PTPN14 Degradation by High-Risk Human Papillomavirus E7 Limits Keratinocyte Differentiation and Contributes to HPV-Mediated Oncogenesis

Abstract

2 6 2 7 Classification: Biological Sciences / Microbiology 2 8 Abstract 3 0High-risk human papillomavirus (HPV) E7 proteins enable oncogenic transformation of HPV-3 1 infected cells by inactivating host cellular proteins. High-risk but not low-risk HPV E7 target 3 2 PTPN14 for proteolytic degradation, suggesting that PTPN14 degradation may be related to 3 3 their oncogenic activity. HPV infects human keratinocytes but the role of PTPN14 in 3 4 keratinocytes and the consequences of PTPN14 degradation are unknown. Using an HPV163 5 E7 variant that can inactivate RB1 but cannot degrade PTPN14 we found that high-risk HPV E7-3 6 mediated PTPN14 degradation impairs keratinocyte differentiation. Deletion of PTPN14 from 3 7 primary human keratinocytes decreased keratinocyte differentiation gene expression. Related to 3 8 oncogenic transformation, both HPV16 E7-mediated PTPN14 degradation and PTPN14 deletion 3 9 promoted keratinocyte survival following detachment from a substrate. PTPN14 degradation 4 0 contributed to high-risk HPV E6/E7-mediated immortalization of primary keratinocytes and HPV-4 1 positive but not HPV-negative cancers exhibit a gene expression signature consistent with 4 2 PTPN14 inactivation. We find that PTPN14 degradation impairs keratinocyte differentiation and 4 3 propose that this contributes to high-risk HPV E7-mediated oncogenic activity independent of 4 4 RB1 inactivation. 4 5 4 6 Significance Statement 4 7 Human papillomaviruses uncouple proliferation from differentiation in order to enable virus 4 8 replication in epithelial cells. HPV E7 proteins are well established to promote proliferation by 4 9 7 1 the proteasome-mediated degradation of RB1 (13-16). RB1 inactivation releases the inhibition 7 2 of E2F transcription factors, thus allowing cell cycle progression and acting as a major driver of 7 3proliferation. HPV E7 also promote proliferation by inhibiting the CDK inhibitors p21 WAF1/CIP1 and 7 4 p27 KIP1 (17)(18)(19). In addition to promoting proliferation, transcriptional studies indicate that human 7 5 cells harboring high-risk HPV genomes express lower levels of differentiation marker genes and 7 6 that both high-risk HPV E6 and E7 likely contribute to this repression (20-26). However, a 7 7 mechanism by which high-risk HPV E6 and/or E7 inhibit differentiation has not been defined. 7 8 RB1 binding by HPV E7 is necessary but insufficient for immortalization and 7 9 transformation, and several observations highlight the need for other contributors to 8 0 transformation. First, in multiple assays, the oncogenic activity of high-risk HPV E7 is disrupted 8 1 4 by mutations in regions that do not include the LxCxE motif (27-31). Second, low-risk HPV E7 8 2 bind RB1 but do not have activity in transformation assays and other E7 such as HPV1 E7 bind 8 3 RB1 with high affinity but do not transform (32-34). Finally, bovine papillomavirus (BPV) E7 8 4does not bind to RB1, but in some assays it is required for BPV-mediated transformation (30, 8 5 35-37). The idea that RB1 inactivation is ins...

Molecular Analysis of the Interaction between Human PTPN21 and the Oncoprotein E7 from Human Papillomavirus Genotype 18

“…PTPN14 or PTPD2 is a cytosolic protein that contains an N-terminal four-point-one, ezrin-radixin-moesin (FERM) domain, central proline-rich motifs, and a C-terminal catalytic PTP domain. Given that PTPN14 is an important antitumor protein that cooperates with the p53 transcription factor and negatively regulates Hippo signaling-involved cell proliferation (Huang et al, 2013;Liu et al, 2013;Mello et al, 2017;Michaloglou et al, 2013;Wilson et al, 2014), it is not surprising that this protein is targeted by HPV E7 via direct binding, which leads to its proteasomal degradation (Hatterschide et al, 2019;Szalmas et al, 2017;White et al, 2016;Yun et al, 2019). Our previous study also revealed that PTPN21 or PTPD1 can directly interact with CR3 of HPV18 E7 (Yun et al, 2019), consistent with previous systematic proteomic analyses (Rozenblatt-Rosen et al, 2012;White et al, 2012).…”

The human papillomavirus oncoproteins: a review of the host pathways targeted on the road to transformation

“…More recently, the interaction between E7 and UBR4 was found to be required for PTPN14 degradation and that this contributes towards delaying keratinocyte differentiation, as discussed previously in this section [212][213][214]. However, PTPN14 knockout also reduced detachmentinduced cell death, perhaps indicating that targeting of this pathway by HPV may concurrently impact upon differentiation and anoikis [214].…”

“…A diverse range of E7 proteins possess the ability to bind the non-receptor protein tyrosine phosphatase PTPN14, with HR-HPV E7 proteins additionally able to target it for proteasomal degradation using the UBR4 ubiquitin ligase [212,213]. This loss of PTPN14 is critical in both delaying the epithelial differentiation programme and for cellular transformation, although evidence that this is achieved via modulation of Hippo pathway signalling remains controversial [214,215]. Interestingly, mutational analyses indicate that this constitutes one of the as yet poorly defined pRb-independent functions of E7: regions in both CR1 and the C-terminus are necessary for formation of the E7/PTPN14/UBR4 complex.…”

The role of Hippo‐YAP signaling in squamous cell carcinomas

“…The HPV E6 protein prevents the YAP1 protein from undergoing proteasome‐mediated degradation 41 . HPV E7 protein binds to and targets PTPN14 for proteasome‐mediated degradation, which likely relieves the cytoplasmic retention of YAP by PTPN14 43,44 . This combination of HPV E6/E7‐induced accumulation of YAP protein plus its release to the nucleus leads to vigorous YAP activation that may promote human cervical epithelial cell proliferation and eventually CIS onset 23,42 .…”