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Maurine E Linder - One of the best experts on this subject based on the ideXlab platform.
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a stat3 Palmitoylation cycle promotes t h 17 differentiation and colitis
Nature, 2020Co-Authors: Maurine E Linder, Mingming Zhang, Lixing Zhou, Min Yang, Garrison Paul Komaniecki, Tatsiana Kosciuk, Xiao Chen, Xiaoping Zou, Hening LinAbstract:Cysteine Palmitoylation (S-Palmitoylation) is a reversible post-translational modification that is installed by the DHHC family of palmitoyltransferases and is reversed by several acyl protein thioesterases1,2. Although thousands of human proteins are known to undergo S-Palmitoylation, how this modification is regulated to modulate specific biological functions is poorly understood. Here we report that the key T helper 17 (TH17) cell differentiation stimulator, STAT33,4, is subject to reversible S-Palmitoylation on cysteine 108. DHHC7 palmitoylates STAT3 and promotes its membrane recruitment and phosphorylation. Acyl protein thioesterase 2 (APT2, also known as LYPLA2) depalmitoylates phosphorylated STAT3 (p-STAT3) and enables it to translocate to the nucleus. This Palmitoylation-dePalmitoylation cycle enhances STAT3 activation and promotes TH17 cell differentiation; perturbation of either Palmitoylation or dePalmitoylation negatively affects TH17 cell differentiation. Overactivation of TH17 cells is associated with several inflammatory diseases, including inflammatory bowel disease (IBD). In a mouse model, pharmacological inhibition of APT2 or knockout of Zdhhc7-which encodes DHHC7-relieves the symptoms of IBD. Our study reveals not only a potential therapeutic strategy for the treatment of IBD but also a model through which S-Palmitoylation regulates cell signalling, which might be broadly applicable for understanding the signalling functions of numerous S-Palmitoylation events.
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abstract b49 role of ras palmitoyl acyltransferase dhhc9 in hematopoiesis and nras leukemogenesis
Molecular Cancer Research, 2014Co-Authors: Bo Jiao, Maurine E LinderAbstract:Hyperactivation of RAS is common in human cancer, including hematological malignancies. Most oncogenic RAS proteins have been difficult to target. Identification of alternative targets that block RAS signaling is critical to develop therapies for RAS-related cancer. The biological activity of RAS proteins relies upon post-translational modifications (PTMs) that anchor RAS to cellular membranes. Protein Palmitoylation regulates the membrane targeting, subcellular trafficking, and functions of proteins. We have previously examined the importance of PTMs in NRAS leukemogenesis and found for the first time that Palmitoylation is essential for NRAS leukemogenesis. These studies suggest that targeting RAS Palmitoylation may be an effective therapy for cancers involving RAS proteins that rely on Palmitoylation for plasma membrane binding. In the previous studies, we blocked NRAS Palmitoylation by mutating the Palmitoylation site in NRAS. Therapeutic intervention of RAS Palmitoylation requires targeting enzymes that mediate RAS Palmitoylation. The reaction of protein S-Palmitoylation is catalyzed by a family DHHC protein palmitoyl-acyltransferases (PATs). Thus far 24 mamalian PATs have been identified. It has been shown that DHHC9 (a 364-amino acid protein encoded by ZDHHC9, an X-linked gene), is the ortholog of yeast Ras2 PAT and constitute a mammalian PAT with specificity for H- and NRAS in vitro. Increased expression of DHHC9 has been found in various cancer. Here we investigate the role of DHHC9 in hematopoiesis and NRAS leukemogenesis in vivo. We found that ectopic expression of DHHC9 in mouse bone marrow cells hinders hematopoiesis, but is incapable of inducing hematological malignancies in mice. We also found that frequency of lineage-specific populations and hematopoietic stem cell phenotype were similar in mice with knockout alleles of ZDHHC9 as that of wild type mice, suggesting that DHHC9 is dispensable for normal hematopoiesis. Expression of oncogenic NRAS in bone marrow cells from ZDHHC9 knockout mice still induced leukemia but the mice survived longer that the wild type control mice. These results suggest that DHHC9 plays a role in the pathogenesis of NRAS-induced leukemia, but it is not the only PATs for RAS Palmitoylation. We are identifying additional RAS PATs and developing cancer therapies targeting RAS Palmitoylation. Citation Format: Ping Liu, Bo Jiao, Maurine Linder, Ruibao Ren. Role of RAS palmitoyl-acyltransferase DHHC9 in hematopoiesis and NRAS leukemogenesis. [abstract]. In: Proceedings of the AACR Special Conference on RAS Oncogenes: From Biology to Therapy; Feb 24-27, 2014; Lake Buena Vista, FL. Philadelphia (PA): AACR; Mol Cancer Res 2014;12(12 Suppl):Abstract nr B49. doi: 10.1158/1557-3125.RASONC14-B49
