The F-BAR Protein CIP4 Inhibits Neurite Formation by Producing Lamellipodial Protrusions

Abstract

Summary Neurite formation is a seminal event in the early development of neurons. However, little is known about the mechanisms by which neurons form neurites. F-BAR proteins function in sensing and inducing membrane curvature [1, 2]. Cdc42-interacting protein 4 (CIP4), a member of the F-BAR family, regulates endocytosis in a variety of cell types [3–9]. However, there is little data on how CIP4 functions in neurons [10, 11]. Here we show that CIP4 plays a novel role in neuronal development by inhibiting neurite formation. Remarkably, CIP4 exerts this effect not through endocytosis, but by producing lamellipodial protrusions. In primary cortical neurons CIP4 is concentrated specifically at the tips of extending lamellipodia and filopodia, instead of endosomes as in other cell types. Overexpression of CIP4 results in lamellipodial protrusions around the cell body, subsequently delaying neurite formation and enlarging growth cones. These effects depend on the F-BAR and SH3 domains of CIP4 and on its ability to multimerize. Conversely, cortical neurons from CIP4-null mice initiate neurites twice as fast as controls. This is the first study to demonstrate that an F-BAR protein functions differently in neuronal vs. non-neuronal cells and induces lamellipodial protrusions instead of invaginations or filopodia-like structures.

Inhibition of TFG function causes hereditary axon degeneration by impairing endoplasmic reticulum structure

“…Cultures were rinsed in PBS solution and blocked with 10% (vol/vol) BSA/PBS solution, permeabilized in 0.2% Triton X-100/PBS solution, and labeled with anti-TFG or anti-Sec16A antibodies and anti-tyrosinated tubulin and Alexa 488-or 546-conjugated secondary antibodies. For wide-field imaging, neurons were imaged on a Nikon TE2000 inverted microscope as described previously (59). Imaging of RPE1 and COS7 cells was conducted following fixation by using a similar procedure to that described for neuron cultures.…”

CIP4 coordinates with phospholipids and actin-associated proteins to localize to the protruding edge and produce actin ribs and veils

“…In nonneuronal cells, CIP4 induces polymerization associated with actin comet tails, endocytosis, and tubulation through its interaction with Cdc42, N-WASP and dynamin (Hartig et al, 2009). Recently, we showed that in cortical neurons CIP4 localizes to the protruding edges of immature neurons and the growth cones of polarized neurons (Saengsawang et al, 2012). We also showed that CIP4 induces lamellipodia and veil formation and increased the density of long, thin actin filaments (defined as actin ribs) around neuronal cell bodies.…”

“…Although the most characterized function of F-BAR proteins is their role in endocytosis, many F-BAR proteins including CIP4 have also been shown to play a role in filopodial and lamellipodial protrusion (Carlson et al, 2011;Guerrier et al, 2009;Lee et al, 2010;Saengsawang et al, 2012;Zaidel-Bar et al, 2010). In nonneuronal cells, CIP4 induces polymerization associated with actin comet tails, endocytosis, and tubulation through its interaction with Cdc42, N-WASP and dynamin (Hartig et al, 2009).…”

Using fly genetics to dissect the cytoskeletal machinery of neurons during axonal growth and maintenance

“…For this, MTs in the neuronal cell bodies become stabilised and push beyond the cell cortex (Brandt, 1998). This is assisted by dynamic rearrangements of the cortical actin cytoskeleton to remove barrier function (Flynn et al, 2012;Neukirchen and Bradke, 2011;Saengsawang et al, 2012) and the formation of filopodia, which provide membrane protrusions into which MTs can extend Gonçalves-Pimentel et al, 2011). In typical vertebrate neurons, several neurites are formed simultaneously on the cell body, but only one of these is selected as the axon.…”