pak2 activation
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2020 ◽  
Vol 295 (9) ◽  
pp. 2640-2649 ◽  
Author(s):  
Madoka Kawaguchi ◽  
Nurmaa Dashzeveg ◽  
Yue Cao ◽  
Yuzhi Jia ◽  
Xia Liu ◽  
...  

CD44 molecule (CD44) is a well-known surface glycoprotein on tumor-initiating cells or cancer stem cells. However, its utility as a therapeutic target for managing metastases remains to be fully evaluated. We previously demonstrated that CD44 mediates homophilic interactions for circulating tumor cell (CTC) cluster formation, which enhances cancer stemness and metastatic potential in association with an unfavorable prognosis. Furthermore, CD44 self-interactions activate the P21-activated kinase 2 (PAK2) signaling pathway. Here, we further examined the biochemical properties of CD44 in homotypic tumor cell aggregation. The standard CD44 form (CD44s) mainly assembled as intercellular homodimers (trans-dimers) in tumor clusters rather than intracellular dimers (cis-dimers) present in single cells. Machine learning–based computational modeling combined with experimental mutagenesis tests revealed that the extracellular Domains I and II of CD44 are essential for its trans-dimerization and predicted high-score residues to be required for dimerization. Substitutions of 10 these residues in Domain I (Ser-45, Glu-48, Phe-74, Cys-77, Arg-78, Tyr-79, Ile-88, Arg-90, Asn-94, and Cys-97) or 5 residues in Domain II (Ile-106, Tyr-155, Val-156, Gln-157, and Lys-158) abolished CD44 dimerization and reduced tumor cell aggregation in vitro. Importantly, the substitutions in Domain II dramatically inhibited lung colonization in mice. The CD44 dimer-disrupting substitutions decreased downstream PAK2 activation without affecting the interaction between CD44 and PAK2, suggesting that PAK2 activation in tumor cell clusters is CD44 trans-dimer–dependent. These results shed critical light on the biochemical mechanisms of CD44-mediated tumor cell cluster formation and may help inform the development of therapeutic strategies to prevent tumor cluster formation and block cluster-mediated metastases.


2019 ◽  
Vol 218 (6) ◽  
pp. 1958-1971 ◽  
Author(s):  
Hannah K. Campbell ◽  
Alicia M. Salvi ◽  
Timothy O’Brien ◽  
Richard Superfine ◽  
Kris A. DeMali

Too little or too much force can trigger cell death, yet factors that ensure the survival of cells remain largely unknown. Here, we demonstrate that E-cadherin responds to force by recruiting and activating p21-activated protein kinase 2 (PAK2) to allow cells to stiffen, metabolize, and survive. Interestingly, PAK2 activation and its control of the apoptotic response are specific for the amplitude of force applied. Specifically, under low amplitudes of physiological force, PAK2 is protected from proteolysis, thereby ensuring cell survival. In contrast, under higher amplitudes of physiological force, PAK2 is left unprotected and stimulates apoptosis, an effect that is prevented by cleavage-resistant forms of the protein. Finally, we demonstrate that PAK2 protection is conferred by direct binding of AMPK. Thus, PAK2 mediates the survival of cells under force. These findings reveal an unexpected paradigm for how mechanotransduction, metabolism, and cell survival are linked.


Virology ◽  
2007 ◽  
Vol 358 (1) ◽  
pp. 119-135 ◽  
Author(s):  
Kristin Agopian ◽  
Bangdong L. Wei ◽  
J. Victor Garcia ◽  
Dana Gabuzda
Keyword(s):  
Mhc I ◽  
Hiv 1 ◽  

2006 ◽  
Vol 80 (6) ◽  
pp. 3050-3061 ◽  
Author(s):  
Kristin Agopian ◽  
Bangdong L. Wei ◽  
J. Victor Garcia ◽  
Dana Gabuzda