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r7bp augments the function of rgs7 gβ5 complexes by a plasma membrane targeting mechanism
Journal of Biological Chemistry, 2006Co-Authors: Ryan M Drenan, Maurine E Linder, Craig A Doupnik, Muralidharan Jayaraman, Abigail Buchwalter, Kevin M Kaltenbronn, James E Huettner, Kendall J BlumerAbstract:the R7 family. We show that unpalmitoylated R7BP undergoes nuclear/cytoplasmic shuttling and that a C-terminal polybasic motif proximal to the Palmitoylation acceptor sites of R7BP mediates nuclear localization, Palmitoylation, and plasma membrane targeting. These results suggest a novel mechanism whereby palmitoyltransferases and nuclear import receptors both utilize the C-terminal domain of R7BP to determine the trafficking fate of R7G5R7BP heterotrimers. Analogous mechanisms may regulate other signaling proteins whose distribution between the plasma membrane and nucleus is controlled by Palmitoylation. Lastly, we show that cytoplasmic RGS7G5R7BP heterotrimers and RGS7G5 heterodimers are equivalently inefficient regulators of G proteincoupled receptor signaling relative to plasma membrane-bound heterotrimers bearing palmitoylated R7BP. Therefore, R7BP augments the function of the complex by a Palmitoylation-regulated plasma membrane-targeting mechanism.
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Palmitoylation of intracellular signaling proteins regulation and function
Annual Review of Biochemistry, 2004Co-Authors: Jessica E Smotrys, Maurine E LinderAbstract:▪ Abstract Protein S-Palmitoylation is the thioester linkage of long-chain fatty acids to cysteine residues in proteins. Addition of palmitate to proteins facilitates their membrane interactions and trafficking, and it modulates protein-protein interactions and enzyme activity. The reversibility of Palmitoylation makes it an attractive mechanism for regulating protein activity, and this feature has generated intensive investigation of this modification. The regulation of Palmitoylation occurs through the actions of protein acyltransferases and protein acylthioesterases. Identification of the protein acyltransferases Erf2/Erf4 and Akr1 in yeast has provided new insight into the Palmitoylation reaction. These molecules work in concert with thioesterases, such as acyl-protein thioesterase 1, to regulate the Palmitoylation status of numerous signaling molecules, ultimately influencing their function. This review discusses the function and regulation of protein Palmitoylation, focusing on intracellular protein...
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signalling functions of protein Palmitoylation
Biochimica et Biophysica Acta, 1998Co-Authors: Julianne T Dunphy, Maurine E LinderAbstract:Abstract Covalent lipid modifications anchor numerous signalling proteins to the cytoplasmic face of the plasma membrane. These modifications mediate protein–membrane and protein–protein interactions and are often essential for function. Protein Palmitoylation, due to its reversible nature, may be particularly important for modulating protein function during cycles of activation and deactivation. Despite intense investigation, the exact functions of protein Palmitoylation are not well understood. However, it is clear that Palmitoylation can affect a protein’s affinity for membranes, subcellular localization, and interactions with other proteins. In this review, recent advances in understanding the functions and mechanisms of protein Palmitoylation are discussed, with particular emphasis on how this lipid affects the biochemistry and cell biology of signalling proteins.
Michael J. Shipston - One of the best experts on this subject based on the ideXlab platform.
