Finding prospective partners in the library: the two-hybrid system and phage display find a match

Allen, James B.; Walberg, Mark W.; Edwards, Michael C.; Elledge, Stephen J.

doi:10.1016/s0968-0004(00)89119-7

Cited by 95 publications

(54 citation statements)

References 33 publications

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“…While nonetheless indicating that interactions between D52-like proteins are highly speci®c, additional factors may have contributed towards this result. Firstly, technical limitations associated with the yeast two-hybrid system (Allen et al, 1995) may have prevented the demonstration of interactions between D52-like baits and other interactor types. Secondly, it may be that interactions between D52-like proteins and other partners are too weak and/or transient to be detected using the twohybrid system.…”

Section: Discussionmentioning

confidence: 99%

“…Presented values have, in all cases, been standardized to the mean number of b-galactosidase activity units obtained for three simultaneously-performed, positive control assays (see Materials and methods), which was to 1000 units the hD52, hD53 and hD54 cDNAs had been isolated (Byrne et al, 1995(Byrne et al, , 1996Nourse et al manuscript in preparation). In this way, interacting proteins identi®ed through library screening would be more likely to be co-expressed with D52-like proteins, and thus to represent biological partners for these proteins (Allen et al, 1995).…”

Section: Interactions Between D52-like Proteins Occur Via Their Predimentioning

confidence: 99%

“…The Hf7c HIS3 reporter is less`leaky' than the Y190 HIS3 reporter (Feilotter et al, 1994;Durfee et al, 1993), and the HIS3 competitor 3-amino-1, 2, 4-triazole is therefore not required to be added to selective media to suppress a basal level of HIS3 expression (Feilotter et al, 1994). Thus, weaker interactions with bait proteins are more likely to be detected (Allen et al, 1995). Screening approximately 44 000 c.f.u.…”

Section: Interactions Between D52-like Proteins Occur Via Their Predimentioning

confidence: 99%

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Identification of homo- and heteromeric interactions between members of the breast carcinoma-associated D52 protein family using the yeast two-hybrid system

et al. 1998

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Section: Discussionmentioning

confidence: 99%

Section: Interactions Between D52-like Proteins Occur Via Their Predimentioning

confidence: 99%

Section: Interactions Between D52-like Proteins Occur Via Their Predimentioning

confidence: 99%

See 1 more Smart Citation

Identification of homo- and heteromeric interactions between members of the breast carcinoma-associated D52 protein family using the yeast two-hybrid system

et al. 1998

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“…The two hybrid system, with its constant improvements during the last decade, provides a very powerful method for the analysis of protein-protein interaction. Nevertheless, it exhibits several limitations and problems (4)(5)(6)(7)(8)(9)(10).…”

Section: Introductionmentioning

confidence: 99%

A novel in vivo assay for the analysis of protein-protein interaction

Maroun¹

1999

Nucleic Acids Research

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The Ras Recruitment System (RRS) is a method for identification and isolation of protein-protein interaction. The method is based on translocation of cytoplasmic mammalian Ras protein to the inner leaflet of the plasma membrane through protein-protein interaction. The system is studied in a temperaturesensitive yeast strain where the yeast Ras guanyl nucleotide exchange factor is inactive at 36°°°°C. Protein-protein interaction results in cell growth at the restrictive temperature. We developed a gene reporter assay for the analysis of protein-protein interaction in mammalian cells. Ras activation in mammalian cells induces the mitogen-activated kinase cascade (MAPK), which can be monitored using Ras-dependent reporter genes. This greatly extends the usefulness of the system and provides a novel assay for protein-protein interaction in mammalian cells.

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“…T he yeast one-hybrid system has been used as a tool for identifying proteins that bind to specific DNA sequences (1,2). The upstream activating sequence of a reporter gene is replaced by a bait sequence, and DNA-binding proteins are then expressed in yeast cells as fusion proteins with a transcriptional activation domain (AD).…”

mentioning

confidence: 99%

Z-DNA-binding proteins can act as potent effectors of gene expression in viv o

Kim²,

Rich³

2002

Proc. Natl. Acad. Sci. U.S.A.

133

102

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The role of Z-DNA-binding proteins in vivo is explored in yeast. A conformation-specific yeast one-hybrid system is made in which formation of Z-DNA is studied near a minimal promoter site where it can be stabilized by negative supercoiling in addition to protein binding. Experiments were carried out with a Z-DNA-binding protein domain from the editing enzyme, double-stranded RNA adenosine deaminase 1. In the one-hybrid system, the reporter gene is activated when a Z-DNA-specific binding domain is fused with an activation domain and expressed in vivo. Significantly, it was found that even in the absence of the activation domain there is substantial transcription of the reporter gene if the Z-DNAbinding protein is expressed in the cell. This result suggests that Z-DNA formation in the promoter region induced or stabilized by a Z-DNA-binding protein can act as a cis-element in gene regulation. Related results have been found recently when the human chromatin-remodeling system converts a segment of DNA in the promoter region of the human colony-stimulating factor 1 gene into the left-handed Z-conformation. The yeast one-hybrid system has been used as a tool for identifying proteins that bind to specific DNA sequences (1, 2). The upstream activating sequence of a reporter gene is replaced by a bait sequence, and DNA-binding proteins are then expressed in yeast cells as fusion proteins with a transcriptional activation domain (AD). Thus, transcription occurs only if the DNA-binding protein fused to the AD interacts with the bait DNA sequence. We have expanded this system to include DNA conformational specificity focusing on left-handed Z-DNA formation. This assay allows us to identify and characterize Z-DNAspecific binding proteins in vivo and study the influence of Z-DNA formation on transcriptional activity.Z-DNA is a left-handed form of the double helix, as revealed in a single-crystal x-ray analysis of duplex d(CGCGCG) (3). The Watson-Crick base pairs in Z-DNA have ''flipped over'' relative to their orientation in B-DNA, resulting in the guanine residues adopting the syn conformation, whereas the cytosine residues remain in the anti conformation. In addition, it produced a zig-zag arrangement of the sugar phosphate backbone, giving rise to the name Z-DNA. Because purines adopt the syn conformation more readily than pyrimidines, Z-DNA formation is favored in sequences with alternations of purines and pyrimidines. The most favored sequence consists of (dC-dG) n although many other sequences can also adopt the Z conformation (4). Although Z-DNA is a higher energy conformation than righthanded B-DNA, it was realized that B-to Z-DNA conversions could occur in vivo when it was discovered that Z-DNA is stabilized by negative supercoiling (5). Formation of Z-DNA removes negative supercoiling, and the energy of supercoiling stabilizes the Z conformation. Movement of RNA polymerase generates negative torsional strain behind it (6), and Z-DNA is maintained near promoter regions in mammalian nuclei as long as the genes are a...

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Finding prospective partners in the library: the two-hybrid system and phage display find a match

Cited by 95 publications

References 33 publications

Identification of homo- and heteromeric interactions between members of the breast carcinoma-associated D52 protein family using the yeast two-hybrid system

Identification of homo- and heteromeric interactions between members of the breast carcinoma-associated D52 protein family using the yeast two-hybrid system

A novel in vivo assay for the analysis of protein-protein interaction

Z-DNA-binding proteins can act as potent effectors of gene expression in viv o

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