European Journal zyxwvutsrqp of Neuroscience, zyxwvutsrqpo Vol. zyxwvutsrq 7, pp. 1810-1821, I995 0 European Neuroscience Association zy Focal Adhesion Kinase in Rat Central Nervous System Ferran Burgaya*, Andrea Menegonl*, Mathias Menegoz, Flavia Valtorta' and Jean-Antoine Girault Chaire de Neuropharmacologie, INSERM U114, College de France, 11, place Marcelin Berthelot, 75005 Paris, France 'Department of Medical Pharmacology, CNR Center of Cytopharmacology and DIBIT, San Raffaele Scientific Institute, University of Milan, Milan, Italy Keywords: tyrosine kinase, development, actin, neuronal cultures, hippocampus Abstract Focal adhesion kinase ( p~125~~~, FAK) is a 125 kDa non-receptor tyrosine kinase enriched in focal adhesions of various cell types, where it is thought to transduce signals triggered by contact with the extracellular matrix. We have studied the expression and localization of FAK in rat CNS. Immunoblotting, immunohistochemistry and in situ hybridization revealed the presence of FAK in all regions of the adult brain and demonstrated its enrichment in specific neuronal populations of the cerebral and cerebellar cortex, as well as in the hippocampus. During development, FAK protein levels were highest around birth in cerebral cortex and caudate putamen and decreased in the adult. In situ hybridization revealed enrichment of FAK mRNA in the ventricular germinative and external layers during the last period of embryonic growth. In primary cultures FAK immunoreactivity was localized in focal adhesions in astrocytes, whereas in developing neurons the highest levels were found in growth cones and perikarya. In the growth cone, FAK immunoreactivitycolocalized with actin filaments. In mature neurons FAK appeared to be distributed in the whole cytoplasm, with no enrichment in any cellular compartment. Our results demonstrate the presence of high levels of FAK in rat CNS, maximal during development but persistent in the adult. Its enrichment in growth cones suggests that it may play a role in neurite outgrowth, as well as in plasticity in the adult. introduction zyxwvutsrqp Phosphorylation of proteins is the most common reversible post- translational modification regulating their properties, and numerous extracellular signals result in its activation or inhibition. Most of the transmembrane receptors for growth factors are tyrosine kinases, or are associated with tyrosine kinases. Tyrosine phosphorylation appears to trigger enzymatic cascades that transduce external signals involved in regulation of growth and differentiation in all cell types. In addition, the fact that many tyrosine kinases and phosphatases are highly expressed in neurons suggests an important regulatory role for protein tyrosine phosphorylation in the context of information processing by the nervous tissues (reviewed in Wagner and Huganir, 1991; Girault, 1993). In neurons, the tyrosine phosphorylation machinery is enriched at the synaptic level, both on the pre- and postsynaptic sides (for refs see Wagner and Huganir, 1991), and is particularly active at the time of synaptogenesis during development (Girault zyxwvut er a/., 1992a). Neuromodulation and the regulation zyxwvutsrq of synaptic efficiency involve phosphorylation of receptors and ion channels. Tyrosine phosphorylation regulates nicotinic acetylcholine receptor desensitiza- tion (Hopfield et al., 1988), NMDA receptors (Wang and Salter, 1994) and cation channels permeability (Huang et a/., 1993; Wilson and Kaczmarek, 1993). Tyrosine phosphorylation of receptors has also been suggested to play a role in receptor aggregation, and hence to be involved in the regulation of synapse formation and stability (Swope et al., 1992). Several synaptic vesicle-specific proteins are also phosphorylated on tyrosine (Pang et al., 1988). The probable role of tyrosine phosphorylation in synaptic plasticity is suggested by the impairment of long-term potentiation zyx (LTP) in the hippocampus, which is observed following application of tyrosine kinase inhibitors (O'Dell et al., 1991) or in mutant mice lacking a functional gene for the tyrosine kinase pp59'Y" (Grant et al., 1992). Focal adhesion kinase (FAK), a 125 kDa tyrosine kinase also known as ppl 25FAK, has been initially characterized as a non-receptor- protein tyrosine kinase associated with focal adhesions in chicken embryo cells (Schaller et al., 1992) and mouse fibroblasts (Hanks et al., 1992). Subsequent studies have shown FAK to be present in most tissues, either by Northern blot hybridization (Hanks et al., 1992; Andre and Becker-Andre, 1993) or by immunoblotting (Turner et al., 1993). Comparison of the sequences of FAK cDNA in chicken (Schaller et al., 1992). mouse (Hanks et al., 1992) and human (Whitney et al., 1993) reveals that it is a highly conserved protein. FAK is thought to be involved in cell adhesion, at the level of the linkage between extracellular matrix (ECM) receptors (the integrins) and actin microfilaments, in association with proteins such as talin, paxillin and alpha-actinin (Schaller and Parsons, 1994). In addition, recent reports demonstrate that FAK activates enzymatic cascades Correspondence to: Ferran Burgaya *The first two authors contributed equally to this work Received 29 March 1995, accepted 8 May I995