In humans, the functional loss of the fragile X mental retardation protein (FMRP) 2 causes the fragile X syndrome (FXS), a severe form of inherited mental retardation (1-4). In the brain of both humans and mice, FMRP deficiency results in a significant change in both dendritic spine morphology and synaptic function (5-9). FMRP is an RNA-binding protein that is thought to act primarily as a repressor of mRNA translation. Among other subcellular regions in neurons, FMRP appears to exercise this control function at postsynaptic sites. It has been hypothesized that in dendrites FMRP locally controls the synthesis of proteins, such as components of the postsynaptic density (PSD), which regulate both dendritic spine morphology and synaptic function (2, 9, 10). The PSD is a complex protein network lying underneath the postsynaptic membrane of excitatory synapses (11-13). It serves to cluster glutamate receptors and cell adhesion molecules, recruit signaling proteins, and anchor these components to the microfilament-based cytoskeleton in dendritic spines. To combine these functions, the central layers of the PSD consist of several scaffold proteins, such as members of the PSD-95, SAPAP/GKAP, and Shank/ProSAP families. Because of their capacity to directly interact with many different PSD components and to regulate the size and shape of dendritic spines, Shanks in particular are assumed to represent master scaffold proteins of the PSD (11). Activity-dependent changes in the PSD composition are thought to represent a molecular basis for most principal brain functions, including learning and memory. Several of these long term synaptic changes and learning paradigms critically depend on dendritic protein synthesis (14 -17). Interestingly, mRNAs encoding some of the central components of the PSD, such as Shank1-3, SAPAP3, PSD-95, and ␣-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid-type glutamate receptor subunits (GluR), are present in dendrites (18 -23).As FMRP has been implicated in the local regulation of mRNA translation at synapses, one crucial question is as follows: which postsynaptic proteins are affected by the loss of FMRP in a quantitative manner and may thus contribute to abnormal dendritic spine morphology and impaired synaptic plasticity? To specifically address this question, we took advan-* This work was supported by the Deutsche Forschungsgemeinschaft Grants Ki488/2-6 (to S. K.) and KR 1321/4-1 (to H.-J. K. and S. K.) and Thyssen-Stiftung Az. 10.05.2.185 (to D. R. and S. K.

Mutations in the polytopic lysosomal membrane glycoprotein CLN3 result in a severe neurodegenerative disorder. Previous studies identified two cytosolic signal structures contributing to lysosomal targeting. We now examined the role of glycosylation and the C-terminal CAAX motif in lysosomal transport of CLN3 in nonneuronal and neuronal cells. Mutational analysis revealed that in COS7 cells, CLN3 is glycosylated at asparagine residues 71 and 85. Both partially and non-glycosylated CLN3 were transported correctly to lysosomes. Mevalonate incorporation and farnesyltransferase inhibitor studies indicate that CLN3 is prenylated most likely at cysteine 435. Substitution of cysteine 435 reduced the steady-state level of CLN3 in lysosomes most likely because of impaired sorting in early endosomal structures, particularly in neuronal cells. Additionally, the cell surface expression of CLN3 was increased in the presence of farnesyltransferase inhibitors. Alteration of the spacing between the transmembrane domain and the CAAX motif or the substitution of the entire C-terminal domain of CLN3 with cytoplasmic tails of mannose 6-phosphate receptors have demonstrated the importance of the C-terminal domain of proper length and composition for exit of the endoplasmic reticulum. The data suggest that co-operative signal structures in different cytoplasmic domains of CLN3 are required for efficient sorting and for transport to the lysosome.

Dendritic mRNA transport coupled with local regulation of translation enables neurons to selectively alter the protein composition of individual postsynaptic sites. We have analyzed dendritic localization of shank1 mRNAs; shank proteins (shank1-3) are scaffolding molecules of the postsynaptic density (PSD) of excitatory synapses, which are crucial for PSD assembly and the formation of dendritic spines. Live cell imaging demonstrates saltatory movements of shank1 mRNA containing granules along microtubules in both anterograde and retrograde directions. A population of brain messenger ribonucleoprotein particles (mRNPs) containing shank1 mRNAs associates with the cargo-binding domain of the motor protein KIF5C. Through expression of dominant negative proteins, we show that dendritic targeting of shank1 mRNA granules involves KIF5C and the KIF5-associated RNA-binding protein staufen1. While transport of shank1 mRNAs follows principles previously outlined for other dendritic transcripts, shank1 mRNAs are distinguished by their translational regulation. Translation is strongly inhibited by a GC-rich 5 untranslated region; in addition, internal ribosomal entry sites previously detected in other dendritic transcripts are absent in the shank1 mRNA. A concept emerges from our data in which dendritic transport of different mRNAs occurs collectively via a staufen1-and KIF5-dependent pathway, whereas their local translation is controlled individually by unique cis-acting elements.