In vitro cultivation of human renal cell cancer

Williams, Richard D.; Elliott, Arthur Y.; Stein, Nell; Fraley, Elwin E.

doi:10.1007/bf02797460

Cited by 52 publications

(19 citation statements)

References 13 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Epimastigotes were maintained in logarithmic growth phase at 26°C in supplemented liver digest-neutralized tryptose medium as described previously (21). Culture-derived amastigotes were obtained by infecting monolayers of the RA786 renal carcinoma cell line (22) grown at 37°C in RPMI supplemented with 10% heat-inactivated fetal bovine serum, 5% L-glutamine, and 5% penicillin/ streptomycin. Amastigotes were purified by centrifugation in metrizamide gradients as described previously (2).…”

Section: Methodsmentioning

confidence: 99%

Amastin mRNA Abundance in Trypanosoma cruzi Is Controlled by a 3′-Untranslated Region Position-dependent cis-Element and an Untranslated Region-binding Protein

Coughlin¹,

Teixeira²,

Kirchhoff³

et al. 2000

Journal of Biological Chemistry

View full text Add to dashboard Cite

The genome of Trypanosoma cruzi contains tandem arrays of alternating genes encoding amastin and tuzin. Amastin is a surface glycoprotein abundantly expressed on the intracellular mammalian amastigote form of the protozoan parasite, and tuzin is a G-like protein. We demonstrated previously that the amastin-tuzin gene cluster is polycistronically transcribed to an equal extent in all parasite life cycle stages. The steady state level of amastin mRNA, however, is 68-fold more abundant in amastigotes than in epimastigotes. Here we show that the half-life of amastin mRNA is 7 times longer in amastigotes than in epimastigotes. Linker replacement experiments demonstrate that the middle onethird of the 630-nucleotide 3-untranslated region (UTR) is responsible for the amastin mRNA up-regulation. This positive effect is dependent on the distance of the 3-UTR segment from the stop codon and the polyadenylation site as well as on its orientation. A protein or protein complex more abundant in amastigotes than in epimastigotes binds to this minimally defined 3-UTR segment and may be involved in its regulatory function.Trypanosoma cruzi is the protozoan parasite that causes Chagas' disease, a debilitating illness that is a major cause of morbidity and mortality in many parts of Latin America. During its life cycle, T. cruzi passes through three developmental stages. In their insect vectors, the parasites multiply as extracellular epimastigotes in the midgut, migrate to the hindgut, and then differentiate into nondividing trypomastigotes. These infective organisms are excreted in the feces after a blood meal. When they contaminate the puncture site or mucous membranes of a mammalian host, they can invade a variety of cell types. Once inside host cells, the trypomastigotes differentiate into amastigotes, which multiply in the cytoplasm. After many cycles of proliferation, the amastigotes differentiate into trypomastigotes and enter the circulation when the host cell ruptures. These trypomastigotes perpetuate either the infection or the life cycle when they invade other host cells or are ingested by insect vectors, respectively.Since the intracellular amastigotes are at the core of the persistence of T. cruzi infection, we previously sought to identify genes that are preferentially expressed by amastigotes in order to develop a better understanding of the molecular biology of this parasite form. We identified two novel genes, encoding the proteins amastin and tuzin, which occur in alternating tandemly arranged pairs (1, 2). Amastin is an abundant glycoprotein on the surface of amastigotes. Less is understood about tuzin's function or location, but it possesses sequence similarity to a heterotrimeric G-protein. Most tandem genes in T. cruzi and other trypanosomatids, such as Leishmania species and African trypanosomes, do not have recognizable upstream promoters and are transcribed into polycistronic precursor RNAs (3-6). The 5Ј ends of the resulting mature monocistronic mRNAs are defined by the trans-splicing of a 39-nucleotide ...

show abstract

Section: Methodsmentioning

confidence: 99%

Amastin mRNA Abundance in Trypanosoma cruzi Is Controlled by a 3′-Untranslated Region Position-dependent cis-Element and an Untranslated Region-binding Protein

