Turnover of protein phosphorylation evolving under stabilizing selection

Landry, Christian R.; Freschi, Luca; Zarin, Taraneh; Moses, Alan M.

doi:10.3389/fgene.2014.00245

Cited by 48 publications

(52 citation statements)

References 57 publications

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“…In summary, the results presented here showed that the PIP 2 sensitivity of Kv7.2 channels was directly affected by post-translational modification. Phosphorylation of a protein as a reversible process usually acts like an on/off switch at a single phosphorylation site (Landry et al 2014). In contrast, the multiple phosphorylation sites in Kv7.2 channels, some of which were phosphorylated by more than one protein kinase, hinted towards an orchestrated contribution to channel function by several phosphoresidues that were concentrated within a functional motif.…”

Section: Discussionmentioning

confidence: 99%

Phosphorylation regulates the sensitivity of voltage‐gated Kv7.2 channels towards phosphatidylinositol‐4,5‐bisphosphate

Salzer

Erdem

Chen

et al. 2016

The Journal of Physiology

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Key points Phosphatidylinositol‐4,5‐bisphosphate (PIP2) is a key regulator of many membrane proteins, including voltage‐gated Kv7.2 channels.In this study, we identified the residues in five phosphorylation sites and their corresponding protein kinases, the former being clustered within one of four putative PIP2‐binding domains in Kv7.2.Dephosphorylation of these residues reduced the sensitivity of Kv7.2 channels towards PIP2.Dephosphorylation of Kv7.2 affected channel inhibition via M1 muscarinic receptors, but not via bradykinin receptors.Our data indicated that phosphorylation of the Kv7.2 channel was necessary to maintain its low affinity for PIP2, thereby ensuring the tight regulation of the channel via G protein‐coupled receptors. AbstractThe function of numerous ion channels is tightly controlled by G protein‐coupled receptors (GPCRs). The underlying signalling mechanisms may involve phosphorylation of channel proteins and participation of phosphatidylinositol‐4,5‐bisphosphate (PIP2). Although the roles of both mechanisms have been investigated extensively, thus far only little has been reported on their interaction in channel modulation. GPCRs govern Kv7 channels, the latter playing a major role in the regulation of neuronal excitability by determining the levels of PIP2 and through phosphorylation. Using liquid chromatography‐coupled mass spectrometry for Kv7.2 immunoprecipitates of rat brain membranes and transfected cells, we mapped a cluster of five phosphorylation sites in one of the PIP2‐binding domains. To evaluate the effect of phosphorylation on PIP2‐mediated Kv7.2 channel regulation, a quintuple alanine mutant of these serines (S427/S436/S438/S446/S455; A5 mutant) was generated to mimic the dephosphorylated state. Currents passing through these mutated channels were less sensitive towards PIP2 depletion via the voltage‐sensitive phosphatase Dr‐VSP than were wild‐type channels. In vitro phosphorylation assays with the purified C‐terminus of Kv7.2 revealed that CDK5, p38 MAPK, CaMKIIα and PKA were able to phosphorylate the five serines. Inhibition of these protein kinases reduced the sensitivity of wild‐type but not mutant Kv7.2 channels towards PIP2 depletion via Dr‐VSP. In superior cervical ganglion neurons, the protein kinase inhibitors attenuated Kv7 current regulation via M1 receptors, but left unaltered the control by B2 receptors. Our results revealed that the phosphorylation status of serines located within a putative PIP2‐binding domain determined the phospholipid sensitivity of Kv7.2 channels and supported GPCR‐mediated channel regulation.

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Section: Discussionmentioning

confidence: 99%

Phosphorylation regulates the sensitivity of voltage‐gated Kv7.2 channels towards phosphatidylinositol‐4,5‐bisphosphate

Salzer

Erdem

Chen

et al. 2016

The Journal of Physiology

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“…Evolutionary turnover within clusters of phosphorylation sites in disordered regions is thought to be widespread (19,28,(35)(36)(37), and the alignment of Ste50 shows that MAPK consensus sites differ in position, spacing, and number, consistent with evolutionary turnover of these sites within the IDR (Fig. 1B) 124 124 142 188 167 111 132 136 127 131 104 180 163 167 124 128 143 143 143 143 215 216 233 291 274 207 225 225 242 235 168 315 269 276 183 227 254 252 254 254 Diverged Orthologous IDRs Recapitulate Multiple Signaling Functions in S. cerevisiae.…”

Section: Significancementioning

confidence: 99%

“…Stabilizing selection could allow for quantitative phenotypes to be maintained within an optimal range while allowing tolerance of mutations or insertions and deletions, as these individually exert weak functional and selective effects (21,26). Although it is likely that some of these highly diverged IDRs, like noncoding regions, are either nonfunctional or sites of lineage-specific evolution (27), at least a portion of these IDRs may be performing quantitative functions that are under stabilizing selection (28).…”

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confidence: 99%

Selection maintains signaling function of a highly diverged intrinsically disordered region

