BackgroundComplex intracellular signaling networks monitor diverse environmental inputs to evoke appropriate and coordinated effector responses. Defective signal transduction underlies many pathologies, including cancer, diabetes, autoimmunity and about 400 other human diseases. Therefore, there is high impetus to define the composition and architecture of cellular communications networks in humans. The major components of intracellular signaling networks are protein kinases and protein phosphatases, which catalyze the reversible phosphorylation of proteins. Here, we have focused on identification of kinase-substrate interactions through prediction of the phosphorylation site specificity from knowledge of the primary amino acid sequence of the catalytic domain of each kinase.ResultsThe presented method predicts 488 different kinase catalytic domain substrate specificity matrices in 478 typical and 4 atypical human kinases that rely on both positive and negative determinants for scoring individual phosphosites for their suitability as kinase substrates. This represents a marked advancement over existing methods such as those used in NetPhorest (179 kinases in 76 groups) and NetworKIN (123 kinases), which consider only positive determinants for kinase substrate prediction. Comparison of our predicted matrices with experimentally-derived matrices from about 9,000 known kinase-phosphosite substrate pairs revealed a high degree of concordance with the established preferences of about 150 well studied protein kinases. Furthermore for many of the better known kinases, the predicted optimal phosphosite sequences were more accurate than the consensus phosphosite sequences inferred by simple alignment of the phosphosites of known kinase substrates.ConclusionsApplication of this improved kinase substrate prediction algorithm to the primary structures of over 23, 000 proteins encoded by the human genome has permitted the identification of about 650, 000 putative phosphosites, which are posted on the open source PhosphoNET website (http://www.phosphonet.ca).

Habituation is a ubiquitous form of non-associative learning observed as a decrement in responding to repeated stimulation that cannot be explained by sensory adaptation or motor fatigue. One of the defining characteristics of habituation is its sensitivity to the rate at which training stimuli are presented—animals habituate faster in response to more rapid stimulation. The molecular mechanisms underlying this interstimulus interval (ISI)-dependent characteristic of habituation remain unknown. In this article, we use behavioural neurogenetic and bioinformatic analyses in the nematode Caenorhabiditis elegans to identify the first molecules that modulate habituation in an ISI-dependent manner. We show that the Caenorhabditis elegans orthologues of Ca 2+ /calmodulin-dependent kinases CaMK1/4, CMK-1 and O-linked N-acetylglucosamine (O-GlcNAc) transferase, OGT-1, both function in primary sensory neurons to inhibit habituation at short ISIs and promote it at long ISIs. In addition, both cmk-1 and ogt-1 mutants display a rare mechanosensory hyper-responsive phenotype (i.e. larger mechanosensory responses than wild-type). Overall, our work identifies two conserved genes that function in sensory neurons to modulate habituation in an ISI-dependent manner, providing the first insights into the molecular mechanisms underlying the universally observed phenomenon that habituation has different properties when stimuli are delivered at different rates.

The reversible phosphorylation of proteins catalyzed by protein kinases in eukaryotes supports an important role for eukaryotic protein kinases (ePKs) in the emergence of nucleated cells in the third superkingdom of life. Choline kinases (ChKs) could also be critical in the early evolution of eukaryotes, because of their function in the biosynthesis of phosphatidylcholine, which is unique to eukaryotic membranes. However, the genomic origins of ePKs and ChKs are unclear. The high degeneracy of protein sequences and broad expansion of ePK families have made this fundamental question difficult to answer. In this study, we identified two class-I aminoacyl-tRNA synthetases with high similarities to consensus amino acid sequences of human protein-serine/threonine kinases. Comparisons of primary and tertiary structures supported that ePKs and ChKs evolved from a common ancestor related to glutaminyl aminoacyl-tRNA synthetases, which may have been one of the key factors in the successful of emergence of ancient eukaryotic cells from bacterial colonies.Protein kinases play a pivotal role in communicating intracellular signals in eukaryotes. The family of eukaryotic protein kinases (ePKs) 3 comprises at least 568 human members, which accounts for more than 2% of the protein coding genes of the entire human genome (1). These kinases are highly conserved both in their primary amino acid sequences (2) and in the threedimensional structures (3) of their catalytic domains. Because of the central regulatory roles and the high conservation of the ePKs, the ancestry of these enzymes has become an important question in the study of the evolution of eukaryotic organisms.The majority of the kinases among the ePKs are responsible for the phosphorylation of proteins on serine or threonine residues, whereas a smaller group of protein kinases catalyzes their tyrosine phosphorylation. This branch of protein-tyrosine kinases (PTKs) arose from protein-serine/threonine kinases (STKs), which is believed to be an important event in early metazoan evolution (4, 5). Of all the STKs, there is another lumped group of diverse kinases that are described as atypical protein kinases. With little sequence identity and structural similarity to typical protein kinases, these atypical protein kinases are suggested to have diverged early in evolution and have distinct evolutionary histories (6, 7). Despite the atypical protein kinases and recently derived PTKs, the rest of the typical protein kinases constitutes a major lineage in protein kinase evolution.Eukaryotic life is believed to have evolved between 1.7 and 2.7 billion years ago, and no living representatives of the earliest eukaryotes survive today. Consequently, the actual origin of protein kinases is difficult to establish with a high degree of confidence. Firstly, protein sequences are highly degenerate, which makes the detection of sequence similarities difficult even at the superfamily level (8). Secondly, the ePKs comprise a group of very broadly expanded proteins. Loss and expansion of kin...

(250 words)Many aspects of neural physiology, including processes that underlie learning (e. OGT-1 function in parallel pathways that may converge on a common substrate to modulate the tap processes present in all animals. peer-reviewed)

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