Rhodopsin Arginine-135 Mutants Are Phosphorylated by Rhodopsin Kinase and Bind Arrestin in the Absence of 11-<i>cis</i>-Retinal

Shi, Wen; Sports, Charlene D.; Raman, Dayanidhi; Shirakawa, Satoko; Osawa, Shôji; Weiss, Ellen R.

doi:10.1021/bi9731100

Cited by 45 publications

(38 citation statements)

References 29 publications

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“…Both complexes appear to involve highly-conserved R3.50, which is also a key residue during binding and activation of the G protein α-subunit (46). However, the binding spectrum of the deprotonated complex indicates that R* exhibits a more open and more flexible conformation than R*H + .…”

Section: Discussionmentioning

confidence: 95%

The arrestin-1 finger loop interacts with two distinct conformations of active rhodopsin

Elgeti

Kazmin

Rose

et al. 2018

Journal of Biological Chemistry

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Signaling of the prototypical G protein coupled receptor (GPCR) rhodopsin through its cognate G protein transducin (G t ) is quenched when arrestin binds to the activated receptor. Although the overall architecture of the rhodopsin-arrestin complex is known, many questions regarding its specificity remain unresolved. Here, using FTIR difference spectroscopy and a dual pH/ peptide titration assay, we show that rhodopsin maintains certain flexibility upon binding the "finger loop" of visual arrestin (prepared as synthetic peptide ArrFL-1). We found that two distinct complexes can be stabilized depending on the protonation state of E3.49 in the conserved (D)ERY motif. Both complexes exhibit different interaction modes and affinities of ArrFL-1 binding. The plasticity of the receptor within the rhodopsin/ArrFL-1 complex stands in contrast to the complex with the C-terminus of the G t α-subunit (GαCT), which stabilizes only one specific substate out of the conformational ensemble. However, GαCT binding and both ArrFL-1 binding modes involve a direct interaction to conserved R3.50, as determined by site-directed mutagenesis. Our findings highlight the importance of receptor conformational flexibility and cytoplasmic proton uptake for modulation of rhodopsin signaling and thereby extend the picture provided by crystal structures of the rhodopsin/arrestin and rhodopsin/ArrFL-1 complexes. Furthermore, the two binding modes of ArrFL-1 identified here involve motifs of conserved amino acids, which indicates that our results may have elucidated a common modulation mechanism of class A GPCR-G protein/-arrestin signaling.Unlike other class A GPCRs the photoreceptor rhodopsin contains the light-sensitive cofactor retinal as a ligand, which is covalently bound to the opsin apoprotein. Upon light-induced cis/trans isomerization of retinal and subsequent alterations within the binding pocket, the signal propagates via GPCR-conserved interhelical networks towards the cytoplasmic surface of the receptor, where major structural rearrangements take place. The most prominent structural change is the rotational outward tilt of transmembrane (TM) helix 6, which opens a cytoplasmic binding crevice and thus constitutes the main prerequisite for binding and activation of signaling via the G protein transducin (1, 2). Even though first identified in rhodopsin, for many other GPCRs homologous structural changes have been shown to exist, and a common structural and functional framework for GPCRs is being developed (3).

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Section: Discussionmentioning

confidence: 95%

The arrestin-1 finger loop interacts with two distinct conformations of active rhodopsin

Elgeti

Kazmin

Rose

et al. 2018

Journal of Biological Chemistry

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“…For example, rhodopsin R135L mutants appear to be phosphorylated by rhodopsin kinase and bound to arrestin in vitro (Shi et al, 1998). When the R135L mutant is introduced into murine retina by in vivo electroporation, it appears to cause arrestin to accumulate around the cell body, suggesting that R135L/arrestin complex may form within these cellular compartments (Chuang et al, 2004).…”

Section: Discussionmentioning

confidence: 99%

Stable Rhodopsin/Arrestin Complex Leads to Retinal Degeneration in a Transgenic Mouse Model of Autosomal Dominant Retinitis Pigmentosa

Chen

Shi²,

Concepcion³

et al. 2006

J. Neurosci.

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Over 100 rhodopsin mutation alleles have been associated with autosomal dominant retinitis pigmentosa (ADRP). These mutations appear to cause photoreceptor cell death through diverse molecular mechanisms. We show that K296E, a rhodopsin mutation associated with ADRP, forms a stable complex with arrestin that is toxic to mouse rod photoreceptors. This cell death pathway appears to be conserved from flies to mammals. A genetics approach to eliminate arrestin unmasked the constitutive activity of K296E and caused photoreceptor cell death through a transducin-dependent mechanism that is similar to light damage. Expressing K296E in the arrestin/ transducin double knock-out background prevented transducin signaling and led to substantially improved retinal morphology but did not fully prevent cell death caused by K296E. The adverse effect of K296E in the arrestin/transducin knock-out background can be mimicked by constant exposure to low light. Furthermore, we found that arrestin binding causes K296E to mislocalize to the wrong cellular compartment. Accumulation of stable rhodopsin/arrestin complex in the inner segment may be an important mechanism for triggering the cell death pathway in the mammalian photoreceptor cell.

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“…The two residues in the intrahelical interaction between Glu-134 and Arg-135 appear to have different roles as concluded from the fact that many substitutions of Glu-134 (Gln, Ile or Ser) did not reduce activity [86]. The importance of Arg-135 in nucleotide exchange catalysis was derived from the severe effect on transducin activation by the R135A, R135L, R135G and E134R/R135E mutations [86,125,162]. In conclusion, the Glu-134/Arg-135 charge-pair, which is located near the cytoplasmic surface, is an important element in regulating the active cytoplasmic conformation.…”

Section: Constitutively Active Mutants Of Rhodopsinmentioning

confidence: 99%

Crystal structure of the ligand-free G-protein-coupled receptor opsin

Park

Scheerer

Hofmann

et al. 2008

Nature

851

925

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Compared with the known structure of inactive rhodopsin, opsin displays prominent structural changes in the conserved E(D)RY and NPxxY(x) 5,6 F regions and TM5-TM7. At the cytoplasmic side, TM6 is tilted outwards by 6-7 Å, whereas the helix structure of TM5 is more elongated and close to TM6. These structural changes, of which some are attributed to an active GPCR state, reorganize the empty retinal binding pocket to disclose two openings for exit and entry of retinal. The opsin structure thus sheds new light on binding of hydrophobic ligands to GPCRs, GPCR activation and signal transfer to the G protein.

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Rhodopsin Arginine-135 Mutants Are Phosphorylated by Rhodopsin Kinase and Bind Arrestin in the Absence of 11-cis-Retinal

Cited by 45 publications

References 29 publications

The arrestin-1 finger loop interacts with two distinct conformations of active rhodopsin

The arrestin-1 finger loop interacts with two distinct conformations of active rhodopsin

Stable Rhodopsin/Arrestin Complex Leads to Retinal Degeneration in a Transgenic Mouse Model of Autosomal Dominant Retinitis Pigmentosa

Crystal structure of the ligand-free G-protein-coupled receptor opsin

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