Prenylation-dependent Association of Ki-Ras with Microtubules

Abstract

We recently identified a prenyl peptide-binding protein in microsomal membranes from bovine brain (Thissen, J. A., and Casey, P. J. (1993)J. Biol. Chem. 268, 13780–13783). Through a variety of approaches, this binding protein has been identified as the cytoskeletal protein tubulin. Prenyl peptides bind to purified tubulin with a K d of 40 nm and also bind to tubulin polymerized into microtubules. Microtubule affinity chromatography of extracts from cells in which the prenyl protein pool was metabolically labeled revealed that prenyl proteins bound to the immobilized microtubules; one, a 24-kDa protein, was tentatively identified as a GTP-binding protein. Of several prenylated GTP-binding proteins tested, including Ki-Ras4B, Ha-Ras, RhoB, RhoA, and Rap1B, only Ki-Ras was found to bind significantly to microtubules, and this was in a prenylation-dependent fashion. A potential significance of the interaction of Ki-Ras4B with microtubules was indicated from analysis of the localization of newly synthesized Ki-Ras4B and Ha-Ras, each tagged with green fluorescence protein (GFP). Treatment of NIH-3T3 cells expressing GFP-Ki-Ras with Taxol (paclitaxel) resulted in accumulation of the expressed protein in intracellular locations, whereas in control cells the protein was correctly targeted to the plasma membrane. Importantly, such treatment with paclitaxel did not affect the cellular localization of expressed GFP-Ha-Ras. These results indicate that an intact microtubule network may be directly involved in Ki-Ras processing and/or targeting and provide direct evidence for a physiological distinction between Ki-Ras and Ha-Ras in cells. Additionally, the finding that paclitaxel treatment of cells disrupts Ki-Ras trafficking suggests an additional mechanism for the anti-proliferative effects of this drug. We recently identified a prenyl peptide-binding protein in microsomal membranes from bovine brain (Thissen, J. A., and Casey, P. J. (1993)J. Biol. Chem. 268, 13780–13783). Through a variety of approaches, this binding protein has been identified as the cytoskeletal protein tubulin. Prenyl peptides bind to purified tubulin with a K d of 40 nm and also bind to tubulin polymerized into microtubules. Microtubule affinity chromatography of extracts from cells in which the prenyl protein pool was metabolically labeled revealed that prenyl proteins bound to the immobilized microtubules; one, a 24-kDa protein, was tentatively identified as a GTP-binding protein. Of several prenylated GTP-binding proteins tested, including Ki-Ras4B, Ha-Ras, RhoB, RhoA, and Rap1B, only Ki-Ras was found to bind significantly to microtubules, and this was in a prenylation-dependent fashion. A potential significance of the interaction of Ki-Ras4B with microtubules was indicated from analysis of the localization of newly synthesized Ki-Ras4B and Ha-Ras, each tagged with green fluorescence protein (GFP). Treatment of NIH-3T3 cells expressing GFP-Ki-Ras with Taxol (paclitaxel) resulted in accumulation of the expressed protein in intracellular locations, whereas in control cells the protein was correctly targeted to the plasma membrane. Importantly, such treatment with paclitaxel did not affect the cellular localization of expressed GFP-Ha-Ras. These results indicate that an intact microtubule network may be directly involved in Ki-Ras processing and/or targeting and provide direct evidence for a physiological distinction between Ki-Ras and Ha-Ras in cells. Additionally, the finding that paclitaxel treatment of cells disrupts Ki-Ras trafficking suggests an additional mechanism for the anti-proliferative effects of this drug. Post-translational addition of isoprenoid lipids via a process termed prenylation is important in the maturation of many proteins that play critical roles in signal transduction and cell growth regulation. In many cases, a requirement for the attached isoprenoid for correct cellular localization and function has been demonstrated (for reviews, see Refs. 1Marshall C.J. Science. 1993; 259: 1865-1866Crossref PubMed Scopus (302) Google Scholar, 2Zhang F.L. Casey P.J. Annu. Rev. Biochem. 1996; 65: 241-269Crossref PubMed Scopus (1733) Google Scholar, 3Der C.J. Cox A.D. Cancer Cells. 