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Distinct Acyl Protein Transferases and Thioesterases Control Surface Expression of Calcium-activated Potassium Channels
Journal of Biological Chemistry, 2012Co-Authors: Lijun Tian, Heather Mcclafferty, Hans-guenther Knaus, Peter Ruth, Michael J. ShipstonAbstract:Abstract Protein Palmitoylation is rapidly emerging as an important determinant in the regulation of ion channels, including large conductance calcium-activated potassium (BK) channels. However, the enzymes that control channel Palmitoylation are largely unknown. Indeed, although Palmitoylation is the only reversible lipid modification of proteins, acyl thioesterases that control ion channel dePalmitoylation have not been identified. Here, we demonstrate that Palmitoylation of the intracellular S0–S1 loop of BK channels is controlled by two of the 23 mammalian palmitoyl-transferases, zDHHC22 and zDHHC23. Palmitoylation by these acyl transferases is essential for efficient cell surface expression of BK channels. In contrast, dePalmitoylation is controlled by the cytosolic thioesterase APT1 (LYPLA1), but not APT2 (LYPLA2). In addition, we identify a splice variant of LYPLAL1, a homolog with ∼30% identity to APT1, that also controls BK channel dePalmitoylation. Thus, both palmitoyl acyltransferases and acyl thioesterases display discrete substrate specificity for BK channels. Because depalmitoylated BK channels are retarded in the trans-Golgi network, reversible protein Palmitoylation provides a critical checkpoint to regulate exit from the trans-Golgi network and thus control BK channel cell surface expression.
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an electrostatic switch controls Palmitoylation of the large conductance voltage and calcium activated potassium bk channel
Journal of Biological Chemistry, 2012Co-Authors: Owen Jeffries, Lijun Tian, Heather Mcclafferty, Michael J. ShipstonAbstract:Protein Palmitoylation is a major dynamic posttranslational regulator of protein function. However, mechanisms that control Palmitoylation are poorly understood. In many proteins, Palmitoylation occurs at cysteine residues juxtaposed to membrane-anchoring domains such as transmembrane helices, sites of irreversible lipid modification, or hydrophobic and/or polybasic domains. In particular, polybasic domains represent an attractive mechanism to dynamically control protein Palmitoylation, as the function of these domains can be dramatically influenced by protein phosphorylation. Here we demonstrate that a polybasic domain immediately upstream of palmitoylated cysteine residues within an alternatively spliced insert in the C terminus of the large conductance calcium- and voltage-activated potassium channel is an important determinant of channel Palmitoylation and function. Mutation of basic amino acids to acidic residues within the polybasic domain results in inhibition of channel Palmitoylation and a significant right-shift in channel half maximal voltage for activation. Importantly, protein kinase A-dependent phosphorylation of a single serine residue within the core of the polybasic domain, which results in channel inhibition, also reduces channel Palmitoylation. These data demonstrate the key role of the polybasic domain in controlling stress-regulated exon Palmitoylation and suggests that phosphorylation controls the domain by acting as an electrostatic switch.
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ion channel regulation by protein Palmitoylation
Journal of Biological Chemistry, 2011Co-Authors: Michael J. ShipstonAbstract:Protein S-Palmitoylation, the reversible thioester linkage of a 16-carbon palmitate lipid to an intracellular cysteine residue, is rapidly emerging as a fundamental, dynamic, and widespread post-translational mechanism to control the properties and function of ligand- and voltage-gated ion channels. Palmitoylation controls multiple stages in the ion channel life cycle, from maturation to trafficking and regulation. An emerging concept is that Palmitoylation is an important determinant of channel regulation by other signaling pathways. The elucidation of enzymes controlling Palmitoylation and developments in proteomics tools now promise to revolutionize our understanding of this fundamental post-translational mechanism in regulating ion channel physiology.
