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Neurotransmitters, Second Messengers and Drug Action in the Nervous System

      Our lab studies diverse signaling systems including those of neurotransmitters and second messengers as well as the actions of drugs upon these processes.  We have been interested in atypical neurotransmitters such as nitric oxide (NO), carbon monoxide (CO), and the D-isomers of certain amino acids, specifically D-serine and D-aspartate.  A particular focus has been upon downstream targets of NO.  Besides its stimulation of cGMP formation, NO acts by nitrosylating a wide range of target proteins including prominent intracellular proteins such as the sodium pump and tubulin.  Recently we showed that nitrosylation mediates a novel interaction between the two major small molecule inflammatory systems in cells, NO formed by inducible NO synthase (iNOS) and the prostaglandin-forming COX2 enzyme.  iNOS and COX2 bind physiologically with NO formed by iNOS nitrosylating and activating COX2.  Inhibitors of iNOS-COX2 binding block prostaglandin formation which may afford a novel means of develoing anti-inflammatory drugs.

        We have discovered and characterized a novel cell death signaling cascade whereby cell stressors activate iNOS or neuronal NOS with the generated NO nitrosylating the glycolytic enzyme glyceraldehyde-3-phosphate-dehydrogenase (GAPDH).  Nitrosylation terminates the catalytic activity of the enzyme and confers upon it the ability to bind to Siah, a ubiquitin-3-ligase, which mediates nuclear translocation of GAPDH.  In the nucleus GAPDH initiates cell death apparently by binding to the protein acetylating enzyme p300 and activating it to stimulate p53, a tumor suppressor that kills cells.  This signaling cascade extends from the outside of the cell to the nucleus in a few simple steps, participates in diverse modes of cell death and provides multiple sites for intervention by drugs to prevent cell death and be potentially therapeutic in conditions such as stroke and neurodegenerative disease.  We showed that Deprenyl, a drug used to treat Parkinson’s disease and which protects neurons from cell death, acts by blocking the nitrosylation of GAPDH and its binding to Siah.  Moreover, the ability of huntingtin, the protein mutated in Huntington’s Disease, to enter the nucleus and kill neurons stems from its binding to GAPDH and Siah.

        We have established that D-serine is a novel neurotransmitter which acts by stimulating the “glycine” site of glutamate-NMDA receptors.  D-serine is formed primarily in glia which are activated by glutamate leading to the stimulation of serine racemase, the enzyme which converts L- to D-serine with release of D-serine. 

        In past years we identified, isolated and elucidated receptors for the second messenger IP3, which releases intracellular calcium.  We have been studying higher inositol phosphates, especially IP7 which contains an energetic pyrophosphate bond.  We discovered that IP7 phosphorylates proteins in a fashion rather different than does ATP.  This phosphorylation is a non-enzymatic auto-phosphorylation.  Most remarkably, the phosphate from IP7 is added only to sites on proteins that are already phosphorylated, hence IP7 pyrophosphorylates protein targets, a novel form of protein phosphorylation and a unique means of signal transduction.  Studies involving deletion of IP6 kinase, the enzyme that forms IP7 by phosphorylating IP6, reveal physiologic roles for IP7 in the trafficking of synaptic and other types of vesicles, apoptosis, chemotaxis, and telomere elongation.  One of the three IP6 kinase enzymes, IP6 kinase 2, appears to be physiologically associated with cell death induction.  It is normally bound in the cytoplasm to the heat shock protein HSP90 which maintains it in an inactive form.  Anti-cancer drugs that block this binding lead to increased IP7 formation in cell death.