The Rac1 hypervariable region in targeting and signaling

Lam, B. Daniel; Hordijk, Peter L.

doi:10.4161/sgtp.23310

Cited by 43 publications

(40 citation statements)

References 144 publications

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“…We found that the construct encoding the HV+CAAX sequence from Rac1 (C-tail) localizes to cell nucleus primarily, with some distribution at cell membrane and cytosol (Fig. S6A), as was previously reported (35,37,38). TIR-FRAP (total internal reflection fluorescence recovery after photobleaching) analysis of the membrane fraction showed an average recovery time constant of 8.5 ± 0.68 s, indicating that the molecules are dynamically exchanging with the cytosolic pool (Fig.…”

Section: C-term Hypervariable Region Is Important For the Regulation supporting

confidence: 53%

“…Studies have shown that the Rac1 binding to the plasma membrane requires prenylation of the C-terminal CAAX domain (7,8,36). However, the hypervariable (HV) region upstream of the CAAX sequence also modulates the subcellular localization of Rac1 (37,38). The affinity of the polybasic sequences in the HV region of Rac1 to different phospholipids has been extensively studied in vitro (39)(40)(41).…”

Section: C-term Hypervariable Region Is Important For the Regulation mentioning

confidence: 99%

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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: C-term Hypervariable Region Is Important For the Regulation supporting

confidence: 53%

Section: C-term Hypervariable Region Is Important For the Regulation mentioning

confidence: 99%

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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“…Rho GTPases share two common features which are crucial for their function: membrane anchorage and binary switch cycle (8). The GTPases can be located at the plasma membrane through their hypervariable region (HVR) and a lipid anchor (9). The binary switch determines whether Rho GTPases are in the active or inactive state.…”

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confidence: 99%

Unraveling the molecular mechanism of interactions of the Rho GTPases Cdc42 and Rac1 with the scaffolding protein IQGAP2

Ozdemir

Jang

Gürsoy

et al. 2018

Journal of Biological Chemistry

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IQ motif-containing GTPase-activating protein (IQGAP) scaffolding proteins play central roles in cell-cell adhesion, polarity, and motility. The Rho GTPases Cdc42 and Rac1, in their GTPbound active forms, interact with all three human IQGAPs. The IQGAP-Cdc42 interaction promotes metastasis by enhancing actin polymerization. However, despite their high sequence identity, Cdc42 and Rac1 differ in their interactions with IQGAP. Two Cdc42 molecules can bind to the Exdomain and the RasGAP site of the GTPaseactivating protein (GAP)-related domain (GRD) of IQGAP and promote IQGAP dimerization. Only one Rac1 molecule might bind to the RasGAP site of GRD and may not facilitate the dimerization, and the exact mechanism of Cdc42 and Rac1 binding to IQGAP is unclear. Using all-atom molecular dynamics simulations, site-directed mutagenesis, and Western blotting, we unraveled the detailed mechanisms of Cdc42 and Rac1 interactions with IQGAP2. We observed that Cdc42 binding to the Ex-domain of GRD of IQGAP2 (GRD2) releases the Ex-domain at the C-terminal region of GRD2, facilitating IQGAP2 dimerization. Cdc42 binding to the Ex-domain promoted allosteric changes in the RasGAP site, providing a binding site for the second Cdc42 in the RasGAP site. Of note, the Cdc42 "insert loop" was important for the interaction of the first Cdc42 with the Ex-domain. By contrast, differences in Rac1 insert loop sequence and structure precluded its interaction with the Ex-domain. Rac1 could bind only to the RasGAP site of apo-GRD2 and could not facilitate IQGAP2 dimerization. Our detailed mechanistic insights help decipher how Cdc42 can stimulate actin polymerization in metastasis.The Rho GTPase family is a subfamily of the Ras GTPase superfamily. Rho GTPases regulate multiple cellular processes such as growth, survival, cell invasion, cell motility and vesicle trafficking by interacting with effector molecules (1-4). To date, the mammalian Rho GTPase family includes 22 members; Cdc42, Rac1, and RhoA are the most studied among them (5).Rho GTPases are distinguished from other GTPases by the presence of an "insert loop" which is rich in charged residues in an α-helical motif (Fig. http://www.jbc.org/cgi

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“…The previous study identified that Rac1 could binding to SET, also known as TAF1β (template activating factor 1β), and suppress the function of PP2A by increasing the phosphorylation level in tumor cells [48, 49]. Also, it has been reported that in cardiac myocytes, Rac1/CDC42 complex promotes PP2A activity by dephosphorylating PP2A through upregulating p21-activated kinase-1 (Pak1) activity [50].…”

Section: Resultsmentioning

confidence: 99%

XIAP BIR domain suppresses miR-200a expression and subsequently promotes EGFR protein translation and anchorage-independent growth of bladder cancer cell

et al. 2017

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BackgroundThe X-linked inhibitor of apoptosis protein (XIAP) is a well-known potent apoptosis suppressor and also participates in cancer cell biological behaviors, therefore attracting great attentions as a potential antineoplastic therapeutic target for past years. Anti-IAP therapy is reported to be closely related to epidermal growth factor receptor (EGFR) expression level. However, whether and how XIAP modulates EGFR expression remains largely unknown.MethodsHuman XIAP was knockdown with short-hairpin RNA in two different bladder cancer cell lines, T24T and UMUC3. Two XIAP mutants, XIAP ∆BIR (deletion of N-terminal three BIR domains) and XIAP ∆RING (deletion of C-terminal RING domain and keeping the function of BIR domains), were generated to determine which domain is involved in regulating EGFR.ResultsWe found here that lacking of XIAP expression resulted in a remarkable suppression of EGFR expression, consequently leading to the deficiency of anchorage-independent cell growth. Further study demonstrated that BIR domain of XIAP was crucial for regulating the EGFR translation by suppressing the transcription and expression of miR-200a. Mechanistic studies indicated that BIR domain activated the protein phosphatase 2 (PP2A) activity by decreasing the phosphorylation of PP2A at Tyr307 in its catalytic subunit, PP2A-C. Such activated PP2A prevented the deviant phosphorylation and activation of MAPK kinases/MAPKs, their downstream effector c-Jun, and in turn inhibiting transcription of c-Jun-regulated the miR-200a.ConclusionsOur study uncovered a novel function of BIR domain of XIAP in regulating the EGFR translation, providing significant insight into the understanding of the XIAP overexpression in the cancer development and progression, further offering a new theoretical support for using XIAP BIR domain and EGFR as targets for cancer therapy.

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The Rac1 hypervariable region in targeting and signaling

Cited by 43 publications

References 144 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

Unraveling the molecular mechanism of interactions of the Rho GTPases Cdc42 and Rac1 with the scaffolding protein IQGAP2

XIAP BIR domain suppresses miR-200a expression and subsequently promotes EGFR protein translation and anchorage-independent growth of bladder cancer cell

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