Symposium on "Phosphatases in the Nervous System"
as part of the FASEB Summer Research Conference on "Protein Phosphatases"
July 17-22, 2004, Snowmass, Aspen, CO.

Overall Objectives of Meeting: The phosphorylation of proteins on serine, threonine and tyrosine residues is an essential element in the control of such fundamental cellular processes as growth and proliferation, differentiation, cytoskeletal function, the cell cycle, cellular metabolism, and in a variety of aspects of regulation of neuronal function. Although work on protein kinases has dominated the field of protein phosphorylation since its inception, recently, there has been an explosion of interest in protein phosphatase structure and function in many areas of biology. The objectives of the eighth FASEB Conference on Protein Phosphatases, held at Snowmass Village, Colorado, July 17-24, 2004, were to present the latest and most exciting topics in the Protein Phosphatase field, to discuss emerging technologies that will impact future research, to recognize outstanding achievement by specific investigators in the field in the form of Plenary and Keynote Lectures, and to encourage new participation in the field, particularly by junior investigators, women and under-represented minorities.

Participants: There were 130 participants, ~35% were representative of 19 countries other than the US: Austria, Australia, Belgium, Canada, China, Denmark, France, Germany, Hungary, India, Israel, Italy, Japan, Portugal, Spain, Switzerland, The Netherlands, Taiwan, and UK. The majority of the participants were from academia, including many graduate students (30) and postdoctoral fellows (35). Thirteen participants were from the pharmaceutical industry. Approximately 40% of the participants were women.

Program for meeting: The program was organized into eight scientific sessions, a Plenary lecture on the opening evening, and a Keynote lecture on Wednesday evening. On the opening evening a plenary lecture was given by Michael G. Rosenfeld, Howard Hughes Medical Institute, University California, San Diego. The Keynote Address presented on Wednesday evening was given by Eric Olson (University of Texas, Southwestern Medical Center). Beginning on Sunday morning, there were 8 regular scientific sessions that typically featured four or five 30 minute talks (long) and one to two 15 minute talks (short). The 8 sessions were, 1 - Structure and regulation of protein phosphatases; 2 - Protein phosphatases and targeting mechanisms; 3 - Phosphatases and the nervous system; 4 - Phosphatases and the cell cycle; 5 - Phosphatases in growth factor action and development; 6 - Protein tyrosine phosphatases in the immune system; 7 - Protein phosphatases and cell signaling; 8 - Phosphatases and disease. To emphasize the integrated nature of the protein phosphatase field, talks on protein tyrosine and protein serine/threonine phosphatases were interspersed. The quality of the presentations ranged from very good to outstanding. In addition to these scientific sessions, there were four poster sessions.

More than 60 posters were presented with 2 groups of 30 being presented for 2 days each. The poster sessions were very well attended and provided a background for a lot of discussion and interactions between participants.

Participation by new speakers, younger investigators and women: We made a strong effort to infuse "new blood" into the meeting, particularly in the form of young/new investigators to the field and women and under-represented minority participants. Of the 51 total speakers, 18 were women (35%). Two of the speakers were of African-American ethnicity. Almost half (25) of 2 the speakers had not spoken at any previous FASEB Summer Conference on Protein Phosphatases, and many of these individuals had not attended a previous meeting. While the majority of the speakers were from USA and Canada, there were a number of speakers (11) who work in other countries - including Australia, Belgium, Canada, Denmark, France, Israel, Japan, The Netherlands, UK. Fifteen of the speakers were either PhD/MD students, postdoctoral fellows or individuals who were recently appointed to junior faculty positions. As in past meetings, the inclusion of younger scientists (postdocs and graduate students) and late-breaking developments in the main program of the meeting was implemented largely from selection from the submitted posters, and the selected individuals gave short talks. There was general agreement that the "short talk" presentations were of very high quality. We also provided a number of "Poster Awards". A committee formed of the past and future chairs of the conference evaluated and selecting posters for awards, based on scientific merit. The "Poster awards" proved to be popular and encouraged active interactions of younger and senior investigators. Based on a grant of $5000 from the March of Dimes, we presented Poster Awards of $500 to 10 young investigators.

