TY - BOOK

T1 - Functional Characterization of CCHamide and Muscarinic Acetylcholine Receptor Signalling in Drosophila melanogaster

AU - Ren, Guilin Robin

PY - 2015

Y1 - 2015

N2 - G-protein coupled receptors (GPCRs) constitute a large and ancient superfamily of membraneproteins responsible for the transduction of extracellular signals to the inside of the cells. In thisPh.D. thesis, Drosophila melanogaster (Dm) was used as a model organism to investigate a numberof topics concerning the pharmacological and the physiological functions of GPCR signalling. Theprimary research in this Ph.D. thesis concerns two topics: (1) functional characterization ofCCHamide-2 signalling and (2) functional characterization of muscarinic acetylcholine receptor(mAChR) signalling.CCHamide-2 is a newly discovered insect peptide hormone. The function of this novel peptide hasnot been well characterised. In this Ph.D. thesis, I identified CCHamide-2 peptides in endocrinecells of the gut and neurones of the brain of larvae and endocrine cells of the gut of adultDrosophila. Behavioural assays in mutants created with the CRISP/Cas9 technique showed thatCCHamide-2 is probly an orexigenic peptide and also that is an important factor for larvaldevelopmental timing.In mammals, muscarinic acetylcholine signalling is involved in the signal transmission of theparasympathetic nervous system. However little is known about muscarinic acetylcholine receptorsignalling in insects. In this study, I found that two types of mAChRs occur in D. melanogaster, onecoupling to Gq (A-type) and the other to Gi (B-type). Both A- and B-type Dm-mAChRs can beactivated by acetylcholine (ACh), but the classical antagonists atropine, scopolamine, and 3-Quinuclidinyl benzilate (QNB) can only block the A-type Dm-mAChRs, while it does not block BtypeDm-mAChRs. Furthermore, the sensitivity to muscarine is 1000× lower in B-type than in AtypeDm-mAChRs. By comparing the intracellular loops-2 and loops-3 of the D.melanogaster, Caenorhabditis elegans, and human mAChRs, I could identify hallmarks for Gi andGq coupling. Appling these hallmarks to other mAChRs from animals with a sequenced genome, Icould predict whether these receptors were Gq and Gi-coupled. In this way, I found that probly allanimals have at least one Gq-coupled and one Gi-coupled mAChR.

AB - G-protein coupled receptors (GPCRs) constitute a large and ancient superfamily of membraneproteins responsible for the transduction of extracellular signals to the inside of the cells. In thisPh.D. thesis, Drosophila melanogaster (Dm) was used as a model organism to investigate a numberof topics concerning the pharmacological and the physiological functions of GPCR signalling. Theprimary research in this Ph.D. thesis concerns two topics: (1) functional characterization ofCCHamide-2 signalling and (2) functional characterization of muscarinic acetylcholine receptor(mAChR) signalling.CCHamide-2 is a newly discovered insect peptide hormone. The function of this novel peptide hasnot been well characterised. In this Ph.D. thesis, I identified CCHamide-2 peptides in endocrinecells of the gut and neurones of the brain of larvae and endocrine cells of the gut of adultDrosophila. Behavioural assays in mutants created with the CRISP/Cas9 technique showed thatCCHamide-2 is probly an orexigenic peptide and also that is an important factor for larvaldevelopmental timing.In mammals, muscarinic acetylcholine signalling is involved in the signal transmission of theparasympathetic nervous system. However little is known about muscarinic acetylcholine receptorsignalling in insects. In this study, I found that two types of mAChRs occur in D. melanogaster, onecoupling to Gq (A-type) and the other to Gi (B-type). Both A- and B-type Dm-mAChRs can beactivated by acetylcholine (ACh), but the classical antagonists atropine, scopolamine, and 3-Quinuclidinyl benzilate (QNB) can only block the A-type Dm-mAChRs, while it does not block BtypeDm-mAChRs. Furthermore, the sensitivity to muscarine is 1000× lower in B-type than in AtypeDm-mAChRs. By comparing the intracellular loops-2 and loops-3 of the D.melanogaster, Caenorhabditis elegans, and human mAChRs, I could identify hallmarks for Gi andGq coupling. Appling these hallmarks to other mAChRs from animals with a sequenced genome, Icould predict whether these receptors were Gq and Gi-coupled. In this way, I found that probly allanimals have at least one Gq-coupled and one Gi-coupled mAChR.

UR - https://soeg.kb.dk/permalink/45KBDK_KGL/fbp0ps/alma99122590385505763

M3 - Ph.D. thesis

BT - Functional Characterization of CCHamide and Muscarinic Acetylcholine Receptor Signalling in Drosophila melanogaster

PB - Department of Biology, Faculty of Science, University of Copenhagen

ER -