Quantitative FRET imaging of leptin receptor oligomerization kinetics in single cells

Biener, Eva; Charlier, Madia; Ramanujan, V. Krishnan; Daniel, Nathalie; Eisenberg, Avital; Bjørbæk, Christian; Herman, Brian; Gertler, Arieh; Djiane, Jean

doi:10.1042/bc20040511

Cited by 42 publications

(42 citation statements)

References 51 publications

(64 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The fluorescence emission was directed back out into the detection unit through a short-pass filter (l < 750 nm). The FLIM unit was composed of a streak camera (Streakscope C4334; Hamamatsu Photonics) coupled to a fast and high-sensitivity CCD camera (model C8800-53C; Hamamatsu) (Krishnan et al, 2003;Biener et al, 2005). For each nucleus, average fluorescence decay profiles were plotted and lifetimes were estimated by fitting data with biexponential function using a nonlinear least squares estimation procedure with Origin 7.5 software (OriginLab).…”

Section: Flim and Data Analysismentioning

confidence: 99%

RD19, anArabidopsisCysteine Protease Required for RRS1-R–Mediated Resistance, Is Relocalized to the Nucleus by theRalstonia solanacearumPopP2 Effector

et al. 2008

View full text Add to dashboard Cite

Bacterial wilt, a disease impacting cultivated crops worldwide, is caused by the pathogenic bacterium Ralstonia solanacearum. PopP2 (for Pseudomonas outer protein P2) is an R. solanacearum type III effector that belongs to the YopJ/AvrRxv protein family and interacts with the Arabidopsis thaliana RESISTANT TO RALSTONIA SOLANACEARUM 1-R (RRS1-R) resistance protein. RRS1-R contains the Toll/Interleukin1 receptor-nucleotide binding site-Leu-rich repeat domains found in several cytoplasmic R proteins and a C-terminal WRKY DNA binding domain. In this study, we identified the Arabidopsis Cys protease RESPONSIVE TO DEHYDRATION19 (RD19) as being a PopP2-interacting protein whose expression is induced during infection by R. solanacearum. An Arabidopsis rd19 mutant in an RRS1-R genetic background is compromised in resistance to the bacterium, indicating that RD19 is required for RRS1-R-mediated resistance. RD19 normally localizes in mobile vacuole-associated compartments and, upon coexpression with PopP2, is specifically relocalized to the plant nucleus, where the two proteins physically interact. No direct physical interaction between RRS1-R and RD19 in the presence of PopP2 was detected in the nucleus as determined by Fö rster resonance energy transfer. We propose that RD19 associates with PopP2 to form a nuclear complex that is required for activation of the RRS1-R-mediated resistance response.

show abstract

Section: Flim and Data Analysismentioning

confidence: 99%

RD19, anArabidopsisCysteine Protease Required for RRS1-R–Mediated Resistance, Is Relocalized to the Nucleus by theRalstonia solanacearumPopP2 Effector

et al. 2008

View full text Add to dashboard Cite

show abstract

“…Homodimerization of unstimulated leptin b receptors yields FRET efficiencies of 4.9% by the acceptor photobleaching method, the same method used in this study (Biener et al, 2005). Homodimers are expected to yield FRET signals that are half the intensity of heterodimers, resulting from the unavoidable formation of YFP-␤3/YFP-␤3 dimer and CFP-␤3/CFP-␤3 dimers that do not yield FRET signals upon acceptor photobleaching.…”

Section: Discussionmentioning

confidence: 89%

Phospholipase C-β3 and -β1 Form Homodimers, but Not Heterodimers, through Catalytic and Carboxyl-Terminal Domains

