C-terminal Di-arginine Motif of Cdc42 Protein Is Essential for Binding to Phosphatidylinositol 4,5-Bisphosphate-containing Membranes and Inducing Cellular Transformation

Johnson, Jared L.; Erickson, Jon W.; Cerione, Richard A.

doi:10.1074/jbc.m111.336487

Cited by 51 publications

(48 citation statements)

References 47 publications

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“…This finding is consistent with previous studies in Rac1GTPases, which provide evidence that the membrane translocation can occur independent of GEF activation, e.g., caRac1 exhibiting polarized distribution in lamellipodia (50), and Rac1 membrane association occurring at the boundaries of rafts and lipid rafts followed by diffusion into the rafts for GEF-mediated activation (56). Similarly, in other RhoGTPases, like cdc42, the critical role of protein−lipid interactions (via the polybasic domain) in modulating cell growth and transformation has also been demonstrated (57). The emergence of the sptPALM technique has made particle tracking in live cells relatively easy to implement (18,19,22).…”

Section: Discussionmentioning

confidence: 89%

Single-molecule tracking of small GTPase Rac1 uncovers spatial regulation of membrane translocation and mechanism for polarized signaling

Das

Yin

Yang

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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Polarized Rac1 signaling is a hallmark of many cellular functions, including cell adhesion, motility, and cell division. The two steps of Rac1 activation are its translocation to the plasma membrane and the exchange of nucleotide from GDP to GTP. It is, however, unclear whether these two processes are regulated independent of each other and what their respective roles are in polarization of Rac1 signaling. We designed a single-particle tracking (SPT) method to quantitatively analyze the kinetics of Rac1 membrane translocation in living cells. We found that the rate of Rac1 translocation was significantly elevated in protrusions during cell spreading on collagen. Furthermore, combining FRET sensor imaging with SPT measurements in the same cell, the recruitment of Rac1 was found to be polarized to an extent similar to that of the nucleotide exchange process. Statistical analysis of singlemolecule trajectories and optogenetic manipulation of membrane lipids revealed that Rac1 membrane translocation precedes nucleotide exchange, and is governed primarily by interactions with phospholipids, particularly PI(3,4,5)P 3 , instead of protein factors. Overall, the study highlights the significance of membrane translocation in spatial Rac1 signaling, which is in addition to the traditional view focusing primarily on GEF distribution and exchange reaction.super-resolution microscopy C ell polarization is critical for many biological processes, such as front−rear polarity during directed cell migration (chemotaxis, haptotaxis, wound healing, etc.), apical−basal polarity in epithelial cells, and axon specification in neuronal cells. The asymmetric distribution of signaling molecules, adhesion components, and cytoskeletal structures is important for the establishment of polarization. Among signaling proteins, the Rho family small GTPase Rac1 is ubiquitously required for cytoskeletal changes leading to a polarized morphology in many cells (1-4).Most small GTPases function as molecular switches, cycling between a GTP-bound active state and a GDP-bound inactive state. Activation of small GTPases typically requires guanine nucleotide exchange factors (GEFs) that promote the exchange of GDP for GTP. Inactivation requires the inherent GTPase activity, enhanced by the GTPase Activating Proteins (GAPs) (5, 6). Thus, each Rac1 molecule continuously cycles between the active and inactive state during their intracellular existence.Besides the guanine nucleotide binding cycle, another cycle that governs Rac1 activity is the membrane/cytoplasm translocation cycle. Rac1 GTPase, as with many other small GTPases, is posttranslationally prenylated at its C terminus and is capable of binding to lipid bilayers (7,8). Under basal conditions, the majority of Rac1 localizes to the cytoplasm in complex with RhoGDI molecules, which block its interactions with GEFs and GAPs and its downstream effectors (9-12). To become active, Rac1 needs to translocate to the membrane and be free from RhoGDI binding, allowing for its activation by GEFs and its inter...