Coughlin¹,

Teixeira²,

Kirchhoff³

et al. 2000

Journal of Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…Both of them contain NotI sites. We subcloned sequences around these NotI sites and used them as probes for Southern blot hybridization of EcoRI/NotI digested DNA from UOK 121, RXF 393 (Stinson et al, 1992), UMRC 6 and 786 ± 0 (Williams et al, 1976) RCC cell lines (Figure 6). In all four samples the downstream NotI site was unmethylated while the upstream NotI site was methylated.…”

Section: Methylation Patterns Of Sequences¯anking the Vhl Genementioning

confidence: 99%

Analysis of aberrant methylation of the VHL gene by transgenes, monochromosome transfer, and cell fusion

Kuzmin

Geil

et al. 1999

Oncogene

View full text Add to dashboard Cite

Several tumor suppressor genes were shown to be inactivated by a process involving aberrant de novo methylation of their GC-rich promoters which is usually associated with transcriptional repression. The mechanisms underlying this process are poorly understood. In particular this abnormal methylation may be caused and/ or maintained by either de®ciency of some trans-acting factor(s) or by various malfunctions acting in cis. Here we studied the nature of aberrant methylation of the von Hippel-Lindau (VHL) disease tumor suppressor gene in a human clear cell renal carcinoma cell line, UOK 121, that contains a silent hypermethylated endogenous VHL allele. First, we transfected unmethylated VHL transgenes, driven by the VHL promoter, into UOK 121 cells. Next, to exclude possible position e ects that may in¯uence methylation of the introduced VHL genes, we transferred a single chromosome 3, carrying an apparently normal hypomethylated VHL allele into the UOK 121 cells. Finally, we created somatic cell hybrids between UOK 121 and UMRC 6 cells containing a mutant VHL-expressing hypomethylated allele. In these three experiments both the methylation of the VHL promoter and the transcriptional status of the introduced and endogenous VHL alleles remained unchanged. Our results demonstrate that the putative trans-acting factors present in the UOK 121 and UMRC 6 cells are unable to induce changes in methylation pattern of the VHL alleles in all cell lines and hybrids studied. Taken together, the results indicate that cis-speci®c local features are pivotal in maintaining and perpetuating aberrant methylation of the VHL CpG island. Contribution of some putative trans-acting factors cannot be excluded during a period when the aberrant VHL methylation pattern was ®rst generated.

show abstract

“…The 769-P cell line, established along with 786-O by [75], harbors mutated vhl and secretes high levels of VEGF, suggesting a ccRCC phenotype (Additional file 1: Table S1 for ref). These cells contain a SP of higher tumorigenicity that can create tumors even after serial passage in nude mice retaining their original histological characteristics [76].…”

Section: Introductionmentioning

confidence: 99%

Choosing the right cell line for renal cell cancer research

Brodaczewska¹,

Szczylik²,

et al. 2016

View full text Add to dashboard Cite

Cell lines are still a tool of choice for many fields of biomedical research, including oncology. Although cancer is a very complex disease, many discoveries have been made using monocultures of established cell lines. Therefore, the proper use of in vitro models is crucial to enhance our understanding of cancer. Therapeutics against renal cell cancer (RCC) are also screened with the use of cell lines. Multiple RCC in vitro cultures are available, allowing in vivo heterogeneity in the laboratory, but at the same time, these can be a source of errors. In this review, we tried to sum up the data on the RCC cell lines used currently. An increasing amount of data on RCC shed new light on the molecular background of the disease; however, it revealed how much still needs to be done. As new types of RCC are being distinguished, novel cell lines and the re-exploration of old ones seems to be indispensable to create effective in vitro tools for drug screening and more.Electronic supplementary materialThe online version of this article (doi:10.1186/s12943-016-0565-8) contains supplementary material, which is available to authorized users.

show abstract

In vitro cultivation of human renal cell cancer

Cited by 52 publications

References 13 publications

Amastin mRNA Abundance in Trypanosoma cruzi Is Controlled by a 3′-Untranslated Region Position-dependent cis-Element and an Untranslated Region-binding Protein

Amastin mRNA Abundance in Trypanosoma cruzi Is Controlled by a 3′-Untranslated Region Position-dependent cis-Element and an Untranslated Region-binding Protein

Analysis of aberrant methylation of the VHL gene by transgenes, monochromosome transfer, and cell fusion

Choosing the right cell line for renal cell cancer research

Contact Info

Product

Resources

About