1991; 3: 331-340PubMed Google Scholar). Most prenylation events involve addition of either a 15-carbon farnesyl or 20-carbon geranylgeranyl isoprenoid to a conserved Cys residue in a C-terminal sequence termed a “CAAX motif.” The X residue of the CAAX motif generally specifies which isoprenoid is to be attached to the protein by one of two cytosolic protein prenyltransferases, designated protein farnesyltransferase and protein geranylgeranyltransferase type I (4Casey P.J. Thissen J.A. Moomaw J.F. Proc. Natl. Acad. Sci. U. S. A. 1991; 88: 8631-8635Crossref PubMed Scopus (156) Google Scholar, 5Reiss Y. Goldstein J.L. Seabra M.C. Casey P.J. Brown M.S. Cell. 1990; 62: 81-88Abstract Full Text PDF PubMed Scopus (702) Google Scholar, 6Yokoyama K. Goodwin G.W. Ghomashchi F. Glomset J.A. Gelb M.H. Proc. Natl. Acad. Sci. U. S. A. 1991; 88: 5302-5306Crossref PubMed Scopus (217) Google Scholar). The addition of the isoprenoid lipid is followed by the proteolytic removal of the three C-terminal amino acids (the AAX) and methylation of the now C-terminal prenylated Cys residue in a process generally thought to occur in the microsomal membrane compartment of cells (7Jang G.F. Yokoyama K. Gelb M.H. Biochemistry. 1993; 32: 9500-9507Crossref PubMed Scopus (37) Google Scholar, 8Ashby M.N. King D.S. Rine J. Proc. Natl. Acad. Sci. U. S. A. 1992; 89: 4613-4617Crossref PubMed Scopus (98) Google Scholar, 9Perez-Sala D. Tan E.W. Canada F.J. Rando R.R. Proc. Natl. Acad. Sci. U. S. A. 1991; 88: 3043-3046Crossref PubMed Scopus (108) Google Scholar, 10Hancock J.F. Cadwallader K. Marshall C.J. EMBO J. 1991; 10: 641-646Crossref PubMed Scopus (249) Google Scholar). In addition to this sequence of protein modifications, many prenyl proteins contain either an upstream polybasic region or an attached palmitoyl lipid near the C terminus that influences membrane binding (11Kato K. Cox A.D. Hisaka M.M. Graham S.M. Buss J.E. Der C.J. Proc. Natl. Acad. Sci. U. S. A. 1992; 89: 6403-6407Crossref PubMed Scopus (555) Google Scholar, 12Hancock J.F. Paterson H. Marshall C.J. Cell. 1990; 63: 133-139Abstract Full Text PDF PubMed Scopus (844) Google Scholar). Several studies implicate prenylated proteins in regulation of cytoskeletal events. Two members of the Ras superfamily of G proteins, Rap1B and Rap1A, associate with the cytoskeleton during agonist-stimulated platelet activation (13Torti M. Ramaschi G. Sinigaglia F. Lapetina E.G. Balduini C. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 7553-7557Crossref PubMed Scopus (40) Google Scholar, 14Fischer T.H. Gatling M.N. McCormick F. Duffy C.M. White G.C. II J. Biol. Chem. 1994; 269: 17257-17261Abstract Full Text PDF PubMed Google Scholar). The related proteins Rac and Rho are thought to be important regulators of actin filament organization. Evidence to support this hypothesis comes primarily from studies demonstrating that Rac and Rho, respectively, are involved in regulation of growth factor-induced membrane ruffling and assembly of focal adhesion and actin stress fibers in Swiss 3T3 cells (15Ridley A.J. Paterson H.F. Johnston C.L. Diekman D. Hall A. Cell. 1992; 70: 401-410Abstract Full Text PDF PubMed Scopus (3071) Google Scholar, 16Ridley A.J. Hall A. Cell. 1992; 70: 389-399Abstract Full Text PDF PubMed Scopus (3824) Google Scholar). Rac1 has also been found to associate with tubulin in the GTP-bound form, the physiological significance is A. S. J. Biol. Chem. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar). The of prenylation in and is of the of the RhoA, with actin that be prenylated for this function K. H. A. Y. J. Cell. Biol. 1994; PubMed Scopus Google Scholar). is also in the for of prenylated proteins in by Ras be by which microtubule and Taxol which of microtubules Proc. Natl. Acad. Sci. U. S. A. 1994; PubMed Scopus Google Scholar, 1991; PubMed Scopus Google Scholar, G. 1996; Google Scholar). In one of cells with a cell that was by treatment of the cells with either or paclitaxel Proc. Natl. Acad. Sci. U. S. A. 1994; PubMed Scopus Google Scholar). evidence for an interaction between prenylated proteins and cellular microtubules comes from studies These such as and the of prenylated proteins and in cell A. J. 1992; Full Text PDF PubMed Google Scholar, Glomset J.A. A.J. J. Biol. PubMed Scopus Google Scholar). In one the of the in cells by was found to with the of the microtubules from the M. C. S.M. C. J. Cell. 1993; PubMed Scopus Google Scholar). of the cells with to the microtubules treatment with that intact microtubules for the M. C. S.M. C. J. Cell. 1993; PubMed Scopus Google Scholar). is now that prenylated proteins are with many membrane in including plasma and intracellular and the cytoskeleton J.F. Paterson H. Marshall C.J. Cell. 1990; 63: 133-139Abstract Full Text PDF PubMed Scopus (844) Google