Abstract
BackgroundIntracellular Ca2+ regulates many aspects of neuronal function through Ca2+ binding to EF hand-containing Ca2+ sensors that in turn bind target proteins to regulate their function. Amongst the sensors are the neuronal calcium sensor (NCS) family of proteins that are involved in multiple neuronal signalling pathways. Each NCS protein has specific and overlapping targets and physiological functions and specificity is likely to be determined by structural features within the proteins. Common to the NCS proteins is the exposure of a hydrophobic groove, allowing target binding in the Ca2+-loaded form. Structural analysis of NCS protein complexes with target peptides has indicated common and distinct aspects of target protein interaction. Two key differences between NCS proteins are the size of the hydrophobic groove that is exposed for interaction and the role of their non-conserved C-terminal tails.ResultsWe characterised the role of NCS-1 in a temperature-dependent locomotion assay in C. elegans and identified a distinct phenotype in the ncs-1 null in which the worms do not show reduced locomotion at actually elevated temperature. Using rescue of this phenotype we showed that NCS-1 functions in AIY neurons. Structure/function analysis introducing single or double mutations within the hydrophobic groove based on information from characterised target complexes established that both N- and C-terminal pockets of the groove are functionally important and that deletion of the C-terminal tail of NCS-1 did not impair its ability to rescue.ConclusionsThe current work has allowed physiological assessment of suggestions from structural studies on the key structural features that underlie the interaction of NCS-1 with its target proteins. The results are consistent with the notion that full length of the hydrophobic groove is required for the regulatory interactions underlying NCS-1 function whereas the C-terminal tail of NCS-1 is not essential. This has allowed discrimination between two potential modes of interaction of NCS-1 with its targets.
Highlights
Intracellular Ca2+ regulates many aspects of neuronal function through Ca2+ binding to EF hand-containing Ca2+ sensors that in turn bind target proteins to regulate their function
Role of neuronal calcium sensor (NCS)-1 in temperature-dependent locomotion in C. elegans C. elegans possesses a single orthologue of human NCS-1
We have recently established a simple assay for temperature-dependent locomotion (TDL) in C. elegans as a read out for thermosensory signalling and behaviour based on acute inhibitory effect of elevated temperature on locomotion
Summary
Intracellular Ca2+ regulates many aspects of neuronal function through Ca2+ binding to EF hand-containing Ca2+ sensors that in turn bind target proteins to regulate their function. Each NCS protein has specific and overlapping targets and physiological functions and specificity is likely to be determined by structural features within the proteins. Common to the NCS proteins is the exposure of a hydrophobic groove, allowing target binding in the Ca2+-loaded form. Structural analysis of NCS protein complexes with target peptides has indicated common and distinct aspects of target protein interaction. The neuronal calcium sensor (NCS) proteins form a family of Ca2+-binding proteins which have been implicated in the control of multiple aspects of neuronal function [1,2,3]. Whilst being similar to calmodulin they are distinguished by usually having a higher affinity for Ca2+ binding and certain family members are membrane targeted through N-terminal myristoylation (NCS-1, VILIPs, recoverin, and KChIP1) or palmitoylation (certain KChIPs). Despite having high levels of sequence and structural similarity the NCS proteins have a number of known non-redundant functions [3,10]
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