FGFR3 Dimer Stabilization Due to a Single Amino Acid Pathogenic Mutation

Li, Edwin; You, Min; Hristova, Kalina

doi:10.1016/j.jmb.2005.11.077

Cited by 98 publications

(222 citation statements)

References 53 publications

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“…For probes restricted to membranes, FRET between unbound diffusible donors and acceptors is more likely (34 -36) and has been quantified by multiple approaches (2,37). For the present study, the minimum FRET observed between YFP-and mCherry-PLB in the presence of a large excess of CFP-PLB competitor (Fig.…”

Section: Discussionmentioning

confidence: 87%

Phospholamban Oligomerization, Quaternary Structure, and Sarco(endo)plasmic Reticulum Calcium ATPase Binding Measured by Fluorescence Resonance Energy Transfer in Living Cells

Kelly

Hou

Bossuyt

et al. 2008

Journal of Biological Chemistry

116

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Phospholamban (PLB) oligomerization, quaternary structure, and sarco(endo)plasmic reticulum calcium ATPase (SERCA) binding were quantified by fluorescence resonance energy transfer (FRET) in an intact cellular environment. FRET between cyan fluorescent protein-PLB and yellow fluorescent protein-PLB in AAV-293 cells showed hyperbolic dependence on protein concentration, with a maximum efficiency of 45.1 ؎ 1.3%. The observed FRET corresponds to a probe separation distance of 58.7 ؎ 0.5 Å , according to a computational model of intrapentameric FRET. This is consistent with models of the PLB pentamer in which cytoplasmic domains fan out from the central bundle of transmembrane helices. An I40A mutation of PLB did not alter pentamer conformation but increased the concentration of half-maximal FRET (K D ) by >4-fold. This is consistent with the previous observation that this putatively monomeric mutant still oligomerizes in intact membranes but forms more dynamic pentamers than wild type PLB. PLB association with SERCA, measured by FRET between cyan fluorescent protein-SERCA and yellow fluorescent protein-PLB, was increased by the I40A mutation without any detectable change in probe separation distance. The data indicate that the regulatory complex conformation is not altered by the I40A mutation. A naturally occurring human mutation (L39Stop) greatly reduced PLB oligomerization and SERCA binding and caused mislocalization of PLB to the cytoplasm and nucleus. Overall, the data suggest that the PLB pentamer adopts a "pinwheel" shape in cell membranes, as opposed to a more compact "bellflower" conformation. I40A mutation decreases oligomerization and increases PLB binding to SERCA. Truncation of the transmembrane domain by L39Stop mutation prevents anchoring of the protein in the membrane, greatly reducing PLB binding to itself or its regulatory target, SERCA.The 52-amino acid protein phospholamban (PLB) 2 is an important regulator of cardiac calcium handling (1). PLB binds avidly (2) but reversibly (3) to the sarco(endo)plasmic reticulum calcium ATPase (SERCA), reducing this calcium pump's affinity for calcium (4). PLB also oligomerizes into pentamers through leucine zipper interactions in its transmembrane domain (5, 6). NMR studies indicate that PLB tertiary structure consists of an N-terminal (cytosolic) ␣-helix (domain IA) connected by a flexible loop (domain IB) to a second ␣-helix (domain II) that is anchored in the membrane of the sarcoplasmic reticulum (7). Several possible configurations of the cytoplasmic domains of pentameric PLB have been described. Some of these portray the domain IA ␣-helix as being nearly normal to the surface of the membrane (8, 9), whereas others indicate axial declination of domain IA (7, 10 -14), permitting contact with the surface of the membrane (15, 16). These previous studies were performed using in vitro preparations of PLB in defined lipids or detergent. To investigate the quaternary structure of PLB in the membranes of living cells and distinguish between these structural mo...

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Section: Discussionmentioning

confidence: 87%

Phospholamban Oligomerization, Quaternary Structure, and Sarco(endo)plasmic Reticulum Calcium ATPase Binding Measured by Fluorescence Resonance Energy Transfer in Living Cells

Kelly

Hou

Bossuyt

et al. 2008

Journal of Biological Chemistry

116

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“…On the other hand, the A393E mutation increases the fraction of FGFR3 dimers resulting from dimer stabilization, measured by the change in the free energy of dimerization. 35,47 In addition, BernardPierrot et al 46 demonstrated that FGFR3b-S249C was able to transform NIH-3T3 cells, inducing their anchorage-independent growth and tumor transformation when injected subcutaneously into nude mice, with the transforming properties classically attributed to oncogenes.…”

Section: Discussionmentioning

confidence: 99%

FGFR3 mutations in prostate cancer: association with low-grade tumors

et al. 2009

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Prostate cancer is the second cause of cancer-related death in men of the Western World. The role of FGFR3 and its abnormalities in prostate cancer are not known. FGFR3 mutations have been reported in some human tumors. Few studies have analyzed the mutations of FGFR3 in prostate tumors, and no mutations have been previously reported. Prevalence of FGFR3 somatic mutations was investigated in a series of prostate tumors. The presence of other tumors in these patients, including urothelial, skin, colon, and lung neoplasms, was recorded. Mutational analysis of exons 7, 10, and 15 of FGFR3 revealed 9 mutations in the 112 prostate tumors studied (8%). Most of them consisted of the missense change S249C. The prevalence of mutations in tumors with combined Gleason score ¼ 6 is 18% (8/45) compared to 3% (1/36) for tumors with grade ¼ 7, and 0% (0/31) for those with grade Z8 and metastases (P ¼ 0.007). The frequency of FGFR3 mutations in autopsy and biopsy samples was 6 and 9%, respectively. The prevalence of FGFR3 mutations in prostate tumors from patients with only prostate cancer was 2% compared to 23% in prostate tumors from patients with other associated neoplasms (P ¼ 0.001). This is the first report of molecular changes of FGFR3 in prostate cancer. This gene does not seem to be central to the pathogenesis of prostate cancer, but it is significantly associated with a subgroup of low-grade prostate tumors, and with the finding of other tumors, mainly arising in bladder and skin.

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“…The measured effect of the order of -1 kcal/mole is similar to results obtained for a model system comprising the isolated FGFR3 TM domains in bilayers of different composition, demonstrating that the Ala391Glu mutation stabilizes the FGFR3 TM domain dimer by -1.3 ± 0.2 kcal/mole. 41 In addition, computational and experimental studies of the structure of the isolated Neu/ErbB2 and FGFR3 TM domain dimers demonstrate dimer stabilization via Glu-mediated hydrogen bonding. 9,12,14,[41][42][43] Taken together, these results strongly suggest that hydrogen bonds stabilize the mutant Neu/Val664Glu and Neu_FGFR3/Ala391Glu receptors in mammalian cells, a mechanism that was originally proposed 18 years ago, 42 but has been debated ever since.…”

Section: Structure Of Rtk Tm Dimersmentioning

confidence: 99%

“…41 In addition, computational and experimental studies of the structure of the isolated Neu/ErbB2 and FGFR3 TM domain dimers demonstrate dimer stabilization via Glu-mediated hydrogen bonding. 9,12,14,[41][42][43] Taken together, these results strongly suggest that hydrogen bonds stabilize the mutant Neu/Val664Glu and Neu_FGFR3/Ala391Glu receptors in mammalian cells, a mechanism that was originally proposed 18 years ago, 42 but has been debated ever since. Furthermore, the results suggest that an increase of the order of -1 kcal/mole may be sufficient to transform normal RTK signaling processes into pathogenic processes.…”

Section: Structure Of Rtk Tm Dimersmentioning

confidence: 99%