Molecular and Functional Characterisation of Sea Urchin ADP-ribosyl Cyclases
Lead Research Organisation:
University College London
Department Name: Physiology
Abstract
All cells of the body are physically separated from their surrounding environment by a membrane. In order for cells to respond to factors in the blood (such as hormones) which normally can not pass through the membrane, special 'messenger' molecules are used that are generated inside the cell upon contact of the stimulus with the cell. Two of these messengers called cADPR and NAADP are produced by a single family of enzymes / the ADP-ribosyl cyclases. Once made, cADPR and NAADP cause large increases in the concentration of calcium within the cell which in turn are important for many processes including for example the nerve function. Understanding how these enzymes work then is important to understand how cells work. In this study we will determine the DNA sequences of the genes for ADP-ribosyl cyclases in the sea urchin a well-studied experimental system. Having this information will allow us to produce large quantities of each protein to study their properties and determine their location within eggs. We will then study how these enzymes change when eggs are activated by sperm and determine what impact blocking these enzymes has on how the embryo develops. This study should provide new information on these critically important proteins.
Technical Summary
ADP-ribosyl cyclases constitute a remarkable family of enzymes capable of catalyzing multiple reactions including the synthesis of the novel and potent Ca2+ mobilizing messengers, cADPR and NAADP. These enzymes are thus likely to play central roles in Ca2+ signaling - a process vitally important for a myriad of cellular events. Not all ADP-ribosyl cyclases however have been characterized at the molecular level. Furthermore, although both cADPR and NAADP have been implicated in fertilization their role during subsequent development is not known. The objective of this project is to provide a complete molecular description of ADP-ribosyl cyclases in a model organism (the sea urchin) and to define the functional role of these enzymes during early development. Building on substantial preliminary data, we will apply molecular and proteomic approaches to isolate sea urchin ADP-ribosyl cyclases and characterize their catalytic properties. Using molecular and immunological assays, we will define the complement of sea urchin ADP-ribosyl cyclases in eggs and the developing embryo and perform parallel biochemical measurements of activity and messenger levels. By inhibiting ADP-ribosyl cyclase expression, we will establish their role in i) the generation in vivo of cADPR and importantly, NAADP (of which little is known) and ii) the course of development. This study will provide novel insight in to the action of these highly versatile multifunctional enzymes.
Organisations
People |
ORCID iD |
Sandip Patel (Principal Investigator) |
Publications
Churamani D
(2008)
Molecular characterization of a novel cell surface ADP-ribosyl cyclase from the sea urchin.
in Cellular signalling
Ramakrishnan L
(2016)
ADP-ribosyl cyclases regulate early development of the sea urchin.
in Messenger (Los Angeles, Calif. : Print)
Schrlau MG
(2008)
Carbon nanopipettes characterize calcium release pathways in breast cancer cells.
in Nanotechnology
Patel S
(2018)
Two-pore channels open up.
in Nature
Churamani D
(2007)
Molecular characterization of a novel intracellular ADP-ribosyl cyclase.
in PloS one
Love NR
(2015)
NAD kinase controls animal NADP biosynthesis and is modulated via evolutionarily divergent calmodulin-dependent mechanisms.
in Proceedings of the National Academy of Sciences of the United States of America
Pandey V
(2009)
Recruitment of NAADP-sensitive acidic Ca2+ stores by glutamate.
in The Biochemical journal
Brailoiu GC
(2009)
NAADP-mediated channel 'chatter' in neurons of the rat medulla oblongata.
in The Biochemical journal
Ramakrishnan L
(2010)
A single residue in a novel ADP-ribosyl cyclase controls production of the calcium-mobilizing messengers cyclic ADP-ribose and nicotinic acid adenine dinucleotide phosphate.
in The Journal of biological chemistry
Brailoiu E
(2010)
An ancestral deuterostome family of two-pore channels mediates nicotinic acid adenine dinucleotide phosphate-dependent calcium release from acidic organelles.
in The Journal of biological chemistry