Universität zu Köln

Abu Obaid, Asmaa (2022). Deciphering the function and evolutionary context of a novel G alpha protein, Gv. PhD thesis, Universität zu Köln.

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Abstract

Heterotrimeric G proteins act as molecular switches inside cells and control a wide range of systematic functions i.e., embryonic development, learning, homeostasis, etc. Loss or gain of function mutations in G alpha proteins result in signalling defects that lead to pleiotropic manifestations; in addition, they are outstanding pharmaceutical targets. Despite the importance of the G alpha subunits in all biological systems, so far there have been no published data about the function of the most recently discovered G alpha family, Gv. Up to now, nothing is known about its role, effector proteins, and signalling pathway, despite Gv being present in zebrafish, a popular vertebrate model organism. Thereby, the purpose of this study is to decipher the so far uncharacterized Gv protein role, particularly in zebrafish. Moreover, the present study was also designed to examine Gv evolutionary origin and dynamics in relation to the other four G alpha families, since only scarce and partially inconsistent information was available for the birth of Gv in the earliest-diverging, unicellular Holozoa, the Teretosporea and no systematic study of its evolutionary dynamic in relation to the other families has been performed. Firstly, we investigated the ontogenetic and tissue-specific expression of Gv mRNA during embryogenesis and larval development by the histochemical technique wholemount in situ hybridization (WISH). Secondly, we evaluated phenotypic and molecular changes in gnav1-/- mutants with an early 13 bp deletion in the mutant gnav1. Thirdly, to address the evolutionary origin and dynamics of Gv, also in relation to the other four families, we constructed phylogenetic trees by rigorously searching the five G alpha families in NCBI genomic databases of nearly a thousand species from many major phylogenetic subdivisions in Holozoa. Our data show for the first time that gnav1 exhibits broad expression in early embryonic stages starting from gastrulation and becomes confined to specific tissues in larval stages. These expression patterns indicate manifold functions for Gv in embryonic organogenesis including sensory and osmoregulatory organs, cartilaginous tissue and muscle. We also demonstrate altered phenotypes in gnav1-/- mutants that include: the number of eggs produced per clutch is markedly decreased; the mutant embryos hatch significantly earlier than the wildtype siblings; three and five dpf mutants show craniofacial defects and overall decrease in alcian blue staining; five and 11 dpf mutant larvae show prominently reduced mineralization in the craniofacial bones. The 5 dpf mutant larvae exhibit highly significant reduction in calcium, magnesium, and potassium levels. The mutant kidneys show significantly increased mRNA levels of NCC and NKA.5 which play a role in calcium and magnesium homeostasis. Taken together, our data suggest novel functions of Gv in cation homeostasis, and subsequently bone and cartilage formation, hatching process, and oviposition. Our phylogenetic studies reveal the birth of the Gv family in the common ancestor of all Holozoa, i.e., before the divergence into Teretosporea and Filozoa. Moreover, we show that Gv evolutionary dynamics are within the range of those observed for the other four G alpha classes. Furthermore, we have identified two novel G alpha clades, i.e., Gf and Gw, which have not been described before.

Item Type:	Thesis (PhD thesis)
Creators:	Creators Email ORCID ORCID Put Code Abu Obaid, Asmaa asmaabuobied@yahoo.com UNSPECIFIED UNSPECIFIED
URN:	urn:nbn:de:hbz:38-617066
Date:	31 May 2022
Language:	English
Faculty:	Faculty of Mathematics and Natural Sciences
Divisions:	Faculty of Mathematics and Natural Sciences > Department of Biology > Institute for Genetics
Subjects:	Natural sciences and mathematics
Uncontrolled Keywords:	Keywords Language G protein coupled receptors English G alpha proteins English gnav1 English Gv protein English zebrafish English G protein signaling English
Date of oral exam:	31 May 2022
Referee:	Name Academic Title Korsching, Sigrun Prof. Dr.
Refereed:	Yes
URI:	http://kups.ub.uni-koeln.de/id/eprint/61706