Scholar, P.J. Science. PubMed Scopus Google Scholar, Goldstein J.L. Brown M.S. J. Biol. Chem. 1994; 269: Full Text PDF PubMed Google Scholar, G. Balduini C. M. Sinigaglia F. 1994; PubMed Scopus Google Scholar). is newly prenylated proteins are from the to for processing to and that additional be involved in the correct targeting of In to involved in this a for proteins that bind prenylated peptides the C of prenylated binding resulted in the of a binding in a microsomal membrane that and peptides J.A. Casey P.J. J. Biol. Chem. 1993; Full Text PDF PubMed Google Scholar). The and of this binding with for a involved in targeting newly prenylated proteins to the compartment processing is We now analysis to a of the binding protein. the cellular protein identified that a prenylated was the microtubule protein which a of the cytoskeletal A variety of indicate a physiological significance for the of a prenylated protein, that Ki-Ras4B with microtubules. The between and and prenyl protein trafficking is farnesyl geranylgeranyl green protein and from and and from synthesized a and purified by chromatography and was from was from The was from and from was from was from was from and cell from was a from for Ha-Ras and Ki-Ras4B from Der of and respectively, and the for Rap1B was from of was as a protein by and was by NIH-3T3 cells with Ha-Ras was by Cox into cytosolic microsomal and plasma by as J.A. Casey P.J. J. Biol. Chem. 1993; Full Text PDF PubMed Google for to plasma membranes and for to cytosolic and microsomal membrane protein of in and a of J.F. F.L. Casey P.J. PubMed Scopus Google Scholar). the cytosolic and membrane in and J.A. Casey P.J. J. Biol. Chem. 1993; Full Text PDF PubMed Google Scholar). The to the C terminus of a G protein was prenylated and purified protein farnesyltransferase J.A. Casey P.J. J. Biol. Chem. 1993; Full Text PDF PubMed Google Scholar). A of the of and of in of and (4Casey P.J. Thissen J.A. Moomaw J.F. Proc. Natl. Acad. Sci. U. S. A. 1991; 88: 8631-8635Crossref PubMed Scopus (156) Google Scholar). The was with and was to a of the prenylated and the to for an additional The was purified a and GTP-binding proteins in Ha-Ras and purified as (4Casey P.J. Thissen J.A. Moomaw J.F. Proc. Natl. Acad. Sci. U. S. A. 1991; 88: 8631-8635Crossref PubMed Scopus (156) Google and was purified to the and and and purified to the GTP-binding protein was prenylated by the isoprenoid as (4Casey P.J. Thissen J.A. Moomaw J.F. Proc. Natl. Acad. Sci. U. S. A. 1991; 88: 8631-8635Crossref PubMed Scopus (156) Google Scholar). Two to peptides in this The a of of the prenyl and was in the of this whereas the results in and was in the of this In the peptides prenylated as S. J. Biol. Chem. 1990; Full Text PDF PubMed Google as by of of was in of followed by the addition of of The was and the prenylation by addition of farnesyl in of farnesyl of The was for and the was purified by and The involved the and farnesyl a in J. Chem. 1991; Scholar). a the was and the purified by analysis was as J.A. Casey P.J. PubMed Scopus Google Scholar). bovine brain microsomal membranes or of with prenyl in of A a was to the indicated the for and the by the addition of was with and the protein the prenylated was by and with proteins in and to PubMed Scopus Google and the for analysis as J.A. Casey P.J. PubMed Scopus Google Scholar). A of either the or the was this was of which been as the The was for followed by an with The was into a and as by the The was in and was purified from extracts of brain as 1991; Scopus Google Scholar). a brain was a of of of brain in and which also and the The was for to membranes and and the with and a to microtubules by for The the microtubules was a by between and The purified tubulin was purified to chromatography as J. Biol. Chem. 259: Full Text PDF PubMed Google Scholar). tubulin was in tubulin was with in and in a of a to a of and with of in the and was by binding was by the between bound to tubulin in the and of prenyl Microtubule affinity as C.M. J. Biol. PubMed Scopus Google Scholar). tubulin in was into microtubules by followed by the addition of the to the the of the the microtubules was to with and a of the microtubule to of the to the The was to and with several of and to The was with C and tubulin the was The was in the assembly and of the of cells J. M. J. Biol. Chem. Full Text PDF PubMed Google in and and with in and to In for cells in of for this the was by of of and and by with a into C and either or in of NIH-3T3 cells in with bovine and and in microtubule affinity