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multiple palmitoyltransferases are required for Palmitoylation dependent regulation of large conductance calcium and voltage activated potassium channels
Journal of Biological Chemistry, 2010Co-Authors: Lijun Tian, Heather Mcclafferty, Owen Jeffries, Michael J. ShipstonAbstract:Palmitoylation is emerging as an important and dynamic regulator of ion channel function; however, the specificity with which the large family of acyl palmitoyltransferases (zinc finger Asp-His-His-Cys type-containing acyl palmitoyltransferase (DHHCs)) control channel Palmitoylation is poorly understood. We have previously demonstrated that the alternatively spliced stress-regulated exon (STREX) variant of the intracellular C-terminal domain of the large conductance calcium- and voltage-activated potassium (BK) channels is palmitoylated and targets the STREX domain to the plasma membrane. Using a combined imaging, biochemical, and functional approach coupled with loss-of-function (small interfering RNA knockdown of endogenous DHHCs) and gain-of-function (overexpression of recombinant DHHCs) assays, we demonstrate that multiple DHHCs control Palmitoylation of the C terminus of STREX channels, the association of the STREX domain with the plasma membrane, and functional channel regulation. Cysteine residues 12 and 13 within the STREX insert were the only endogenously palmitoylated residues in the entire C terminus of the STREX channel. Palmitoylation of this dicysteine motif was controlled by DHHCs 3, 5, 7, 9, and 17, although DHHC17 showed the greatest specificity for this site upon overexpression of the cognate DHHC. DHHCs that palmitoylated the channel also co-assembled with the channel in co-immunoprecipitation experiments, and knockdown of any of these DHHCs blocked regulation of the channel by protein kinase A-dependent phosphorylation. Taken together our data reveal that a subset of DHHCs controls STREX Palmitoylation and function and suggest that DHHC17 may preferentially target cysteine-rich domains. Finally, our approach may prove useful in elucidating the specificity of DHHC Palmitoylation of intracellular domains of other ion channels and transmembrane proteins.
Xu Wu - One of the best experts on this subject based on the ideXlab platform.
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Fatty acids and cancer-amplified ZDHHC19 promote STAT3 activation through S-Palmitoylation
Nature, 2019Co-Authors: Baoen Chen, Gopala K. Jarugumilli, Baohui Zheng, Sarah R Walker, David A Frank, Mari Mino-kenudson, Aaron N Hata, Xu WuAbstract:Signal transducer and activator of transcription 3 (STAT3) has a critical role in regulating cell fate, inflammation and immunity^ 1 , 2 . Cytokines and growth factors activate STAT3 through kinase-mediated tyrosine phosphorylation and dimerization^ 3 , 4 . It remains unknown whether other factors promote STAT3 activation through different mechanisms. Here we show that STAT3 is post-translationally S -palmitoylated at the SRC homology 2 (SH2) domain, which promotes the dimerization and transcriptional activation of STAT3. Fatty acids can directly activate STAT3 by enhancing its Palmitoylation, in synergy with cytokine stimulation. We further identified ZDHHC19 as a palmitoyl acyltransferase that regulates STAT3. Cytokine stimulation increases STAT3 Palmitoylation by promoting the association between ZDHHC19 and STAT3, which is mediated by the SH3 domain of GRB2. Silencing ZDHHC19 blocks STAT3 Palmitoylation and dimerization, and impairs the cytokine- and fatty-acid-induced activation of STAT3. ZDHHC19 is frequently amplified in multiple human cancers, including in 39% of lung squamous cell carcinomas. High levels of ZDHHC19 correlate with high levels of nuclear STAT3 in patient samples. In addition, knockout of ZDHHC19 in lung squamous cell carcinoma cells significantly blocks STAT3 activity, and inhibits the fatty-acid-induced formation of tumour spheres as well as tumorigenesis induced by high-fat diets in an in vivo mouse model. Our studies reveal that fatty-acid- and ZDHHC19-mediated Palmitoylation are signals that regulate STAT3, which provides evidence linking the deregulation of Palmitoylation to inflammation and cancer. The Palmitoylation of STAT3 is mediated by fatty acids and/or the palmitoyl acyltransferase ZDHHC19, and deregulation of this Palmitoylation has a role in inflammation and tumorigenesis.
