Cutaneous Human Papillomavirus Type 38 E7 Regulates Actin Cytoskeleton Structure for Increasing Cell Proliferation through CK2 and the Eukaryotic Elongation Factor 1A

Abstract

We previously reported that the oncoproteins E6 and E7 from cutaneous human papillomavirus type 38 (HPV38) can immortalize primary human keratinocytes in vitro and sensitize transgenic mice to develop skin cancer in vivo. Immunofluorescence staining revealed that human keratinocytes immortalized by HPV38 E6 and E7 display fewer actin stress fibers than do control primary keratinocyte cells, raising the possibility of a role of the viral oncoproteins in the remodeling of the actin cytoskeleton. In this study, we show that HPV38 E7 induces actin stress fiber disruption and that this phenomenon correlates with its ability to downregulate Rho activity. The downregulation of Rho activity by HPV38 E7 is mediated through the activation of the CK2-MEK-extracellular signal-regulated kinase (ERK) pathway. In addition, HPV38 E7 is able to induce actin fiber disruption by binding directly to eukaryotic elongation factor 1A (eEF1A) and abolishing its effects on actin fiber formation. Finally, we found that the downregulation of Rho activity by HPV38 E7 through the CK2-MEK-ERK pathway facilitates cell growth proliferation. Taken together, our data support the conclusion that HPV38 E7 promotes keratinocyte proliferation in part by negatively regulating actin cytoskeleton fiber formation through the CK2-MEK-ERK-Rho pathway and by binding to eEF1A and inhibiting its effects on actin cytoskeleton remodeling.Human papillomaviruses (HPVs) are double-stranded small DNA viruses, comprising more than 100 members, that are the causative agents of several human diseases, including cancers (25). According to their tropisms, HPVs are subdivided into mucosal and cutaneous types. High-risk mucosal HPV types, such as HPV type 16 (HPV16) and HPV18, are etiological agents of cervical cancer and other anogenital cancers (25). Their ability to induce cellular immortalization and transformation is attributed primarily to the viral oncoproteins E6 and E7 (20,25). While E6 prevents apoptosis by inducing the degradation of the tumor suppressor p53 through the proteasome system, E7 disrupts cell cycle regulation by inactivating pRb (20,25). In addition, HPV16 E6 and E7 alter several other cellular signaling pathways by interacting with a number of cellular proteins, enhancing their carcinogenic properties (20,38,39,41).A large subgroup of cutaneous HPV types belonging to the beta genus of the HPV phylogenetic tree has been proposed to be involved in the development of nonmelanoma skin cancer (NMSC), since they were isolated for the first time from patients suffering from a rare autosomal recessive genetic disorder called epidermodysplasia verruciformis (EV), characterized by susceptibility to beta HPV infection and NMSC development at sun-exposed anatomical regions (7). Subsequent epidemiological studies have also provided lines of evidence for a possible oncogenic role of beta HPV types in non-EV patients, including the healthy population and immunocompromised individuals, e.g., organ transplant recipients (7, 51).Our group has previously reported t...

The Human Papillomavirus E7 Proteins Associate with p190RhoGAP and Alter Its Function

“…Therefore, we compared the status of stress fibers in cells expressing HPV16 E7 to those of control, GFP-expressing cells and cell expressing dominant negative p190 (p190RA). For the purposes of these functional assays, we utilized human osteosarcoma U2OS cells, since they were previously utilized for similar assays and were shown to behave similarly to human keratinocytes in these experiments (47). In addition, we also confirmed the results of these assays in human keratinocyte HaCat cells.…”

“…The HPV16 E7 reduction in RhoA-GTP was shown to be due in part to E7 modulation of the cytoplasmic localization of p27, which can in turn bind and dysregulate RhoA (46). On the other hand, HPV38 E7 uses two distinct mechanisms to downregulate Rho activity: (i) mediating the activation of the CK2-MEK-extracellular signalregulated kinase (ERK) pathway and (ii) directly binding to eukaryotic elongation factor 1A (eEF1A) and abolishing its effects on actin fiber formation (47). Due in part to these published observations and considering that p190 is a direct regulator of the Rho pathway, we wanted to assess the biological consequences of E7-p190 interaction.…”

The Interaction between Human Papillomavirus Proteins and Cytoskeletal Filaments

“…In addition, HPV38 E7 is able to induce actin fiber disruption by directly binding to the eukaryotic elongation factor 1A (eEF1A) and abolishing its effects on actin fiber formation. Their data support the conclusion that HPV38 E7 promotes keratinocyte proliferation in part by negatively regulating actin cytoskeleton fiber formation and by binding to eEF1A and inhibiting its effects on actin cytoskeleton remodeling (Yue et al, 2011). Cells from laryngeal papillomas and normal epithelium were cultured in vitro.…”

“…HPV proteins are interacted with cell skeletal filaments. Some of these proteins are E6 (HPV16,18), E7 (HPV16,18,38), E5 (high and low risk HPV types), E1^E4 (HPV16), E4 proteins (Lee & Dominguez, 2010;McIntosh et al, 2010;Nguyen 2008;Rey et al, 2000, Safi Oz, 2010Stanley, 2001;Uribe & Jay., 2009;Yue et al, 2011).…”

The Ubiquitin-like Protein FAT10 Stabilizes eEF1A1 Expression to Promote Tumor Proliferation in a Complex Manner

“…We first used HEK 293T cells to determine which sites in eEF1A1 binding to FAT10. eEF1A1 was divided into various structural domains, C-terminally truncated eEF1A1 mutants and truncated eEF1A1 fragments (24,25). Co-IP experiments indicated that the eEF1A1 region in aa 284-332 and aa 394-462 contains the FAT10 interaction region (Fig.…”