Keratin-17 Promotes p27KIP1 Nuclear Export and Degradation and Offers Potential Prognostic Utility

Abstract

Keratins that are overexpressed selectively in human carcinomas may offer diagnostic and prognostic utility. In this study, we show that high expression of keratin-17 (K17) predicts poor outcome in patients with cervical cancer, at early or late stages of disease, surpassing in accuracy either tumor staging or loss of p27 KIP1 as a negative prognostic marker in this setting. We investigated the mechanistic basis for the biologic impact of K17 through loss-and gain-of-function experiments in human cervix, breast, and pancreatic cancer cells. Specifically, we determined that K17 functions as an oncoprotein by regulating the subcellular localization and degradation of p27 KIP1 . We found that K17 was released from intermediate filaments and translocated into the nucleus via a nuclear localization signal (NLS), specific among keratins, where it bound p27 KIP1 during G 1 phase of the cell cycle. p27 KIP1 lacks a nuclear export signal (NES) and requires an adaptor for CRM1 binding for nuclear export. In K17, we defined and validated a leucine-rich NES that mediated CRM1 binding for export. Cervical cancer cells expressing K17 mutations in its NLS or NES signals exhibited an increase in levels of nuclear p27 KIP1 , whereas cells expressing wild-type K17 exhibited a depletion in total endogenous p27 KIP1 . In clinical specimens of cervical cancer, we confirmed that the expressions of K17 and p27 KIP1 were inversely correlated, both across tumors and within individual tumors. Overall, our findings establish that K17 functions specially among keratins as an oncoprotein by controlling the ability of p27 KIP1 to influence cervical cancer pathogenesis. Cancer Res; 75(17); 3650-62. Ó2015 AACR.

Loss of Keratin 17 induces tissue-specific cytokine polarization and cellular differentiation in HPV16-driven cervical tumorigenesis in vivo

“…There were no obvious differences between genotypes with regards to the expression of the related type I keratin 14 (K14) (Supplementary Figure S1E). Consistent with findings in human cervical carcinoma (Smedts et al , 13 Escobar-Hoyos et al 15,29 ), therefore, a clear induction of K17 and expansion into the suprabasal layers coincides with lesion progression in the HPV16 tg/+ cervical epithelium. Based on these observations, we next assessed the functional roles for K17 toward the early stages of cervical tumorigenesis in estrogen-implanted HPV16 tg/+ mice.…”

“…Nuclear-localized K17 also occurs in biopsies from cases of basal cell carcinoma in human skin 22 and human cervical carcinoma, and in human cervical carcinoma-derived cell lines in culture. 29 K17 has been reported to be critical for the nuclear-to-cytoplasmic translocation of the cell cycle inhibitor p27KIP1 in human cervical epithelial cells in culture, 29 an influence that may have a direct bearing on the proliferation-differentiation balance in the cervix. Whether a nuclear-localized form of K17 and a primary impact on cell cycle regulation have a role in the setting of HPV16-induced and estrogen-promoted tumorigenesis in the mouse cervical epithelium represent open issues worth examining in the future.…”

Keratins Are Going Nuclear

“…With this in mind, how could one account for the puzzling observation, independently made by two laboratories, that the type I IF protein keratin 17 (K17) impacts the nuclear localization and function of the transcriptional regulator autoimmune regulator (AIRE) (Hobbs et al, 2015) or the cell-cycle inhibitor p27 KIP1 (CDKN1B) (Escobar-Hoyos et al, 2015)? Whereas a number of intricate mechanisms could account for this finding, a simple explanation is that K17 would itself be present and function inside the nucleus, a possibility that had not been considered until recently.…”

“…Whereas a number of intricate mechanisms could account for this finding, a simple explanation is that K17 would itself be present and function inside the nucleus, a possibility that had not been considered until recently. The two aforementioned studies (Hobbs et al, 2015; Escobar-Hoyos et al, 2015), in addition to an unbiased screen to identify nucleocytoplasmic shuttling proteins (Kumeta et al, 2013), have separately converged in demonstrating the presence of endogenous K17 and other keratin proteins inside the nucleus of tumor epithelial cells. This unexpected finding raises several questions and calls for new ideas regarding the rationale underlying the context-dependent regulation and significance of K17 and other IF proteins for cells, tissues, and organs, in both health and disease.…”

“…A key observation in the study by Hobbs et al was that manipulating K17 expression had a marked impact on the distribution of the AIRE protein inside the nucleus, i.e., punctate versus diffuse, in response to relevant stimuli. Escobar-Hoyos et al (Escobar-Hoyos et al, 2015) found K17 to occur in the nucleus of cervical tumor epithelial cells, following up on the observation of a slower proliferation rate and increased incidence of G1 cell-cycle arrest when K17 expression is silenced via RNAi. In all three studies, uncovering the presence of keratin proteins inside the nucleus of cultured tumor cells necessitated the use of a potent inhibitor of CRM1/Exportin 1-mediated nuclear export, namely Leptomycin B (LMB), so as to ‘‘trap’’ the protein of interest inside the nucleus and thus facilitate its detection.…”