chromatography cells by the addition of to and to followed by for and for The with C to the and to a microtubule affinity with C and and as in the Ha-Ras, RhoA, RhoA, and Rap1B proteins prenylated with as Ha-Ras and Ki-Ras and a microtubule affinity as for the cell that of was to the Ras as a protein. In proteins with of the microtubule affinity as for in C and the and with C and The was in and by as Most was to PubMed Scopus Google Scholar). of the and of tubulin was by a polymerized in the of Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar). Several tubulin in These an from a of the from M. of and two C-terminal from G. M.H. Cell. Full Text PDF PubMed Scopus Google Scholar). binding was the analysis of Ras proteins was binding was by an as Biochem. J. PubMed Scopus Google Scholar). by and to The was in and for The was in and for the the was for each in and to with an attached to proteins by analysis of the as P.J. Der C.J. Buss J.E. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar, Casey P.J. Glomset J.A. Gelb M.H. 1990; Scopus Google Scholar). prenyl proteins with and the proteins with The to followed by and by Ki-Ras4B was into to the GFP-Ki-Ras H. 1996; PubMed Scopus Google Scholar). Ha-Ras was into and was as a by a by The of Ki-Ras and Ha-Ras and the of the by analysis and of the and processing of to the in H. 1996; PubMed Scopus Google NIH-3T3 cells as and each three with an and and with paclitaxel or for The or into cells a and J. was three each 40 and the cells with with either paclitaxel or and and in the in a a In the by peptides to the C of prenylated proteins recently to with a protein in microsomal membranes from bovine brain J.A. Casey P.J. J. Biol. Chem. 1993; Full Text PDF PubMed Google Scholar). the involved in this a was Several for to a labeled to proteins in the microsomal membrane Most of the tested, which and the to a protein with an of whereas a protein with an of was J.A. Casey P.J. PubMed Scopus Google Scholar). the of the of to two proteins, analysis was The for binding to the and proteins, and was in the of either the the three C-terminal or a Additionally, the significantly for binding not These results indicated that the binding of the to two proteins was and that this interaction was the farnesyl as as the upstream region the and the three C-terminal amino to proteins in bovine brain membranes and protein and was with microsomal membranes of in the or of followed by with as proteins by in the of and with or with with of to proteins in bovine brain was with bovine brain of followed by the addition of the indicated of as and by of of the in A and was The binding membrane to not been to the cytosolic J.A. Casey P.J. J. Biol. Chem. 1993; Full Text PDF PubMed Google Scholar). the cytosolic to revealed two proteins with the as identified in the microsomal membranes of protein from microsomal membrane and proteins found to be in in the cytosolic in the microsomal membrane not We to the to the of proteins and to potential in cellular prenyl affinity chromatography has been as an in proteins involved in the two Y. Goldstein J.L. Seabra M.C. Casey P.J. Brown M.S. Cell. 1990; 62: 81-88Abstract Full Text PDF PubMed Scopus (702) Google Scholar, J.F. Casey P.J. J. Biol. Chem. 1992; Full Text PDF PubMed Google Scholar). We an affinity the to the cytosolic of bovine brain was the affinity proteins of and in a to that for the and proteins by Most the protein was significantly from the cytosolic proteins to the as revealed by a in the of this protein in the and A of the and proteins be from the affinity with and the with the These that two of each of proteins bound to the affinity one which be with and a that was In the only found to the bound proteins from the affinity was with The of binding of two proteins to the prenylated was demonstrated a affinity the which did not proteins the protein by the affinity was a protein in the bovine brain cytosolic The of this protein, in with that be as this protein is in brain this several tubulin to the from the prenyl affinity by analysis of from the affinity the of the protein as C results one such and the was with three additional tubulin The tubulin also the protein in the a of this was not The of tubulin by the prenyl affinity with to be to the indicated that tubulin a binding for the prenylated direct binding of to purified tubulin. binding was by the binding in the of an of The results of this analysis revealed that tubulin a binding for the with an d of 40 nm binding is the is with bovine brain an of tubulin