Fund raising: Fund raising for the conference was quite strong. We received a total of 11 contributions from pharmaceutical and biotechnology companies, including two contributions of over $5,000 and 3 donations of $2,000-4,999 (Pfizer, Novo Nordisk, Ceptyr, Proctor & Gamble Pharmaceuticals, Hoffman-La Roche Inc., Berlex, Intra-Cellular Therapies, Inc., Merck & Co., Inc, Amgen, Cell Signaling Technology, Inc., Bristol-Myers Squibb). We also obtained grant support from the NIH (NCI) in the amount of $8,000. Finally we obtained two awards of $5000 each from the March of Dimes and the International Society for Neurochemistry. The award from the March of Dimes was used to support the poster awards. The grant from the International Society for Neurochemistry was used to support exclusively the speakers in the session on "Protein Phosphatases in the Nervous System". From these resources, plus the amount provided by FASEB (total $54,000), we were able to reimburse the full registration for the speakers who gave long talks, and the Plenary and Keynote lecturers. We provided partial travel support for all these speakers, which represented 50-100% of their airfares.

Review of symposium session on "Phosphatases and the nervous system" supported by the International Society for Neurochemistry: This session, organized and chaired by Angus Nairn, Yale Univ., was held on Monday morning and was one of the highlights of the meeting. There were 6 speakers in the session who discussed various aspects of serine/threonine and tyrosine protein phosphatases in neuronal development, synaptic plasticity, and neuronal disease. These were: Isabelle Mansuy (Inst Cell Biol. Zurich) "Phosphatases in brain ischemia and recovery"; David Van Vactor (Harvard Medical School) "LAR signaling and synapse morphogenesis"; Mark DellAcqua (Univ. Colorado) "PP2B targeting at synapses"; Shirish Shenolikar (Duke Univ.) "Neurabin-I/PP1 and maturation of dendritic spines"; Paul Lombroso (Yale Univ.) "Taking STEPs to limit MAP kinase signaling"; Estelle Sontag (UT Southwestern) "PP2A and Alzheimers disease" (Mark DellAcqua and Estelle Sontag presented short talks). Details of the presentations are included in the summaries appended below. One notable feature of the presentation of given by Paul Lombroso that is not included in the summary, is that he gave a translational presentation that bridged his clinical work in child psychiatric illness (autism and Tourette's) and basic research into the protein tyrosine phosphatase, STEP, in learning processes related to these psychiatric illnesses.

Summaries of presentations in ISN sponsored symposium on Protein Phosphatases and the nervous system.

Isabelle M. Mansuy. (Swiss Federal Inst. Tech. Zürich, Switzerland) discussed the role of Protein Phosphatase 1 in brain ischemia and recovery. The hippocampus is a brain area essential for learning and memory that exhibits a high vulnerability to energy deprivation such as experienced during ischemic episodes. The mechanisms of such vulnerability are not well understood but it is recognized that brain ischemia can lead to the induction of pathological forms of synaptic plasticity causing delayed neuronal death, and also to changes in plasticity during recovery. The role of the Ser/Thr protein phosphatase PP1 in these processes was investigated using an in vitro model of hippocampal ischemia and a combination of genetic, pharmacological and physiological techniques. The results show that manipulations resulting in the inhibition of PP1 worsen the effects of ischemia on synaptic transmission. They diminish the recovery of field excitatory postsynaptic potentials (fEPSPs) following ischemia in area CA1 of the hippocampus. By contrast, manipulations that increase PP1 activity improve recovery. These results suggest that PP1 may be a natural neuroprotectant that plays an important role in the control of brain hemostasis and plasticity.

David Van Vactor (Dept. Cell Biology, Harvard Medical School, Boston) discussed the role of the LAR signaling pathway in synapse morphogenesis. The LAR receptor protein tyrosine phosphatase and its intracellular binding partner Liprin act to promote synapse growth and to control morphogenesis at the active zones that orchestrate neurotransmitter release. Parallel studies in mammalian synapses suggest this role has been well conserved, and raise questions as to what effectors collaborate with Liprin to control synapse growth and synapse maturation. Using a combination of genetics, cell biology and biochemistry, LAR has been found to interact with the tyrosine kinase, Abl, and to act antagonistically to regulate synapse growth. Analysis of Liprin also has revealed a failure in the assembly of T-bar structures at synapses suggesting a role not only in active zone assembly, but also for the correct delivery of synaptic components to sites of release.

