The Phospholipase C Signalling System: Study of the role of the G₀ protein and identification of a novel variant of Phospholipase C β4

De Filippis, Lidia (2003). The Phospholipase C Signalling System: Study of the role of the G₀ protein and identification of a novel variant of Phospholipase C β4. PhD thesis The Open University.

DOI: https://doi.org/10.21954/ou.ro.0000f6f8

Abstract

The thesis focuses on the properties and regulation of phospholipase C β (PLC β). Studies concerning the effect of Gαo subunit on PLC activity revealed that mutationally activated Gαo inhibits PLC activity in mouse Ltk clones. The observed effect seemed to be specific of Gαo, as either wild-type Gαo or the wild-type and mutationally activated Gαil subunits were not able to affect receptor-mediated activation of PLC. Known second messengers as Ca2+ and cAMP were not involved in this inhibitory process since they were not able to mimic or prevent the Gαo effect on PLC. Transient transfection experiments in COS7 cells did not reproduce the same results; therefore, either a direct or an indirect mechanism of PLC activity regulation by Gαo remain feasible hypothesis. The thesis also presents results concerning the identification of a novel C-terminal variant of PLC β4 (PLC β4c), that has a 37-nucleotide insertion which results in replacement of 22 aminoacid residues at the carboxyl terminal tail of PLC β4a with 41 unique residues. RT-PCR and Western Blot experiments did show that, in contrast with the known enzyme (PLC β4c) detectable only in brain, PLC β4c is present in several tissues and cell lines. Expression of full-length cDNAs in COS7 cells revealed that the two forms of PLC β are associated with the particulate fraction and equally responsive to G protein subunits. However, the different carboxyl terminal tails of PLC β4 isoforms may allow for differential targeting and subcellular localization, contributing to regulation of PLC β4 - mediated specific pathways. Even the presence of a PDZ domain binding sequence in PLC β4a but not in PLC β4c suggests functional differences between the two variants.

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