Interaction of Zyxin, a Focal Adhesion Protein, with the E6 Protein from Human Papillomavirus Type 6 Results in Its Nuclear Translocation

Abstract

Zyxin, a focal adhesion molecule, interacts specifically with the E6 protein from human papillomavirus (HPV) type 6 in a yeast two-hybrid screen of a cDNA library prepared from human keratinocytes. Zyxin does not interact significantly with E6 proteins from HPV types 11, 16, or 18. The interaction was confirmed by in vitro and in vivo analyses and it requires the LIM domains (Lin-11, Isl-1, and Mec-3 [G. Freyd, S. K. Kim, and H. R. Horvitz, Nature 344:876-879, 1990]) found at the carboxyl terminus of zyxin. Cotransfection of E6 from HPV ( 6 E6) and zyxin results in the accumulation of zyxin in the nucleus where it can function as a transcriptional activator. 6 E6 can also mobilize endogenous zyxin to the nucleus.Human papillomaviruses (HPVs) are responsible for hyperproliferation of cutaneous and mucocutaneous epithelial cells that can lead to propagation of benign (30) or malignant (81) lesions depending on the virus type. The E6 and E7 proteins encoded by mucocutaneous high-risk types interact with p53 (65, 77) and the retinoblastoma (Rb) protein family (50), respectively, and transform cells in culture (10,35,39,49,74). In contrast, the E6 and E7 proteins encoded by low-risk viruses do not interact with these proteins and are not typically associated with events that lead to cell transformation (6).The E6 proteins encoded by HPVs contain about 150 amino acids and possess two Cys-X-X-Cys zinc fingers that bind zinc (5). While host proteins that interact with the E6 protein from both low-and high-risk HPVs (22,44,45,54,69) or only from high-risk HPVs have been identified (16, 26-28, 40, 42, 47, 57, 60, 70), no specific interaction between low-risk E6 and host proteins has been described. Here we report that low-risk E6 from HPV type 6 ( 6 E6) interacts with zyxin, a focal adhesion protein (7).Focal adhesion plaques are discrete areas on the cell membrane where the cells contact the underlying substratum or each other (36, 75). They are also the sites where multiple protein complexes involved in signaling assemble (15). Focal adhesions appear to represent transmembrane connections between the extracellular matrix and the cytoskeleton. Thus, it is not surprising that disrupted focal adhesions are frequently associated with the transformed phenotype (14). The E6 proteins from bovine papillomavirus and high-risk HPV interact with paxillin, another focal adhesion protein (13,70,71). This interaction may in part account for the disruption of actin fiber organization when bovine papillomavirus type 1 E6 is overexpressed in cells (70).Zyxin has features reminiscent of a signaling protein. Relative to the structural components of focal adhesions such as vinculin and ␣-actinin it is present at low abundance in cells and it is phosphorylated at multiple sites in vivo (18). Structurally, it has a proline-rich domain at its N terminus and multiple LIM ) domains in its carboxy-terminal half (8). Both domains are thought to be involved in protein binding (59,66). The proline-rich domain associates with SH3 domains that are f...

Retrograde Fluxes of Focal Adhesion Proteins in Response to Cell Migration and Mechanical Signals

“…A diffusion-like mechanism seems inefficient within a crowded cytoplasmic environment, and transport along the flux of actin filaments may serve as a much more effective mechanism. Particularly relevant is the relocation of zyxin from focal adhesions into the nucleus of vascular smooth muscle cells in response to mechanical signals, where zyxin itself may serve as a transcription activator (Degenhardt and Silverstein, 2001). In addition, retrograde flux of FAK may have a profound effect on the state of phosphorylation at focal adhesions and actin filaments, which in turn affects the maturation and turnover of focal adhesions (Webb et al, 2004).…”

“…Recent studies further indicated that mechanical stimulation can induce the change in distribution of zyxin from focal adhesions to either actin stress fibers in cultured fibroblasts (Yoshigi et al, 2005), or the nucleus of vascular smooth muscle cells (Cattaruzza et al, 2004). The latter may be related to the suspected ability of zyxin to regulate gene expression (Degenhardt and Silverstein, 2001). However, the mechanism of redistribution from focal adhesions is unclear, because these studies showed only the steady-state localization after mechanical stimulation.…”

The human papillomavirus 16 E6 protein can either protect or further sensitize cells to TNF: effect of dose

“…The first well-characterized activity of E6 was to bind to and accelerate the degradation of the p53 tumor suppressor, 2 but E6 also interacts with a number of additional cellular proteins that are engaged in a variety of cellular processes. These include proteins involved in the regulation of transcription, DNA replication and proliferation such as p300/CBP, 3,4 IRF-3, 5 hMcm 7, 6,7 E6TP1, 8 ADA3, 9,10 TRIP-Br1 11 and tuberin 12 ; proteins involved in apoptosis and immune evasion such as Bak, 13 c-Myc, 14,15 tumor necrosis factor (TNF) R1 16 and FADD 17 ; proteins involved with epithelial organization and differentiation such as paxillin, 18 E6BP/ERC-55, 19 zyxin 20 and fibulin-1; 21 proteins involved in cell-cell adhesion, polarity and proliferation control that contain a PDZ-binding motif such as hDLG, 22,23 hScrib, 24 MAGI-1, 25,26 MAGI-2, MAGI-3 27 and MUPP1; 28 and proteins involved in DNA repair such as XRCC1 29 and 6-O-methylguanine-DNA methyltransferase. 30 However, with some exceptions, the mechanisms and consequences of the interactions between E6 and its cellular partners for either the host or the virus are not well characterized.…”

The Human Papillomavirus 16 E6 Protein Binds to Fas-associated Death Domain and Protects Cells from Fas-triggered Apoptosis

“…12). Identified E6 partners include the following: proteins involved in the regulation of transcription and DNA replication, such as p300/CBP (13,14), , hMcm7 (16,17), E6TP1 (18), and ADA3 (19,20); proteins involved in apoptosis and immune evasion such as Bak (21), c-Myc (22), and TNF receptor 1 (TNF R1) (23); proteins involved with epithelial organization and differentiation such as paxillin (24), E6BP/ERC-55 (25), zyxin (26), HPV 6 and fibulin-1 (27); proteins involved in cell-cell adhesion, polarity, and proliferation control that contain a PDZ-binding motif such as hDLG (28,29), hScrib (30), MAGI-1 (31,32), MAGI-2, MAGI-3 (33), and MUPP1 (34); and proteins involved in DNA repair such as XRCC1 (35) and 6-O-methylguanine-DNA methyltransferase (36). However, with some exceptions, the mechanisms and the consequences of the interactions between E6 and its reported cellular partners on either the host or the virus life cycle are not well understood.…”