Mark L. Dell'Acqua (Dept. Pharm. Univ. Colorado Hlth. Sci. Ctr., Denver) discussed the NMDA receptor-PP2B pathways implicated in regulation of AKAP79/150 and PKA at synapses. At the postsynaptic membrane of glutamatergic synapses, protein kinase A (PKA) and protein phosphatase 2B/calcineurin (PP2B/CaN) in association with the anchoring protein AKAP79/150 are recruited to NMDA and AMPA receptors by PSD-95 family MAGUK scaffold proteins. These complexes may function in NMDA receptor regulation of AMPA receptor phosphorylation and trafficking in hippocampal long-term potentiation (LTP) and epression (LTD) synaptic plasticity. Thus, it is important to understand mechanisms of AKAP79/150 targeting to synapses and recruitment to PSD-MAGUKs. Using fluorescence resonance energy transfer (FRET) imaging living neurons, PP2B and PKA-RII localization was studied. These studies indicated that NMDA-LTD treatment of hippocampal slices (that induces AMPA receptor GluR1-Ser845 dephosphorylation and internalization) also causes a shift in the distribution of AKAP79/150 and PKA-RII, but not PSD-95 or PP2B/CaNA, away from synaptic membrane fractions into cytosolic fractions all in a PP2B regulated manner. Based on these studies, NMDA-regulated removal of the AKAP79/150-PKA complex from dendritic spines 4 likely functions to shift the local balance of kinase and phosphatase activity thus favoring dephosphorylation of synaptic substrates such as GluR1that are important for LTD induction.

S. Shenolikar (Dept. Pharm., Duke Univ. Med. Ctr., Durham) discussed the role of Neurabin-I (neuronal actin-binding protein) which is highly enriched in growth cones and dendritic spines of mammalian neurons. Within spines, neurabin acts as a protein scaffold that targets protein phosphatase-1 (PP1) to the actin cytoskeleton, specifically the structure known as the postsynaptic density. Biochemical and cell-biological analyses have established that neurabin-I reorganizes F-actin fibers and elicits morphological changes in both neurons and non-neuronal cells. Neurabins also homo- and heterodimerize via their C-terminal coiled-coil domains and their dimerization functions to negatively regulate the generation of filopodia. Mutagenesis studies have highlighted the key role of the N-terminal domain (1-286) of neurabin in actin bundling. Other cellular studies have suggested that the neurabin-I-associated PP1 also played a critical role in modulating neuronal morphology. Together these studies suggest that neurabin-1 targets PP1 and other signaling proteins to the actin cytoskeleton during the development and regulation of neuronal circuitry.

Paul Lombroso (Child Study Ctr., Yale Univ. Sch. Med., New Haven) discussed the role of the protein tyrosine phosphatase STEP, a brain specific PTP that is only expressed in neurons. STEP is highly enriched in the basal ganglia and related structures, brain regions involved in learning and memory. Substrates for STEP include the tyrosine kinase, fyn, NMDA receptors, and the MAP kinase family of signaling proteins. STEP binds to the ERKs and dephosphorylates and inactivates their regulatory tyrosine residue. In this way, STEP is able to limit the duration that the ERKs are active. Recent results indicate that STEP is involved in learning and memory through its ability to regulate the ERK pathway. STEP is itself regulated by phosphorylation and two important neurotransmitters are involved in this process. Dopamine stimulation leads to a DA/cAMP/PKA-mediated phosphorylation of STEP at specific serine residues. STEP is now unable to interact with downstream effectors due to steric interference at its ERK binding site. Glutamate stimulation leads to an NMDAR/calcineurin-mediated dephosphorylation of STEP at the same regulatory serine residues. STEP thus acts as a switch within neurons to modulate ERK signaling, and has long-term effects on learning and behavior.

Estelle Sontag (Dept. Pathol., UT Southwestern Medical Center, Dallas) discussed the role of alterations in protein phosphatase 2A (PP2A) expression and post-translational modification in Alzheimer's disease pathology. The formation of ß-amyloid-containing senile plaques and taurich neurofibrillary tangles are central events in Alzheimer's disease (AD) pathogenesis. The major brain protein phosphatase 2A (PP2A), specifically binds to and dephosphorylates tau. Notably, the methylation of PP2A catalytic subunit is critically required for ABaC holoenzyme assembly. Immunoblotting analyses revealed that there was a significant and selective reduction in the total amounts of ABaC in AD-affected brain regions, which matched the decrease in PP2A activity and methylation levels measured in the same brain homogenates. Immunohistochemical studies showed that neuronal ABaC expression levels were significantly reduced in AD-affected regions, but not in normal controls and non-AD dementias. The regional loss of neuronal ABaC immunoreactivity closely reflected the severity of tau pathology, but not amyloid plaque burden. Thus deregulation of PP2A methylation/demethylation cycles and downregulation of ABaC holoenzymes may contribute to AD pathogenesis.