Zarin

Tsai

et al. 2017

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

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Intrinsically disordered regions (IDRs) are characterized by their lack of stable secondary or tertiary structure and comprise a large part of the eukaryotic proteome. Although these regions play a variety of signaling and regulatory roles, they appear to be rapidly evolving at the primary sequence level. To understand the functional implications of this rapid evolution, we focused on a highly diverged IDR in Saccharomyces cerevisiae that is involved in regulating multiple conserved MAPK pathways. We hypothesized that under stabilizing selection, the functional output of orthologous IDRs could be maintained, such that diverse genotypes could lead to similar function and fitness. Consistent with the stabilizing selection hypothesis, we find that diverged, orthologous IDRs can mostly recapitulate wildtype function and fitness in S. cerevisiae. We also find that the electrostatic charge of the IDR is correlated with signaling output and, using phylogenetic comparative methods, find evidence for selection maintaining this quantitative molecular trait despite underlying genotypic divergence.C urrent predictions suggest that close to 40% of all proteins in eukaryotic organisms are either entirely disordered or contain sizeable regions that are disordered, meaning they do not autonomously fold into defined secondary or tertiary structures (1, 2). These intrinsically disordered regions (IDRs) are thought to have important implications for protein function (3, 4) and are known to play regulatory roles, often through short linear motifs (SLiMs) that control protein-protein interactions, localization, degradation, and posttranslational modifications (5, 6). Although proteome-wide studies have provided in silico evidence for conservation of length (7) and composition (8) in some IDRs, reports of increased rates of insertions and deletions (9-13) and amino acid substitutions (14) in IDRs are indicative of their rapid evolution compared with ordered regions. In addition, although some SLiMs are indeed conserved in IDRs (15-17), others appear in clusters where precise position and number are not conserved (18)(19)(20). Although it is reasonable to assume that conservation of sequence in IDRs is indicative of functional conservation of SLiMs, it is more difficult to interpret the functional consequences of IDRs that are highly diverged at the sequence level: These may represent either nonfunctional sequences evolving in the absence of constraint or weakly constrained functional elements that are gained or lost in a compensatory manner [undergoing evolutionary turnover (as described in refs. 18, 21)], such that they are not conserved at the amino acid sequence level.Like IDRs, noncoding DNA often shows relatively rapid evolution and weak constraints at the sequence level (22). Interestingly, IDRs show other parallels with noncoding DNA (18,23,24). For example, nonconserved clusters of phosphorylation sites in IDRs are reminiscent of nonconserved transcription factor binding sites in enhancers. Although these enhancers and the bi...

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“…The binding interfaces of multimeric proteins can vary substantially among species, sometimes with no overlap at all (4,5). The key amino acid sequences involved in intermolecular cross-talk in signal-transduction systems can evolve at high rates (6,7), and growing evidence suggests that the locations of sites involved in posttranslational modification in individual proteins are under much weaker selective constraints than their absolute numbers (8). Although it is often argued that subtle differences in the motifs involved in intermolecular interactions are molded by the demands of natural selection, seldom has any direct evidence ever been provided in support of such arguments.…”

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confidence: 99%

Evolutionary meandering of intermolecular interactions along the drift barrier

Lynch¹,

Hagner

2014

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

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Many cellular functions depend on highly specific intermolecular interactions, for example transcription factors and their DNA binding sites, microRNAs and their RNA binding sites, the interfaces between heterodimeric protein molecules, the stems in RNA molecules, and kinases and their response regulators in signaltransduction systems. Despite the need for complementarity between interacting partners, such pairwise systems seem to be capable of high levels of evolutionary divergence, even when subject to strong selection. Such behavior is a consequence of the diminishing advantages of increasing binding affinity between partners, the multiplicity of evolutionary pathways between selectively equivalent alternatives, and the stochastic nature of evolutionary processes. Because mutation pressure toward reduced affinity conflicts with selective pressure for greater interaction, situations can arise in which the expected distribution of the degree of matching between interacting partners is bimodal, even in the face of constant selection. Although biomolecules with larger numbers of interacting partners are subject to increased levels of evolutionary conservation, their more numerous partners need not converge on a single sequence motif or be increasingly constrained in more complex systems. These results suggest that most phylogenetic differences in the sequences of binding interfaces are not the result of adaptive fine tuning but a simple consequence of random genetic drift. cellular evolution | molecular interaction | transcription | random genetic drift | coevolution M uch of biology relies on the specificity of intermolecular interactions-the regulation of gene expression, the transmission of information via signal transduction, the assembly of monomeric subunits into multimers, vesicle sorting in eukaryotic cells, toxin-antitoxin systems in microbes, mating-type recognition, and many other cellular features. Although the basic structural features of such fitness-related traits would seem to be under strong purifying selection, molecular specificity often seems to be highly flexible in evolutionary time. For example, transcription-factor binding-site motifs often vary dramatically among orthologous genes in different species and even among similarly regulated genes within the same species (1-3). The binding interfaces of multimeric proteins can vary substantially among species, sometimes with no overlap at all (4, 5). The key amino acid sequences involved in intermolecular cross-talk in signal-transduction systems can evolve at high rates (6, 7), and growing evidence suggests that the locations of sites involved in posttranslational modification in individual proteins are under much weaker selective constraints than their absolute numbers (8). Although it is often argued that subtle differences in the motifs involved in intermolecular interactions are molded by the demands of natural selection, seldom has any direct evidence ever been provided in support of such arguments.The theory provided below makes the case...

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Turnover of protein phosphorylation evolving under stabilizing selection

Cited by 48 publications

References 57 publications

Phosphorylation regulates the sensitivity of voltage‐gated Kv7.2 channels towards phosphatidylinositol‐4,5‐bisphosphate

Phosphorylation regulates the sensitivity of voltage‐gated Kv7.2 channels towards phosphatidylinositol‐4,5‐bisphosphate

Selection maintains signaling function of a highly diverged intrinsically disordered region

Evolutionary meandering of intermolecular interactions along the drift barrier

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