Paralemmin, a Prenyl-Palmitoyl–anchored Phosphoprotein Abundant in Neurons and Implicated in Plasma Membrane Dynamics and Cell Process Formation

Abstract

We report the identification and initial characterization of paralemmin, a putative new morphoregulatory protein associated with the plasma membrane. Paralemmin is highly expressed in the brain but also less abundantly in many other tissues and cell types. cDNAs from chicken, human, and mouse predict acidic proteins of 42 kD that display a pattern of sequence cassettes with high inter-species conservation separated by poorly conserved linker sequences. Prenylation and palmitoylation of a COOH-terminal cluster of three cysteine residues confers hydrophobicity and membrane association to paralemmin. Paralemmin is also phosphorylated, and its mRNA is differentially spliced in a tissue-specific and developmentally regulated manner. Differential splicing, lipidation, and phosphorylation contribute to electrophoretic heterogeneity that results in an array of multiple bands on Western blots, most notably in brain. Paralemmin is associated with the cytoplasmic face of the plasma membranes of postsynaptic specializations, axonal and dendritic processes and perikarya, and also appears to be associated with an intracellular vesicle pool. It does not line the neuronal plasmalemma continuously but in clusters and patches. Its molecular and morphological properties are reminiscent of GAP-43, CAP-23, and MARCKS, proteins implicated in plasma membrane dynamics. Overexpression in several cell lines shows that paralemmin concentrates at sites of plasma membrane activity such as filopodia and microspikes, and induces cell expansion and process formation. The lipidation motif is essential for this morphogenic activity. We propose a function for paralemmin in the control of cell shape, e.g., through an involvement in membrane flow or in membrane–cytoskeleton interaction.

Paralemmin-1, a Modulator of Filopodia Induction Is Required for Spine Maturation

“…Consistent with a potential role for paralemmin-1 in filopodia induction, endogenous paralemmin-1 is detected in filopodia and spines in both immature (DIV 10) and mature (DIV 26) hippocampal neurons ( Figure 1A). Alternative splicing of paralemmin-1 is developmentally regulated (Kutzleb et al, 1998). The expression of a short splice variant (paralemmin-S) lacking exon 8 occurs early in development, preceding spine formation, Knockdown of paralemmin-1 influences the number of filopodia formed at DIV 7.…”

“…The following primary antibodies were used: green fluorescent protein (GFP; chicken; 1:1000; Abcam, Cambridge, MA), GluR1 (rabbit; 1:500; Upstate Biotech, Lake Placid, NY) and hemagglutinin (HA; mouse; 1:1000; Synaptic Systems, Gö ttingen, Germany). For endogenous paralemmin-1 detection, rabbit anti-paralemmin-1 sera 2 and 10 were used (Kutzleb et al, 1998). We used the following secondary antibodies: Alexa 488 -conjugated anti-chicken (1:1000, Molecular Probes, Eugene, OR), Alexa 568 -conjugated anti-mouse (1:1000, Molecular Probes), and Alexa 568 -conjugated anti-rabbit (1:1000, Molecular Probes).…”

“…Paralemmin-1 is abundantly expressed in the brain and concentrated at sites of plasma membrane activity, where it is anchored to the plasma membrane through lipid modifications. (Burwinkel et al, 1998;Kutzleb et al, 1998;Gauthier-Campbell et al, 2004;Castellini et al, 2005;Basile et al, 2006; This article was published online ahead of print in MBC in Press (http://www.molbiolcell.org/cgi/doi/10.1091/mbc.E07-08 -0802) on February 20, 2008February 20, . 2007).…”

“…Future experiments focused on identification of molecules that specifically associate with the paralemmin-1 isoform containing exon 8 may help clarify the differential effects induced by paralemmin-1 splice variants on spine maturation and AMPA receptor recruitment. Interestingly, the variant lacking exon 8 (paralemmin-S) is expressed at high levels at early stages of postnatal development, whereas the expression of the variant containing exon 8 (paralemmin-L) peaks at P14 (Kutzleb et al, 1998). Thus, sequential expression of paralemmin-1 splice variants may contribute to filopodia induction and their subsequent transformation to spines.…”

“…Paralemmin-1 is abundantly expressed in the brain and concentrated at sites of plasma membrane activity, where it is anchored to the plasma membrane through lipid modifications. (Burwinkel et al, 1998;Kutzleb et al, 1998;Gauthier-Campbell et al, 2004 ). This protein localizes to the plasma membranes of postsynaptic specializations, axonal and dendritic processes, and perikarya.…”