132H cells adopted a different transcriptome with changes in stress-related pathways that were reversible with the mutant IDH1 inhibitor, GSK864. The combination of GSK-J4 and Belinostat was highly effective on IDH1R132H cells, but not on wt glioma cells or nonmalignant fibroblasts and astrocytes. The cell death induced by GSK-J4 and Belinostat combination involved the induction of cell cycle arrest and apoptosis. RNA sequencing analyses revealed activation of inflammatory and unfolded protein response pathways in IDH1-mutant cells upon treatment with GSK-J4 and Belinostat conferring increased stress to glioma cells. Specifically, GSK-J4 induced ATF4-mediated integrated stress response and Belinostat induced cell cycle arrest in primary IDH1-mutant glioma cells; which were accompanied by DDIT3/CHOP-dependent upregulation of apoptosis. Moreover, to dissect out the responsible target histone demethylase, we undertook genetic approach and demonstrated that CRISPR/Cas9 mediated ablation of both KDM6A and KDM6B genes phenocopied the effects of GSK-J4 in IDH1-mutant cells. Finally, GSK-J4 and Belinostat combination significantly decreased tumor growth and increased survival in an orthotopic model in mice. Together, these results suggest a potential combination epigenetic therapy against IDH1-mutant gliomas.

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NIGMS human genetic mutant cell repository

Mammalian Genome ◽

10.1007/bf00356547 ◽

1994 ◽

Vol 5 (6) ◽

pp. 323-323

Keyword(s):

Mutant Cell

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Impaired clearance of free cystine from lysosome-enriched granular fractions of I-cell-disease fibroblasts

Biochemical Journal ◽

10.1042/bj2370009 ◽

1986 ◽

Vol 237 (1) ◽

pp. 9-15 ◽

Cited By ~ 5

Author(s):

F Tietze ◽

L H Rome ◽

J D Butler ◽

G S Harper ◽

W A Gahl

Keyword(s):

Mutant Cell ◽

Cell Types ◽

Nephropathic Cystinosis ◽

Cell Disease ◽

Rapid Loss ◽

Normal Cells ◽

Cultured Fibroblasts ◽

Carrier Mechanism ◽

Mucolipidosis Ii

Cultured fibroblasts from patients with I-cell disease (mucolipidosis II) accumulate excessive amounts of free cystine, similarly to cells from patients with nephropathic cystinosis, a disorder of lysosomal cystine transport. To clarify whether the intralysosomal accumulation of cystine in I-cell-disease fibroblasts was due to a defective disposal mechanism, we measured the rates of clearance of free [35S]cystine from intact normal, cystinotic and I-cell-disease fibroblasts. Loss of radioactivity from the two mutant cell types occurred slowly (t 1/2 = 500 min) compared with the rapid loss from normal cells (t 1/2 = 40 min). Lysosome-rich granular fractions isolated from three different cystine-loaded normal, cystinotic and I-cell-disease fibroblast strains were similarly examined for non-radioactive cystine egress. Normal granular fractions lost cystine rapidly (mean t 1/2 = 43 min), whereas cystinotic granular fractions did not lose any cystine (mean t 1/2 = infinity). I-cell-disease granular fractions displayed prolonged half-times for cystine disposal (mean = 108 min), suggesting that I-cell-disease fibroblasts, like cystinotic cells, possess a defective carrier mechanism for cystine transport.

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Identification of the ATPase Subunit of the Primary Maltose Transporter in the Hyperthermophilic Anaerobe Thermotoga maritima

Applied and Environmental Microbiology ◽

10.1128/aem.00930-17 ◽

2017 ◽

Vol 83 (18) ◽

Cited By ~ 4

Author(s):

Raghuveer Singh ◽

Derrick White ◽

Paul Blum

Keyword(s):

Thermotoga Maritima ◽

Mutant Cell ◽

Sugar Metabolism ◽

Genetic System ◽

Gene Replacement ◽

Atpase Subunit ◽

Content Type ◽

Fermentative Hydrogen Production ◽

Atpase Subunits ◽

Fermentative Hydrogen

ABSTRACT Thermotoga maritima is a hyperthermophilic anaerobic bacterium that produces molecular hydrogen (H2) by fermentation. It catabolizes a broad range of carbohydrates through the action of diverse ABC transporters. However, in T. maritima and related species, highly similar genes with ambiguous annotation obscure a precise understanding of genome function. In T. maritima, three putative malK genes, all annotated as ATPase subunits, exhibited high identity to each other. To distinguish between these genes, malK disruption mutants were constructed by gene replacement, and the resulting mutant cell lines were characterized. Only a disruption of malK3 produced a defect in maltose catabolism. To verify that the mutant phenotype arose specifically from malK3 inactivation, the malK3 mutation was repaired by recombination, and maltose catabolism was restored. This study demonstrates the importance of a maltose ABC-type transporter and its relationship to sugar metabolism in T. maritima. IMPORTANCE The application and further development of a genetic system was used here to investigate gene paralogs in the hyperthermophile Thermotoga maritima. The occurrence of three ABC transporter ATPase subunits all annotated as malK was evaluated using a combination of genetic and bioinformatic approaches. The results clarify the role of only one malK gene in maltose catabolism in a nonmodel organism noted for fermentative hydrogen production.

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