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The Peptides Mimicking the Third Intracellular Loop of 5-Hydroxytryptamine Receptors of the Types 1B and 6 Selectively Activate G Proteins and Receptor-Specifically Inhibit Serotonin Signaling via the Adenylyl Cyclase System

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Abstract

The third intracellular loop (ICL3) of G protein-coupled receptors has, as a rule, a key role in their interaction with heterotrimeric G proteins. We synthesized peptides corresponding to the C-terminal region of the ICL3 (C-ICL3) of 5-hydroxytryptamine receptors of the type 1B (5-HT1BR) and 6 (5-HT6R) and studied their influence on the functional activity of adenylyl cyclase signaling system (ACSS) in synaptosomal membranes isolated from the rat brain. The 5-HT1BR-peptide ARERKATKTL307–316K-amide mimicking agonist-activated 5-HT1BR reduced forskolin-stimulated adenylyl cyclase (AC) activity and activated pertussis toxin-sensitive G proteins. It lowered inhibitory effects of serotonin and 5-HT1BR-agonists on forskolin-stimulated AC activity and their stimulating effects on GTP binding. This was not the case in the presence of 5-HT1BR-antagonists. The 5-HT6R-peptides mimicking 5-HT6R activated both the basal AC activity and GTP binding of cholera toxin-sensitive G proteins. They lowered the stimulating effect of serotonin and 5-HT6R-agonists on AC and Gs proteins, but in the presence of 5-HT6R-antagonists their action was blocked. Of all the 5-HT6R-peptides with linear and dimeric structure we studied the palmitoylated peptide KHSRKALKASL258–268K(Pal)A-amide had a most pronounced effect both on the basal and 5-HT6R-agonist-stimulated ACSS. The data was obtained indicating that the peptides corresponding to C-ICL3 of 5-HT1BR and 5-HT6R selectively activate Gi and Gs proteins, respectively, and in a receptor-specific manner reduce signal transduction via serotonin-sensitive ACSS in the rat brain. The results of the study give strong evidence in favor of active participation of C-ICL3 of these 5-HTRs in their coupling with the G proteins.

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Abbreviations

AC:

Adenylyl cyclase

ACSS:

Adenylyl cyclase signaling system

t-BOC:

tert-Butyloxycarbonyl group

ChT:

Cholera toxin

EMD-386088:

5-Chloro-2-methyl-3-(1, 2, 3, 6-tetrahydro-4-pyridinyl)-1H-indole

GPCR:

G protein-coupled receptor

GppNHp:

β,γ-Imidoguanosine-5′-triphosphate

Gs, Gi :

G protein of the stimulatory and inhibitory types

5-HT1BR, 5-HT6R:

5-Hydroxytryptamine receptor of the types 1B and 6

ICL:

Intracellular loop

N-ICL3, C-ICL3:

N- and C-terminal regions of the third ICL

8-OH-DPAT:

8-Hydroxy-2-(di-n-propylamino)tetraline

Pal:

Palmitate

PAR:

Protease-activated receptor

PT:

Pertussis toxin

SB-271046:

5-Chloro-N-(4-methoxy-3-(piperazin-1-yl)phenyl)-3-methylbenzo[b]thiophene-2-sulfonamide

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Acknowledgements

This work was supported by Grant No. 09-04-00746 from the Russian Foundation of Basic Research and Program of Russian Academy of Sciences “Fundamental Sciences––Medicine” (2009–2010).

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Authors and Affiliations

  1. Department of Biochemistry, I.M. Sechenov Institute of Evolutionary Physiology and Biochemistry, Russian Academy of Science, Thorez av. 44, 194223, St. Petersburg, Russia

    Alexander O. Shpakov & Kira V. Derkach

  2. Institute of Macromolecular Compounds, Russian Academy of Science, St. Petersburg, Russia

    Elena A. Shpakova, Irina I. Tarasenko & Gennady P. Vlasov

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  1. Alexander O. Shpakov
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  2. Elena A. Shpakova
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  5. Gennady P. Vlasov
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Correspondence to Alexander O. Shpakov.

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Shpakov, A.O., Shpakova, E.A., Tarasenko, I.I. et al. The Peptides Mimicking the Third Intracellular Loop of 5-Hydroxytryptamine Receptors of the Types 1B and 6 Selectively Activate G Proteins and Receptor-Specifically Inhibit Serotonin Signaling via the Adenylyl Cyclase System. Int J Pept Res Ther 16, 95–105 (2010). https://doi.org/10.1007/s10989-010-9208-x

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