IFN-κ, a novel type I IFN, is undetectable in HPV-positive human cervical keratinocytes

Abstract

Interferons (IFNs) are expressed by many cell types and play a pivotal role in the generation of immune responses against viral infections. IFN-k, a novel type I IFN, displays a tight tropism for keratinocytes and specific lymphoid populations and exhibits functional similarities with other type I IFNs. The human papillomavirus (HPV), the etiological agent for cervical cancer, infects keratinocytes of the uterine cervix and has been shown to directly inhibit the IFN pathway. We evaluated IFN-k, -b, and -g gene expression in HPV-negative normal and HPV-positive pre-malignant and malignant ex vivo cervical tissue covering the entire spectrum of cervical disease. Quantitative real-time polymerase chain reaction and methods previously optimized for detecting low-expressing genes in cervical tissue were used. In contrast to IFN-b and -g, IFN-k mRNA prevalence and levels were unexpectedly higher in diseased compared with normal whole cervical tissue with highest levels observed in invasive carcinoma tissue. Strikingly, laser capture microdissection revealed an absence of IFN-k mRNA in diseased epithelium, whereas stromal IFN-k was found exclusively in diseased tissue. IFN-g and IFN-b were likewise found to be upregulated in diseased cervical stroma. Immunofluorescence supports the involvement of monocytes and dendritic cells in the stromal induction of IFNs in diseased tissue. Further, using three-dimensional raft cultures in which the viral life cycle can be mimicked, human keratinocytes transfected with full-length HPV16 displayed a significant decrease in IFN-k mRNA compared with non-transfected human keratinocytes. Altogether, these findings show that IFN-k is down-regulated in cervical keratinocytes harboring HPV, which may be a contributing factor in the progression of a cervical lesion. Interferons (IFNs) have a critical role in the generation of an innate immune response against invading pathogens and, as a consequence, are an approved therapy for many diseases 1,2 including human papillomavirus (HPV)-associated diseases. 3,4 On infection, IFNs are synthesized and secreted to evoke anti-viral, 5 anti-tumor, 6 and immunoregulatory (reviewed in Stetson and Medzhitov 7 ) activity conferring protection to neighboring cells. Two main classes of IFNs exist. The type I IFNs, consisting of IFN-a, -b, -k, -d, -e, -o, and -t, share a common receptor and downstream signaling pathways (reviewed in Pestka et al. 8 ) and are chiefly involved in generating anti-viral activity in response to pathogens. The sole member of the type II IFN subclass, IFN-g, uses a discrete type II IFN receptor, 9 distinct signaling pathways and is primarily involved in the regulation of immune and inflammatory responses. IFN-k, a newly identified type I IFN, has been shown to signal through the type I IFN pathway 10 and displays a tight tropism for keratinocytes, monocytes (MCs), and dendritic cells (DCs). 10 IFN-k has been shown to display similar anti-viral activity to other type I IFNs. 11 However, it is distinct from other type I IFNs, as it...

High-Risk Human Papillomaviruses Repress Constitutive Kappa Interferon Transcription via E6 To Prevent Pathogen Recognition Receptor and Antiviral-Gene Expression

“…Our analyses revealed that the constitutive transcription of IFN-is inhibited in HPV-positive keratinocytes, which is consistent with recent in situ analyses of HPV-positive biopsy specimen material and findings in HPV-positive cervical cancer cells (12,37). Since a good correlation between ISGs reduced in HPV-positive cells (Table 1) and ISGs induced by IFN-in HPV18-positive cells can be observed, we postulate that the major cause for the reduction of ISGs in HPV-positive cells is the inhibition of IFN-transcription.…”

Interferon Kappa Inhibits Human Papillomavirus 31 Transcription by Inducing Sp100 Proteins

“…Classical type I IFNs such as IFN-␣s and IFN-␤, as well as type III IFNs, are strongly induced upon virus infection, which activates pattern recognition receptor pathways. In contrast, IFN-and IFN-ε are constitutively expressed in a cell type-specific manner and are only very moderately induced by pattern recognition receptor pathways, indicating a different way of regulation (13)(14)(15)(16)48). Recent studies demonstrated that IFN-expression is inhibited by hr-HPV both in tissue culture and in vivo (13,15,16).…”

“…In contrast, IFN-and IFN-ε are constitutively expressed in a cell type-specific manner and are only very moderately induced by pattern recognition receptor pathways, indicating a different way of regulation (13)(14)(15)(16)48). Recent studies demonstrated that IFN-expression is inhibited by hr-HPV both in tissue culture and in vivo (13,15,16). It had been previously shown that IFN-␤ inhibits the growth of cells lines that maintain replicating HPV16 or -31 genomes (4,10).…”

“…The reduction of ISG expression in HPV-positive keratinocytes correlates with the inhibition of IFN-, which is an unusual member of the type I IFN family as it is constitutively expressed at high levels in normal keratinocytes and only weakly responds to inducers of IFN-␣s and -␤ (13)(14)(15)(16). The inhibition of IFN-expression is caused by the hr-HPV E6 oncoprotein and involves the induction of DNA methylation at the IFN-promoter (15,16).…”

Early Defensive Mechanisms against Human Papillomavirus Infection

“…Furthermore, knockdown of STING moderately reduced IFN-expression in primary human KC, and both proteins were reduced in HPV-positive cervical tissue and CIN1 (60). IFN-downregulation in HPV-positive precursor lesions and cervical cancer has been observed before (64,65). Interestingly, De Carlo et al (65) found that while IFN-␤ and IFN-␥ mRNAs were detectable in HPV16-positive cervical epithelium from LSIL, high-grade squamous intraepithelial lesion, and carcinoma biopsy specimens, IFN-mRNA was absent.…”