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ZDHHC7-mediated S-Palmitoylation of Scribble regulates cell polarity
Nature Chemical Biology, 2016Co-Authors: Baoen Chen, Michael Deran, Gopala K. Jarugumilli, Yang S Brooks, Baohui Zheng, Jianjun Fu, Xu WuAbstract:Scribble (SCRIB) is a tumor-suppressor protein, playing critical roles in establishing and maintaining epithelial cell polarity. SCRIB is frequently amplified in human cancers but does not localize properly to cell-cell junctions, suggesting that mislocalization of SCRIB disrupts its tumor-suppressive activities. Using chemical reporters, here we showed that SCRIB localization was regulated by S-Palmitoylation at conserved cysteine residues. Palmitoylation-deficient mutants of SCRIB were mislocalized, leading to disruption of cell polarity and loss of their tumor-suppressive activities to oncogenic YAP, MAPK and PI3K/AKT pathways. We further found that ZDHHC7 was the major palmitoyl acyltransferase regulating SCRIB. Knockout of ZDHHC7 led to SCRIB mislocalization and YAP activation, and disruption of SCRIB's suppressive activities in HRas^V12-induced cell invasion. In summary, we demonstrated that ZDHHC7-mediated SCRIB Palmitoylation is critical for SCRIB membrane targeting, cell polarity and tumor suppression, providing new mechanistic insights of how dynamic protein Palmitoylation regulates cell polarity and tumorigenesis. The use of activity-based chemical probes revealed that Scribble is palmitoylated at cysteine residues by the palmitoyl acyltransferase ZDHHC7. Loss of Scribble Palmitoylation results in loss of cell polarity and its tumor suppressor activity.
Baoen Chen - One of the best experts on this subject based on the ideXlab platform.
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retraction note fatty acids and cancer amplified zdhhc19 promote stat3 activation through s Palmitoylation
Nature, 2020Co-Authors: Jixiao Niu, Baoen Chen, Gopala K. Jarugumilli, Baohui Zheng, Sarah R Walker, Aaron N Hata, Yang Sun, Mari Minokenudson, David A FrankAbstract:Signal transducer and activator of transcription 3 (STAT3) has a critical role in regulating cell fate, inflammation and immunity1,2. Cytokines and growth factors activate STAT3 through kinase-mediated tyrosine phosphorylation and dimerization3,4. It remains unknown whether other factors promote STAT3 activation through different mechanisms. Here we show that STAT3 is post-translationally S-palmitoylated at the SRC homology 2 (SH2) domain, which promotes the dimerization and transcriptional activation of STAT3. Fatty acids can directly activate STAT3 by enhancing its Palmitoylation, in synergy with cytokine stimulation. We further identified ZDHHC19 as a palmitoyl acyltransferase that regulates STAT3. Cytokine stimulation increases STAT3 Palmitoylation by promoting the association between ZDHHC19 and STAT3, which is mediated by the SH3 domain of GRB2. Silencing ZDHHC19 blocks STAT3 Palmitoylation and dimerization, and impairs the cytokine- and fatty-acid-induced activation of STAT3. ZDHHC19 is frequently amplified in multiple human cancers, including in 39% of lung squamous cell carcinomas. High levels of ZDHHC19 correlate with high levels of nuclear STAT3 in patient samples. In addition, knockout of ZDHHC19 in lung squamous cell carcinoma cells significantly blocks STAT3 activity, and inhibits the fatty-acid-induced formation of tumour spheres as well as tumorigenesis induced by high-fat diets in an in vivo mouse model. Our studies reveal that fatty-acid- and ZDHHC19-mediated Palmitoylation are signals that regulate STAT3, which provides evidence linking the deregulation of Palmitoylation to inflammation and cancer. The Palmitoylation of STAT3 is mediated by fatty acids and/or the palmitoyl acyltransferase ZDHHC19, and deregulation of this Palmitoylation has a role in inflammation and tumorigenesis.
