Linked Life Data

15469499

Source:http://linkedlifedata.com/resource/pubmed/id/15469499

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
3
pubmed:dateCreated
2004-10-7
pubmed:abstractText
Protein function is often mediated via formation of stable or transient complexes. Here we report the determination of protein-protein interactions in plants using bimolecular fluorescence complementation (BiFC). The yellow fluorescent protein (YFP) was split into two non-overlapping N-terminal (YN) and C-terminal (YC) fragments. Each fragment was cloned in-frame to a gene of interest, enabling expression of fusion proteins. To demonstrate the feasibility of BiFC in plants, two pairs of interacting proteins were utilized: (i) the alpha and beta subunits of the Arabidopsis protein farnesyltransferase (PFT), and (ii) the polycomb proteins, FERTILIZATION-INDEPENDENT ENDOSPERM (FIE) and MEDEA (MEA). Members of each protein pair were transiently co-expressed in leaf epidermal cells of Nicotiana benthamiana or Arabidopsis. Reconstitution of a fluorescing YFP chromophore occurred only when the inquest proteins interacted. No fluorescence was detected following co-expression of free non-fused YN and YC or non-interacting protein pairs. Yellow fluorescence was detected in the cytoplasm of cells that expressed PFT alpha and beta subunits, or in nuclei and cytoplasm of cells that expressed FIE and MEA. In vivo measurements of fluorescence spectra emitted from reconstituted YFPs were identical to that of a non-split YFP, confirming reconstitution of the chromophore. Expression of the inquest proteins was verified by immunoblot analysis using monoclonal antibodies directed against tags within the hybrid proteins. In addition, protein interactions were confirmed by immunoprecipitations. These results demonstrate that plant BiFC is a simple, reliable and relatively fast method for determining protein-protein interactions in plants.
pubmed:commentsCorrections
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Nov
pubmed:issn
0960-7412
pubmed:author
pubmed:issnType
Print
pubmed:volume
40
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
419-27
pubmed:dateRevised
2006-11-15
pubmed:meshHeading
pubmed:year
2004
pubmed:articleTitle
Detection of protein-protein interactions in plants using bimolecular fluorescence complementation.
pubmed:affiliation
Department of Plant Sciences, Tel Aviv University, Tel Aviv 69978, Israel.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't