Zhang¹,

Vogel²,

McCullar³

et al. 2006

Mol Pharmacol

View full text Add to dashboard Cite

Phospholipase C-␤ (PLC-␤) isoenzymes are key effectors in G protein-coupled signaling pathways. Prior research suggests that some isoforms of PLC-␤ may exist and function as dimers. Using coimmunoprecipitation assays of differentially tagged PLC-␤ constructs and size-exclusion chromatography of native PLC-␤, we observed homodimerization of PLC-␤3 and PLC-␤1 isoenzymes but failed to detect heterodimerization of these isoenzymes. Size-exclusion chromatography data suggest that PLC-␤3 and PLC-␤1 form higher affinity homodimers than PLC-␤2. Evidence supportive of limited PLC-␤ monomerhomodimer equilibrium appears at Յ100 nM. Further assessment of homodimerization status by coimmunoprecipitation assays with differentially tagged PLC-␤3 fragments demonstrated that at least two subdomains of PLC-␤3 are involved in dimer formation, one in the catalytic X and Y domains and the other in the G protein-regulated carboxyl-terminal domain. In addition, we provide evidence consistent with the existence of PLC-␤ homodimers in a whole-cell context, using fluorescent protein-tagged constructs and microscopic fluorescence resonance energy transfer assays.Phospholipase C (PLC) is a key effector enzyme in multiple cellular signaling pathways, hydrolyzing phosphatidylinositol 4,5-bisphosphate to inositol 1,4,5-triphosphate and diacylglycerol, second messengers that subsequently regulate Ca 2ϩ release from intracellular stores and protein kinase C activation. PLC isozymes are grouped into five families based on structure and regulatory mechanism: PLC-␤, PLC-␥, PLC-␦, PLC-, and PLC-. The PLC-␤ family exists as four known homologs in mammals, ␤1-␤4, all of which are involved in G protein-coupled receptor-mediated signaling cascades (Rhee, 2001;Saunders et al., 2002). Of the four isoenzymes, PLC-␤1 and PLC-␤3 are widely distributed and are often found coexpressed in a variety of tissues, including brain, liver, uterus, parotid gland, lung, platelets, and heart (Hansen et al., 1995;Fukami, 2002).PLC-␤ isoenzymes contain sequences with homology to several characterized protein structural motifs commonly found in membrane associated, Ca 2ϩ and lipid binding proteins. These structural domains include pleckstrin homology (PH), EF hand, and C2 homology regions, as well as a PDZbinding domain at the C terminus. Highly conserved among PLC family members are two domains required for catalytic activity, designated X and Y, that structurally form a triose phosphate isomerase barrel (TIM) to create the catalytic pocket in the crystal form of PLC-␦ (Rhee, 2001).Unique among PLC family members, PLC-␤ isoenzymes contain a carboxyl-terminal (C-tail) domain of approximately 400 amino acids that is involved in membrane association and regulation by G␣ q subunits. Crystallization of the isolated C-tail domain of a turkey PLC-␤ revealed a structure of three ␣-helices forming a coiled coil that associates as an antiparallel homodimer along its long axis (Singer et al., 2002). Size-exclusion chromatography data suggested that purified, concentrated, f...

show abstract

“…Earlier studies suggested ligand-independent dimerization or oligomerization of the LR ECD in solution , but recent evidence obtained by multi-angle laser light scattering (MALLS) and smallangle X-ray scattering (SAXS) experiments has now indicated that it is monomeric in the absence of leptin (Mancour et al 2012, Moharana et al 2014. On the membrane, however, the LR can assemble as preformed dimers or oligomers, as evidenced by a high basal signal in the absence of leptin in both fluorescence resonance energy transfer (FRET) and bioluminescence resonance energy transfer (BRET) between differently tagged LRs and by co-immunoprecipitation of tagged LRs (Nakashima et al 1997, White & Tartaglia 1999, Couturier & Jockers 2003, Biener et al 2005. Different isoforms of the LR can heterodimerize with each other both in the presence and absence of leptin (White & Tartaglia 1999, Bacart et al 2010.…”

Section: Disulfide Network and Glycosylation Of The Ecdmentioning

confidence: 99%

“…Different isoforms of the LR can heterodimerize with each other both in the presence and absence of leptin (White & Tartaglia 1999, Bacart et al 2010. Leptin treatment induces an increase in BRET and FRET signals, which are probably a consequence of reorganization within the pre-formed complexes and/or de novo oligomerization (Couturier & Jockers 2003, Biener et al 2005.…”

Section: Disulfide Network and Glycosylation Of The Ecdmentioning

confidence: 99%

20 YEARS OF LEPTIN: Insights into signaling assemblies of the leptin receptor

Peelman¹,

Zabeau²,

Moharana³

et al. 2014

Journal of Endocrinology

View full text Add to dashboard Cite

Leptin plays a central role in the control of body weight and energy homeostasis, but is a pleiotropic cytokine with activities on many peripheral cell types. In this review, we discuss the interaction of leptin with its receptor, and focus on the structural and mechanistic aspects of the extracellular aspects of leptin receptor (LR) activation. We provide an extensive overview of all structural information that has been obtained for leptin and its receptor via X-ray crystallography, electron microscopy, small-angle X-ray scattering, homology modeling, and mutagenesis studies. The available knowledge is integrated into putative models toward a recapitulation of the LR activation mechanism.

show abstract

Quantitative FRET imaging of leptin receptor oligomerization kinetics in single cells

Cited by 42 publications

References 51 publications

RD19, anArabidopsisCysteine Protease Required for RRS1-R–Mediated Resistance, Is Relocalized to the Nucleus by theRalstonia solanacearumPopP2 Effector

RD19, anArabidopsisCysteine Protease Required for RRS1-R–Mediated Resistance, Is Relocalized to the Nucleus by theRalstonia solanacearumPopP2 Effector

Phospholipase C-β3 and -β1 Form Homodimers, but Not Heterodimers, through Catalytic and Carboxyl-Terminal Domains

20 YEARS OF LEPTIN: Insights into signaling assemblies of the leptin receptor

Contact Info

Product

Resources

About