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Fatty acids and cancer-amplified ZDHHC19 promote STAT3 activation through S-Palmitoylation
Nature, 2019Co-Authors: Baoen Chen, Gopala K. Jarugumilli, Baohui Zheng, Sarah R Walker, David A Frank, Mari Mino-kenudson, Aaron N Hata, Xu WuAbstract:Signal transducer and activator of transcription 3 (STAT3) has a critical role in regulating cell fate, inflammation and immunity^ 1 , 2 . Cytokines and growth factors activate STAT3 through kinase-mediated tyrosine phosphorylation and dimerization^ 3 , 4 . It remains unknown whether other factors promote STAT3 activation through different mechanisms. Here we show that STAT3 is post-translationally S -palmitoylated at the SRC homology 2 (SH2) domain, which promotes the dimerization and transcriptional activation of STAT3. Fatty acids can directly activate STAT3 by enhancing its Palmitoylation, in synergy with cytokine stimulation. We further identified ZDHHC19 as a palmitoyl acyltransferase that regulates STAT3. Cytokine stimulation increases STAT3 Palmitoylation by promoting the association between ZDHHC19 and STAT3, which is mediated by the SH3 domain of GRB2. Silencing ZDHHC19 blocks STAT3 Palmitoylation and dimerization, and impairs the cytokine- and fatty-acid-induced activation of STAT3. ZDHHC19 is frequently amplified in multiple human cancers, including in 39% of lung squamous cell carcinomas. High levels of ZDHHC19 correlate with high levels of nuclear STAT3 in patient samples. In addition, knockout of ZDHHC19 in lung squamous cell carcinoma cells significantly blocks STAT3 activity, and inhibits the fatty-acid-induced formation of tumour spheres as well as tumorigenesis induced by high-fat diets in an in vivo mouse model. Our studies reveal that fatty-acid- and ZDHHC19-mediated Palmitoylation are signals that regulate STAT3, which provides evidence linking the deregulation of Palmitoylation to inflammation and cancer. The Palmitoylation of STAT3 is mediated by fatty acids and/or the palmitoyl acyltransferase ZDHHC19, and deregulation of this Palmitoylation has a role in inflammation and tumorigenesis.
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ZDHHC7-mediated S-Palmitoylation of Scribble regulates cell polarity
Nature Chemical Biology, 2016Co-Authors: Baoen Chen, Michael Deran, Gopala K. Jarugumilli, Yang S Brooks, Baohui Zheng, Jianjun Fu, Xu WuAbstract:Scribble (SCRIB) is a tumor-suppressor protein, playing critical roles in establishing and maintaining epithelial cell polarity. SCRIB is frequently amplified in human cancers but does not localize properly to cell-cell junctions, suggesting that mislocalization of SCRIB disrupts its tumor-suppressive activities. Using chemical reporters, here we showed that SCRIB localization was regulated by S-Palmitoylation at conserved cysteine residues. Palmitoylation-deficient mutants of SCRIB were mislocalized, leading to disruption of cell polarity and loss of their tumor-suppressive activities to oncogenic YAP, MAPK and PI3K/AKT pathways. We further found that ZDHHC7 was the major palmitoyl acyltransferase regulating SCRIB. Knockout of ZDHHC7 led to SCRIB mislocalization and YAP activation, and disruption of SCRIB's suppressive activities in HRas^V12-induced cell invasion. In summary, we demonstrated that ZDHHC7-mediated SCRIB Palmitoylation is critical for SCRIB membrane targeting, cell polarity and tumor suppression, providing new mechanistic insights of how dynamic protein Palmitoylation regulates cell polarity and tumorigenesis. The use of activity-based chemical probes revealed that Scribble is palmitoylated at cysteine residues by the palmitoyl acyltransferase ZDHHC7. Loss of Scribble Palmitoylation results in loss of cell polarity and its tumor suppressor activity.
Martin E Hemler - One of the best experts on this subject based on the ideXlab platform.