that the of proteins in bovine brain not with the prenyl as the tubulin by the prenyl affinity and to bound to of from analysis revealed that the and of tubulin to the prenyl as by analysis of proteins to the two tubulin was also of to the also to microtubules or the binding the of tubulin. We microtubules from purified tubulin of the analysis revealed that the also to such microtubules not that the prenyl binding tubulin The results from the in studies the of the of prenylated proteins in the cell that bind to tubulin and/or microtubules. to such proteins, a microtubule affinity was this has been to a of proteins that associate with microtubules C.M. J. Biol. PubMed Scopus Google Scholar, G. A. D. Biol. Cell. 1992; 3: PubMed Scopus Google Scholar). for the of prenyl proteins in labeled the of which of the isoprenoid with to prenylated proteins J. M. J. Biol. Chem. Full Text PDF PubMed Google Scholar). cells with to prenylated proteins, to and the extracts and a microtubule affinity The of proteins the affinity and in the and of protein was by the microtubule affinity several and proteins be with of revealed that three prenylated proteins of and in the We which of the two or was attached to the The the three prenylated proteins to isoprenoid analysis only the 15-carbon farnesyl isoprenoid was of the prenylated proteins the contain the geranylgeranyl in the binding of proteins to microtubules. These the that are proteins that bind to microtubules, as such proteins may be in or not prenylated the to cells. prenylated proteins are GTP-binding proteins with in the of proteins are important regulators of cellular was of to of the prenylated proteins was a GTP-binding protein. cells as and the extracts by microtubule affinity chromatography and for binding an with this type of analysis only G proteins that GTP-binding is a in such analysis revealed the of a GTP-binding protein that from the microtubule affinity the as the prenylated proteins The of the GTP-binding proteins, did not associate with the microtubules as indicated by the of GTP-binding in the and and of a GTP-binding by the immobilized microtubules. evidence was by the finding that several a protein of in the not finding was not to the of the protein as in of with several Ras members not The results from the analysis revealed the of three proteins, one of which was a GTP-binding protein, that bind to microtubules. The of the was not in the binding of the proteins to microtubules was prenylation-dependent or this interaction was with proteins or a of related GTP-binding In an to to the of prenylated G proteins to with microtubules. Ki-Ras4B, which a polybasic sequence near the C-terminal prenyl and Ha-Ras, which has such polybasic region J.F. J.E. Marshall C.J. Cell. Full Text PDF PubMed Scopus Google Scholar). Ki-Ras and Ha-Ras in and a of protein in the of Ha-Ras, and of the Ki-Ras was These two of and Ras proteins the microtubule affinity as for the cell Ras proteins by of whereas Ras protein was by not Ha-Ras, was found to associate with the microtubule affinity in a and the bound proteins be with and The requirement for the farnesyl lipid was by the affinity chromatography with Ki-Ras and of proteins with the microtubule C and The of Ha-Ras to bind to microtubules was not to an additional requirement that this protein also be as Ha-Ras in NIH-3T3 which be and also did not bind to the microtubule affinity not The from studies was that microtubules in a prenylation-dependent a protein that a polybasic C-terminal the of Ki-Ras with microtubules was or prenylated GTP-binding proteins with also bind microtubules, several additional proteins Additionally, GTP-binding proteins that contain a polybasic sequence upstream from prenyl Cys residue are primarily and Ki-Ras also be Goldstein J.L. Brown M.S. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google the the of the microtubule binding for proteins the farnesyl isoprenoid be proteins, which RhoA, Rap1B, and Ki-Ras A for C-terminal sequence and isoprenoid of the GTP-binding proteins in A was prenylated with the and with an of microtubule affinity as The binding of proteins to the microtubule was by in the to that bound to the by and analysis of an of each The results of revealed that only either in a or with the microtubule revealed that the farnesyl and geranylgeranyl isoprenoid is by microtubules by the protein, The that proteins a polybasic C terminus and not bind to the microtubule suggests that the interaction between Ki-Ras and microtubules is a one and that in