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Section: Discussionmentioning

confidence: 89%

Single-molecule tracking of small GTPase Rac1 uncovers spatial regulation of membrane translocation and mechanism for polarized signaling

Das

Yin

Yang

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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“…Moreover, isoprenylated Cdc42 was found to bind directly and tightly to liposomes as long as they contain PI(4,5)P 2 (ref. 37).…”

Section: Discussionmentioning

confidence: 99%

HIV-1 Tat inhibits phagocytosis by preventing the recruitment of Cdc42 to the phagocytic cup

et al. 2015

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“…Previous studies on Ras, another prenylated protein, revealed that prenylation alone is not sufficient for stable plasma membrane association, and that additional palmitoylation (in the cases of N-Ras, H-Ras, and K-RasA) or positively charged residues (in the case of K-RasB) is required [30]. In the plasma membrane, negatively charged phospholipids such as phosphoinositides, phosphatidylinositol, and phosphatidylserine are asymmetrically enriched in the inner leaflets [36] and interact with positively charged residues [31]. We demonstrated here that ALDH3B1 is palmitoylated at Cys462 and Cys463 ( Figure 3D) and that the positively charged residues Arg462 and Arg463 are important for the plasma membrane localization of ALDH3B3 ( Figure 5B).…”

Section: Discussionmentioning

confidence: 99%

“…It has been reported that clusters of positively charged residues near prenylation sites are important for the plasma membrane localization of K-RasB and Cdc42 [30,31]. Therefore, to examine the role of Arg residues in the plasma membrane localization of ALDH3B3, we created a double Ala-substituted mutant (R462/463A), in which both the Arg462 and Arg463 residues were substituted with Ala residues.…”

Section: The C-terminal Regions Of Aldh3b2 and Aldh3b3 Determine Theimentioning

confidence: 99%

Mouse aldehyde dehydrogenase ALDH3B2 is localized to lipid droplets via two C-terminal tryptophan residues and lipid modification

Kitamura

Takagi

Naganuma

et al. 2014

Biochemical Journal

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Short title: Lipid droplet localization of ALDH3B2Summary statement: The mouse aldehyde dehydrogenases ALDH3B2 and ALDH3B3 exhibit similar substrate specificity but distinct intracellular localization (ALDH3B2, lipid droplets; ALDH3B3, plasma membrane). The C-terminal prenylation and two Trp residues are important for the lipid droplet localization of ALDH3B2.2 ABSTRACT Aldehyde dehydrogenases (ALDHs) catalyze the conversion of toxic aldehydes to non-toxic carboxylic acids. Of the 21 ALDHs in mice, it is the ALDH3 family members (ALDH3A1, ALDH3A2, ALDH3B1, ALDH3B2, and ALDH3B3) that are responsible for the removal of lipid-derived aldehydes. However, ALDH3B2 and ALDH3B3 have yet to be characterized.Here, we examined the enzyme activity, tissue distribution, and subcellular localization of ALDH3B2 and ALDH3B3. Both were found to exhibit broad substrate preferences from medium-chain to long-chain aldehydes, resembling ALDH3A2 and ALDH3B1. Although ALDH3B2 and ALDH3B3 share extremely high sequence similarity, their localizations differed, with ALDH3B2 found in lipid droplets and ALDH3B3 localized to the plasma membrane. Both ALDH3B2 and ALDH3B3 were modified by prenylation at their C-termini; this modification greatly influenced their membrane localization and enzymatic activity toward hexadecanal. We found that their C-terminal regions, particularly the two Trp residues (Trp462 and Trp469) of ALDH3B2 and the two Arg residues (Arg462 and Arg463) of ALDH3B3, were important for the determination of their specific localization. Abnormal quantity and perhaps quality of lipid droplets are implicated in several metabolic diseases. We speculate that ALDH3B2 acts to remove lipid-derived aldehydes in lipid droplets generated via oxidative stress as a quality control mechanism.

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C-terminal Di-arginine Motif of Cdc42 Protein Is Essential for Binding to Phosphatidylinositol 4,5-Bisphosphate-containing Membranes and Inducing Cellular Transformation

Cited by 51 publications

References 47 publications

Single-molecule tracking of small GTPase Rac1 uncovers spatial regulation of membrane translocation and mechanism for polarized signaling

Single-molecule tracking of small GTPase Rac1 uncovers spatial regulation of membrane translocation and mechanism for polarized signaling

HIV-1 Tat inhibits phagocytosis by preventing the recruitment of Cdc42 to the phagocytic cup

Mouse aldehyde dehydrogenase ALDH3B2 is localized to lipid droplets via two C-terminal tryptophan residues and lipid modification

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