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dhhc2 affects Palmitoylation stability and functions of tetraspanins cd9 and cd151
Molecular Biology of the Cell, 2008Co-Authors: Chandan Sharma, Xiuwei H Yang, Martin E HemlerAbstract:Although Palmitoylation markedly affects tetraspanin protein biochemistry and functions, relevant palmitoylating enzymes were not known. There are 23 mammalian “DHHC” (Asp-His-His-Cys) proteins, which presumably palmitoylate different sets of protein substrates. Among DHHC proteins tested, DHHC2 best stimulated Palmitoylation of tetraspanins CD9 and CD151, whereas inactive DHHC2 (containing DH→AA or C→S mutations within the DHHC motif) failed to promote Palmitoylation. Furthermore, DHHC2 associated with CD9 and CD151, but not other cell surface proteins, and DHHC2 knockdown diminished CD9 and CD151 Palmitoylation. Knockdown of six other Golgi-resident DHHC proteins (DHHC3, -4, -8, -17, -18, and -21) had no effect on CD9 or CD151. DHHC2 selectively affected tetraspanin Palmitoylation, but not the Palmitoylations of integrin β4 subunit and bulk proteins visible in [3H]palmitate-labeled whole cell lysates. DHHC2-dependent Palmitoylation also had multiple functional effects. First, it promoted physical associations between CD9 and CD151, and between α3 integrin and other proteins. Second, it protected CD151 and CD9 from lysosomal degradation. Third, the presence of DHHC2, but not other DHHC proteins, shifted cells away from a dispersed state and toward increased cell–cell contacts.
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Palmitoylation supports assembly and function of integrin tetraspanin complexes
Journal of Cell Biology, 2004Co-Authors: Xiuwei Yang, Oleg V Kovalenko, Christoph Claas, Christopher S Stipp, Wei Tang, Martin E HemlerAbstract:As observed previously, tetraspanin Palmitoylation promotes tetraspanin microdomain assembly. Here, we show that palmitoylated integrins (α3, α6, and β4 subunits) and tetraspanins (CD9, CD81, and CD63) coexist in substantially overlapping complexes. Removal of β4 Palmitoylation sites markedly impaired cell spreading and signaling through p130Cas on laminin substrate. Also in Palmitoylation-deficient β4, secondary associations with tetraspanins (CD9, CD81, and CD63) were diminished and cell surface CD9 clustering was decreased, whereas core α6β4–CD151 complex formation was unaltered. There is also a functional connection between CD9 and β4 integrins, as evidenced by anti-CD9 antibody effects on β4-dependent cell spreading. Notably, β4 Palmitoylation neither increased localization into “light membrane” fractions of sucrose gradients nor decreased solubility in nonionic detergents—hence it does not promote lipid raft association. Instead, Palmitoylation of β4 (and of the closely associated tetraspanin CD151) promotes CD151–α6β4 incorporation into a network of secondary tetraspanin interactions (with CD9, CD81, CD63, etc.), which provides a novel framework for functional regulation.
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Palmitoylation of tetraspanin proteins modulation of cd151 lateral interactions subcellular distribution and integrin dependent cell morphology
Molecular Biology of the Cell, 2002Co-Authors: Xiuwei Yang, Christoph Claas, Stinekathrein Kraeft, Lan Bo Chen, Zemin Wang, Jordan A Kreidberg, Martin E HemlerAbstract:Here we demonstrate that multiple tetraspanin (transmembrane 4 superfamily) proteins are palmitoylated, in either the Golgi or a post-Golgi compartment. Using CD151 as a model tetraspanin, we identified and mutated intracellular N-terminal and C-terminal cysteine Palmitoylation sites. Simultaneous mutations of C11, C15, C242, and C243 (each to serine) eliminated >90% of CD151 Palmitoylation. Notably, Palmitoylation had minimal influence on the density of tetraspanin protein complexes, did not promote tetraspanin localization into detergent-resistant microdomains, and was not required for CD151-α3β1 integrin association. However, the CD151 tetra mutant showed markedly diminished associations with other cell surface proteins, including other transmembrane 4 superfamily proteins (CD9, CD63). Thus, Palmitoylation may be critical for assembly of the large network of cell surface tetraspanin-protein interactions, sometimes called the “tetraspanin web.” Also, compared with wild-type CD151, the tetra mutant was much more diffusely distributed and showed markedly diminished stability during biosynthesis. Finally, expression of the tetra-CD151 mutant profoundly altered α3 integrin-deficient kidney epithelial cells, such that they converted from a dispersed, elongated morphology to an epithelium-like cobblestone clustering. These results point to novel biochemical and biological functions for tetraspanin Palmitoylation.
