Actinfilin, a Brain-specific Actin-binding Protein in Postsynaptic Density

Abstract

The dynamic assembly and disassembly of actin-based cytoskeleton is closely linked to the changes in the postsynaptic density in both number and shape, which is thought to be important in forming long-term memory. Thus, regulation of actin filaments may play a critical role in contributing to the formation of long-term memory. Here, we report the cloning of actinfilin, a brain-specific Kelch protein, which interacts with F-actin. Actinfilin contains an amino-terminal POZ/BTB domain and carboxyl positioned six tandem Kelch repeats that presumably form six blades of ␤-propeller structure of the Kelch domain. Co-immunoprecipitation analyses showed that the amino-terminal POZ domain mediated actinfilin-actinfilin interaction. The recombinant Kelch domain alone was sufficient to mediate binding to Factin. Immunohistochemistry studies of rat brain sections suggested that actinfilin is broadly expressed in neurons of most regions of the brain. The subcellular localization of actinfilin was studied by biochemical fractionation and immunogold labeling. The results showed the postsynaptic density distribution of actinfilin. Together, these results indicate that actinfilin may be a key player in the actin-based neuronal function.Cytoskeleton structures play a role in determining the static and dynamic morphology of subcellular structures in neurons. Activity-induced actin assembly and disassembly is thought to play a critical role in shaping the dendritic spine, a subcellular protrusion that is found in more than 90% of glutamatergic synapses. The changes in number, size, and morphology of spines are linked to long-term memory (1-6). Increasing evidence suggests that the dynamic reorganization of actin filament is probably mediated by several classes of molecules including small second messages and regulatory protein factors that interact with G-or F-actin. Thus, molecular identification and functional understanding of regulatory components may provide important insights into both the dynamic morphology of synaptic structure and the activities that are coupled with polymerization, stability, and subcellular anchoring of actin filaments. CAP70 1 is a scaffolding protein that was purified from the kidney on the basis of its interaction with the carboxyl terminus of cystic fibrosis transmembrane conductance regulator (7). Like other PDZ domain-containing proteins, such as PSD95 or INAD (inactivation no afterpotential D), the CAP70 coding sequence is essentially composed of four tandemly positioned PDZ domains. In contrast to other known PDZ domain-containing proteins, CAP70 has the activity to dynamically alter the oligomerization state of the cystic fibrosis transmembrane conductance regulator channel, thereby inducing potentiation of cystic fibrosis transmembrane conductance regulator-induced chloride channel activity (7). The native CAP70 protein was found in actin-rich domains, such as the apical surfaces of epithelial cells in the small intestine and kidney as well as flopodia-like structures in cultured epithelial cells...

KRIP6: A novel BTB/kelch protein regulating function of kainate receptors

“…BTB/kelch proteins are highly conserved across species, yet their functional roles appear quite diverse, ranging from organization of cytoskeletal elements to control of protein trafficking/ ubiquitination and regulation of gene expression. Two brain-enriched BTB/kelch proteins closely related to KRIP6, Mayven and actinfilin, associate with the actin cytoskeleton (Chen et al, 2002;Soltysik-Espanola et al, 1999). However, unlike Mayven and actinfilin, KRIP6 does not colocalize with F-actin in COS-7 cells or in neurons, and thus probably does not bind actin.…”

Actinfilin Is a Cul3 Substrate Adaptor, Linking GluR6 Kainate Receptor Subunits to the Ubiquitin-Proteasome Pathway

“…The myc-tagged actinfilin and its two truncations are described in Ref. 26. nAchR␥-HA was received from Dr. Edward Hawrot (Brown University, Providence, RI).…”

“…The rabbit anti-actinfilin antibody (1:5000 for immunoblotting) is described in Ref. 26. The rabbit anti-Cul3 antibody was developed as previously described (40) and used at 1:500 for immunoblotting.…”

“…Actinfilin, an actin-binding protein of unknown cellular function, contains a BTB domain, six Kelch domain repeats, and a C-terminal PDZ binding motif (26,27). Using a yeast…”

Fetal Alz-50 Clone 1 Interacts with the Human Orthologue of the Kelch-like Ech-Associated Protein

“…However, several related proteins with Kelch domains and homologies to hKeap1 have been reported to have important neuronal functions. Both Actinfilin and Mayven are Kelch repeat containing proteins that bind to f-actin and are expressed in neurons (27,38). Another Kelch-containing protein, NRP/B, is involved in neurite outgrowth and neuronal differentiation (26).…”