ABSTRACT The interaction of human immunodeficiency virus type 1 (HIV-1) Nef with p21-activated kinase 2 (Pak2) has been proposed to play an important role in T-cell activation and disease progression during viral infection. However, the mechanism by which Nef activates Pak2 is poorly understood. Mutations in most Nef motifs previously reported to be required for Pak2 activation (G2, PxxP72, and RR105) also affect other Nef functions, such as CD4 or major histocompatibility complex class I (MHC-I) downregulation. To better understand Nef interactions with Pak2, we performed mutational analysis of three primary HIV-1 Nef clones that exhibited similar capacities for downregulation of CD4 and MHC-I but variable abilities to associate with activated Pak2. Our results demonstrate that Nef amino acids at positions 85, 89, 187, 188, and 191 (L, H, S, R, and F in the clade B consensus, respectively) are critical for Pak2 association. Mutation of these Nef residues dramatically altered association with Pak2 without affecting Nef expression levels or CD4 and MHC-I downregulation. Furthermore, compensation occurred at positions 89 and 191 when both amino acids were substituted. Since residues 85, 89, 187, 188, and 191 cluster on the surface of the Nef core domain in a region distinct from the dimerization and SH3-binding domains, we propose that these Nef residues form part of a unique binding surface specifically involved in association with Pak2. This binding surface includes exposed and recessed hydrophobic residues and may participate in an as-yet-unidentified protein-protein interaction to facilitate Pak2 activation.


2005 ◽  
Vol 79 (23) ◽  
pp. 14976-14980 ◽  
Author(s):  
Bangdong L. Wei ◽  
Vivek K. Arora ◽  
Alexa Raney ◽  
Lillian S. Kuo ◽  
Guang-Hui Xiao ◽  
...  

ABSTRACT The accessory human immunodeficiency virus type 1 (HIV-1) protein Nef activates the autophosphorylation activity of p21-activated kinase 2 (PAK2). Merlin, a cellular substrate of PAK2, is homologous to the ezrin-radixin-moesin family and plays a critical role in Rac signaling. To assess the possible impact on host cell metabolism of Nef-induced PAK2 activation, we investigated the phosphorylation of merlin in Nef expressing cells. Here we report that Nef induces merlin phosphorylation in multiple cell lines independently of protein kinase A. This intracellular phosphorylation of merlin directly correlates with in vitro assay of the autophosphorylation activity of Nef-activated PAK2. Importantly, merlin phosphorylation induced by Nef was also observed in human primary T cells. The finding that Nef induces phosphorylation of the key signaling molecule merlin suggests several possible roles for PAK2 activation in HIV pathogenesis.


2004 ◽  
Vol 78 (23) ◽  
pp. 12773-12780 ◽  
Author(s):  
Kati Pulkkinen ◽  
G. Herma Renkema ◽  
Frank Kirchhoff ◽  
Kalle Saksela

ABSTRACT We have previously reported that Nef specifically interacts with a small but highly active subpopulation of p21-activated kinase 2 (PAK2). Here we show that this is due to a transient association of Nef with a PAK2 activation complex within a detergent-insoluble membrane compartment containing the lipid raft marker GM1. The low abundance of this Nef-associated kinase (NAK) complex was found to be due to an autoregulatory mechanism. Although activation of PAK2 was required for assembly of the NAK complex, catalytic activity of PAK2 also promoted dissociation of this complex. Testing different constitutively active PAK2 mutants indicated that the conformation associated with p21-mediated activation rather than kinase activity per se was required for PAK2 to become NAK. Although association with PAK2 is one of the most conserved properties of Nef, we found that the ability to stimulate PAK2 activity differed markedly among divergent Nef alleles, suggesting that PAK2 association and activation are distinct functions of Nef. However, mutations introduced into the p21-binding domain of PAK2 revealed that p21-GTPases are involved in both of these Nef functions and, in addition to promoting PAK2 activation, also help to physically stabilize the NAK complex.


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