addition to the prenylated C terminus are involved in this Several for a significance of the interaction between Ki-Ras with microtubules. is that the microtubule network may play a in the processing and cellular targeting of is not the many prenyl proteins in cells are to correct cellular and is that to be identified are involved in this and studies that Ki-Ras is to the plasma membrane and that prenylated C and the polybasic region is for this localization J.F. Paterson H. Marshall C.J. Cell. 1990; 63: 133-139Abstract Full Text PDF PubMed Scopus (844) Google Scholar). an was in which of a protein between a green protein and Ki-Ras in cells demonstrated localization to the plasma membrane in cells H. 1996; PubMed Scopus Google Scholar). We this to the plasma membrane targeting of Ki-Ras be by of the microtubule NIH-3T3 cells with which disrupts the cells microtubule network by the microtubules, or with of an either the protein or GFP-Ha-Ras. The localization of the newly synthesized and proteins by fluorescence In control cells expressing paclitaxel treatment affect the localization of A and in cells not with the of GFP-Ki-Ras and to the plasma membrane C and In GFP-Ki-Ras synthesized in cells was found in intracellular locations, that not to the plasma membrane whereas paclitaxel treatment affect the localization of These results that an intact microtubule network is for the correct localization of not Ha-Ras, in cells. a in which a prenyl peptide-binding protein was in bovine brain J.A. Casey P.J. J. Biol. Chem. 1993; Full Text PDF PubMed Google a to a protein as the protein in this that the of this protein by prenyl affinity chromatography to as a cytoskeletal protein. of studies revealed that a of cellular prenylated proteins be immobilized microtubules, the of microtubules in cellular events by prenyl The that one of the cellular prenyl proteins that bind microtubules was a GTP-binding protein of and that several also a 24-kDa protein that in the to the of Ras proteins, and a variety of prenylated GTP-binding proteins, with microtubules. The finding that not Ha-Ras or a of related proteins, to microtubules in a prenylation evidence for of this interaction and for the of in addition to the prenylated C terminus for the of Ki-Ras with microtubules. A direct between Ki-Ras4B and microtubules in cells was by the finding that Ki-Ras4B is not correctly targeted to the plasma membrane in cells in which the network is by treatment with a paclitaxel treatment affect the cellular localization of the significance of this finding is not are several for Ki-Ras in cells is that an intact network is to newly prenylated Ki-Ras to the plasma membrane. In this the of the microtubule network by paclitaxel may a of binding for Ki-Ras that the protein from processing and targeting members of the prenylated protein are targeted to many membranes in that several are involved in the members of this of proteins processing and to in cells. is that in Ki-Ras that is newly prenylated not is targeted to the microtubules, a function processing and targeted to the plasma membrane. In this the of paclitaxel Ki-Ras localization may be to this the process the results from the a direct interaction between Ki-Ras and the microtubule network in cells. In to a between cell and in that a physiological of paclitaxel Ki-Ras targeting to the plasma membrane. Several studies in the of prenylated proteins, including in In one such by Ras by which of microtubules, and which microtubule Proc. Natl. Acad. Sci. U. S. A. 1994; PubMed Scopus Google Scholar). has also been to Ki-Ras in and cells in G. 1996; Google Scholar). studies that indicated a between Ras and microtubules the that paclitaxel affect the of activation of and was in whereas of has been in cells M. J. 70: PubMed Scopus Google Scholar, K. H. H. H. M. M. J. 63: PubMed Scopus Google Scholar). the for the effects of paclitaxel in the two has not been studies provide evidence for a between microtubules and which the In the provide evidence for of the network in the trafficking and/or cellular function of In this also direct evidence for a physiological distinction between Ha-Ras and Ki-Ras in cells. are to the by which paclitaxel treatment of cells targeting of Ki-Ras to the plasma membrane and/or for a potential function of from studies may provide additional as to function as We Moomaw and for of the protein and the Ras proteins, and and for with tubulin and for We Der and for Ras Cox for NIH-3T3 cells expressing Ras and for and for the and and for critical the