Krüppel-like factor 5 mediates the transforming activity of oncogenic H-Ras

Abstract

Previous studies indicate that Kru¨ppel-like factor 5 (KLF5), also known as intestinal-enriched Kru¨ppel-like factor (IKLF), is a positive regulator of cell proliferation and gives rise to a transformed phenotype when overexpressed. Here we demonstrate that levels of KLF5 transcript and protein are significantly elevated in oncogenic H-Ras-transformed NIH3T3 cells. These cells display an accelerated rate of proliferation in both serum-containing and serum-deprived media and form anchorage-independent colonies in soft agar assays. H-Ras-transformed cells also contain elevated mitogenactivated protein kinase (MAPK) activity. When treated with inhibitors of MEK (MAPK kinase), H-Ras-transformed cells lose their growth advantage and no longer form colonies. Significantly, levels of KLF5 transcript and protein are substantially reduced in H-Ras-transformed cells treated with MEK inhibitors. Moreover, inhibition of KLF5 expression in H-Ras-transformed cells with KLF5-specific small interfering RNA (siRNA) leads to a decreased rate of proliferation and a significant reduction in colony formation. H-Ras-transformed cells also contain elevated levels of Egr1 that are diminished by MEK inhibitors. Inhibition of Egr1 by siRNA results in a reduced level of KLF5, indicating that Egr1 mediates the inductive action of MAPK on KLF5. Lastly, KLF5 activates expression of cyclin D1. These findings indicate that the increased expression of KLF5 in H-Rastransformed cells is secondary to increased MAPK activity from H-Ras overexpression and that the elevated level of KLF5 is in part responsible for the proproliferative and transforming activities of oncogenic H-Ras.

Krüppel-like factors 4 and 5: the yin and yang regulators of cellular proliferation

“…Constitutive expression of KLF5 in NIH3T3 [26] and intestinal epithelial cells, IEC6 [27], results in an increased rate of proliferation and, eventually, a transformed phenotype as characterized by anchorageindependent growth [26] (Chanchevalap and Yang, unpublished observations). The level of KLF5 transcript is also elevated in NIH3T3 cells transformed by oncogenic H-Ras, and this increase mediates the transforming effect of oncogenic H-Ras [28]. Fig.…”

“…The increase in KLF5 expression is a direct result of Egr1, which is activated by mitogenactivated protein kinase (MAPK), which is itself stimulated by oncogenic H-Ras. The increase in KLF5 then stimulates expression of the genes encoding cyclin D1 [28], cyclin B1, and Cdc2 (Nandan and Yang, unpublished observations), a change which culminates in accelerated cell proliferation and subsequent transformation. The importance of KLF5 in mediating cell proliferation is further demonstrated by the recent finding that the inhibitory effect of all-trans retinoic acid (ATRA) on proliferation of the intestinal epithelial cells, IEC6, is mediated by suppression of KLF5 expression [27].…”

Up-Regulation of Krüppel-Like Factor 5 in Pancreatic Cancer Is Promoted by Interleukin-1β Signaling and Hypoxia-Inducible Factor-1α

“…This is of particular interest, as recent studies showed that oncogenic HRAS may induce KLF5 overexpression through a mitogenactivated protein kinase (MAPK) signaling cascade, involving extracellular signal-regulated kinase (ERK) and early growth response gene 1 (4,10). Because pancreatic cancers frequently harbor activating KRAS mutations (11), which consecutively activate MEK/MAPK/Erk signaling (12), we hypothesized that KLF5 could be up-regulated in this cancer entity.…”

Ubiquitin–proteasome degradation of KLF5 transcription factor in cancer and untransformed epithelial cells

“…These studies establish KLF5 as a strong candidate for the tumor suppressor gene located at the q21 band of chromosome 13 (13q21), whose deletion frequently occurs in many types of human tumors (Knuutila et al, 1999). In fibroblasts, however, overexpression of KLF5 stimulates cell growth, enhances transformation, and likely mediates Ras oncogene's transforming activity (Sun et al, 2001;Nandan et al, 2004). Therefore, KLF5 appears to have different functions in epithelial cells versus fibroblasts.…”

“…It plays an important role in various aspects of carcinogenesis including cell proliferation, differentiation, cell cycle regulation, and angiogenesis (Dang et al, 2000;Black et al, 2001;Sun et al, 2001;Chen et al, 2002Chen et al, , 2003Shindo et al, 2002;Bateman et al, 2004;Nandan et al, 2004). Through interactions with other proteins such as CBP/P300 (Miyamoto et al, 2003), retinoid acid receptor (Shindo et al, 2002), and NF-kB (Aizawa et al, 2004) in different cell systems, KLF5 has been shown to induce the expression of different genes including smooth muscle cell differentiation marker SM22a (Adam et al, 2000), cyclin D1 (Bateman et al, 2004;Nandan et al, 2004), platelet-derived growth factor a (PDGFa) (Aizawa et al, 2004;Usui et al, 2004), and possibly transforming growth factor b (TGF-b) (Shindo et al, 2002).…”