Survival in a dynamic external environment demands the ability to monitor and respond to a wide range of internal and external variables, or signals. The binding of a signal to its receptor initiates a sequence or "cascade" of molecular events inside the cell that modulate relevant metabolic, nuclear, motile, or other processes. The transmission of an often extracellular receptor binding event into the interior of the cell and its translation into a catalytic or other response is called signal transduction. To be consistently successful in nature's continual competition for scarce resources, such responses must be rapid and efficient. Efficiency demands that the response be comprehensive in scope to insure against wastage of either materials or energy. This task is rendered challengingly complex by the cacophony of competing and contradictory signals that may bombard the cell at any single instant. A microbe floating in a pond is continually subjected to variations in temperature, sunlight, external pH, oxygen tension, and the availability of the compounds that supply the carbon, nitrogen, sulfur, phosphorous, etc. needed to sustain life. Mammalian cells are bathed in a churning humoral sea containing scores of hormones whose shifting levels are punctuated by flashes of neuronal excitation. Cells need mechanisms for integrating information and making decisions, for selecting the response(s) most appropriate for a particular combination of signal inputs. In other words, signal transduction demands much more than the construction of molecular switches for turning particular cellular processes "on" and "off". Simple linear connections between a receptor and its primary site of action are not enough. Successful signal transduction demands the creation of highly branched, tightly interwoven, integrated command and control networks - organic microprocessors - in which the signaling pathways triggered by particular effectors actively interact with and influence one another.

Nature creates sophisticated microprocessors from the simple organic components that reside within cells in essentially the same way in which man builds computers with inorganic materials. In man-made computers bits are electromagnetic in nature, as are their functional outputs. In the biological world, bits are chemical in nature. One of the most frequently employed organic bits found in nature is the addition and removal of a phosphate group from a protein, a process referred to as protein phosphorylation-dephosphorylation. Its high charge density renders the phosphate group a potent perturber of protein structure and function. Thus, the phosphorylation of a protein through the action of a protein kinase can have a dramatic effect on its catalytic capabilities, location within the cell, affinity for a ligand, etc. The phosphoprotein can be restored to its unmodified state on demand via the hydrolytic removal of the phosphoryl group through the action of a protein phosphatase.

The long-range objective of my research group is to understand how cells integrate protein phosphorylation-dephosphorylation processes to create information processing networks. To accomplish this, we are mapping the physical and functional architecture of the protein phosphorylation networks from two evolutionarily distinct prokaryotes, the cyanobacterium Synechocystis sp. PCC 6803 and the thermophilic archaeon Sulfolobus solfataricus. Work on the cyanobacterium is being carried out in collaboration with Malcolm Potts' laboratory. Prokaryotes offer several advantages for achieving this goal. These include the availability of complete genome sequences for numerous prokaryotes, their limited and non-redundant gene population, their susceptibility to genetic manipulation, and the ability to trace the development of regulatory networks over evolutionary time. Our approach to this problem is a highly multidisciplinary one. It involves:
a) Identifying prokaryotic protein kinases, protein phosphatases, and phosphoproteins by a combination of sequence analysis techniques (genomics) and protein chemistry.

b) Cloning and expression of the genes encoding these proteins and evaluation of their functional properties (substrate specificity, functional consequences of phosphorylation, etc.) in vitro.

c) Evaluating the physiological role of individual phosphorylation events by analyzing the phenotypic consequences of genetically knocking out individual protein kinases or protein phosphatases.

d) The construction and evaluation of computer models that replicate the architecture and function of this information processing network.

The successful dissection and modeling of protein phosphorylation-dephosphorylation networks as complete, integrated wholes will open a door to many new technologies. These include more efficient and versatile strategies for the genetic engineering of microorganisms, the fuller exploitation of microorganisms as environmental monitors, and the creation of man-made microprocessors based on organic technology. This also represents an important step in unravelling the much more quantitatively complex organic computers found in human cells, the understanding of which will lead to new breakthroughs in human health and well-being.

Wurgler-Murphy, S. M., King, D. M., and Kennelly, P. J. (2004) The phosphorylation site database: A guide to the serine-, threonine-, and/or tyrosine-phosphorylated proteins in prokaryotic organisms. Proteomics 4:1562-1570.   [Abstract]

Lower, B. H., Potters, M. B., and Kennelly, P. J. (2004) A Phosphoprotein from the Archaeon Sulfolobus solfataricus with Protein-Serine/Threonine Kinase Activity. J. Bacteriol. 186:463-472.   [Abstract]

Lower, B.H. and Kennelly, P.J. (2003) Open Reading Frame sso2387 from the Archaeon Sulfolobus solfataricus Encodes a Polypeptide with Protein-Serine Kinase Activity. J. Bacteriol. 185(11):3436-3445.   [Abstract]

Kennelly, P.J. (2003) Archaeal Protein Kinases and Protein Phosphatases: Insights from Genomics and Biochemistry. Biochem. J. 370:373-389.   [Abstract]

Potters, M.B., Solow, B.T., Bischoff, K.M., Graham, D.E., Lower, B.H., Helm, R., and Kennelly, P. J. (2003) Phosphoprotein with Phosphoglycerate Mutase Activity from the Archaeon Sulfolobus solftaricus. J. Bacteriol. 185(7):2112-2121.   [Abstract]

Li, R., Haile, J. D., and Kennelly, P. J. (2003) An Arsenate Reductase from Synechocystis sp. Strain PCC 6803 Exhibits a Novel Combination of Catalytic Characteristics. J. Bacteriol. 185:6780-6789.   [Abstract]

Lower, B. H., and Kennelly, P. J. (2002) The Membrane-Associated Protein-Serine/Threonine Kinase from Sulfolobus solfataricus Is a Glycoprotein. J. Bacteriol. 184:2614-2619.   [Abstract]

Kennelly, P. J. (2001) Protein Kinases and Protein Phosphatases in Prokaryotes: A Genomic Perspective. FEMS Microbiol. Lett. 206:1-8.   [Abstract]

Kennelly, P. J. (2001) Protein Phosphatases: A Phylogenetic Perspective. Chemical Reviews 101:2291-2312.   [Abstract]

Lower, B. H., Bischoff, K. M., and Kennelly, P. J. (2000) The Archaeon Sulfolobus solfataricus Contains a Membrane-Associated Protein Kinase Activity that Preferentially Phosphorylates Threonine Residues In Vitro. J. Bacteriol. 182:3452-3459.   [Abstract]