The EF-hand Ca²⁺-binding Protein p22 Associates with Microtubules in an N-Myristoylation–dependent Manner

Abstract

Proteins containing the EF-hand Ca(2+)-binding motif, such as calmodulin and calcineurin B, function as regulators of various cellular processes. Here we focus on p22, an N-myristoylated, widely expressed EF-hand Ca(2+)-binding protein conserved throughout evolution, which was shown previously to be required for membrane traffic. Immunofluorescence studies show that p22 distributes along microtubules during interphase and mitosis in various cell lines. Moreover, we report that p22 associates with the microtubule cytoskeleton indirectly via a cytosolic microtubule-binding factor. Gel filtration studies indicate that the p22-microtubule-binding activity behaves as a 70- to 30-kDa globular protein. Our results indicate that p22 associates with microtubules via a novel N-myristoylation-dependent mechanism that does not involve classic microtubule-associated proteins and motor proteins. The association of p22 with microtubules requires the N-myristoylation of p22 but does not involve p22's Ca(2+)-binding activity, suggesting that the p22-microtubule association and the role of p22 in membrane traffic are functionally related, because N-myristoylation is required for both events. Therefore, p22 is an excellent candidate for a protein that can mediate interactions between the microtubule cytoskeleton and membrane traffic.

Acute Regulation of Na/H Exchanger NHE3 by Adenosine A1 Receptors Is Mediated by Calcineurin Homologous Protein

“…Cells were exposed to transfection reagent either without siRNA (Untransfected) or with siRNA against CHP (CHP siRNA), protein tyrosine kinase (Pyk2 siRNA) or non-mammalian protein (GL2 siRNA). been suggested to influence membrane trafficking by effecting either directly the components of the vesicle transport machinery or indirectly the organization of the cytoskeleton (30). NHE3 is sensitive to the organization of the cytoskeleton (70).…”

“…The ability of CHP to alter NHE1 transport independent of surface NHE1 protein (28) renders it an attractive candidate for supporting adenosine-mediated change in intrinsic transport activity of NHE3. CHP associates with microtubules via an N-myristoylation-dependent mechanism that does not involve conventional microtubule-associated proteins (30) and is involved in vesicular transport (25). CHP also interacts with kinesin-related protein KIF1B␤2, a molecular motor in neurons (31).…”

Expression of Calcineurin B Homologous Protein 2 Protects Serum Deprivation-induced Cell Death by Serum-independent Activation of Na+/H+ Exchanger

“…Ubiquitous CHP1 was previously suggested to be involved in various cell functions, such as inhibition of calcineurin activity (37), vesicular transport of proteins (38), interaction with microtubules (39), and interaction with a death-associated protein kinase-related apoptosis-inducing protein kinase 2 (DRAK2) (40). Although we do not know whether CHP2 is also involved in these cellular functions, they may not be relevant to the observed effects of CHP2 because the functions of CHP1/NHE1 and CHP2/NHE1 were compared in this study.…”

Solution Structure of the Cytoplasmic Region of Na+/H+ Exchanger 1 Complexed with Essential Cofactor Calcineurin B Homologous Protein 1

“…3B). The secondary structure consists of ␣A (residues [11][12][13][14][15][16][17][18][19][20][21][22], ␣B (residues 26 -37), ␣C (residues 48 -51), ␣D (residues 64 -70), ␣E (residues 80 -88), ␣F (residues 111-122), ␣G (residues 132-143), ␣H (residues 149 -162), ␣I (residues 174 -180), and ␣J (residues 185-188) (Figs. 1B and 3B).…”