title: The interplay between epigenetic silencing, oncogenic KRas and HIF-1 regulatory pathways in control of BNIP3 expression in human colorectal cancer cells.
abstract: Bcl-2/adenovirus E1B-19kDa-interacting protein 3 (BNIP3) is an important mediator of cell survival and a member of the Bcl-2 family of proteins that regulate programmed cell death and autophagy. We have previously established a link between the expression of oncogenic HRas and up-regulation of BNIP3 and the control of autophagy in cancer cells. However, in view of varied expression of BNIP3 in different tumor types and emerging uncertainties as to the role of epigenetic silencing, oncogenic regulation and the role of BNIP3 in cancer are still poorly understood. In the present study we describe profound effect of KRas on the expression of methylated BNIP3 in colorectal cancer cells and explore the interplay between HIF-1, hypoxia pathway and oncogenic KRas in this context. We observed that BNIP3 mRNA remains undetectable in aggressive DLD-1 cells harboring G13D mutant KRAS and HT-29 colorectal cancer cells unless the cells are exposed to demethylating agents such as 5-aza-2'-deoxycytidine. Following this treatment BNIP3 expression remains uniquely dependent on the Ras activity. We found that hypoxia or pharmacological activation of HIF-1 alone contributes to, but is not sufficient for efficient induction of BNIP3 mRNA transcription in cells lacking mutant KRas activity. The up-regulation of BNIP3 by KRas in this setting is mediated by the MAPK pathway, and is attenuated by the respective inhibitors (PD98059, U0126). Thus, we demonstrate the novel mechanism where activity of Ras is essential for 5-aza-2'-deoxycytidine-mediated BNIP3 expression. Moreover, we found that 5-aza-2'-deoxycytidine-mediated or enforced up-regulation of BNIP3 in DLD-1 cells results in KRas-dependent resistance to 5-Fluorouracil.
authors.affiliation: Ludwik Hirszfeld Institute of Immunology and Experimental Therapy, Polish Academy of Sciences, Wroclaw, Poland.
authors.forename: Ewelina
authors.initials: E
authors.lastname: Swiderek
authors.forename: Wojciech
authors.initials: W
authors.lastname: Kalas
authors.forename: Edyta
authors.initials: E
authors.lastname: Wysokinska
authors.forename: Alicja
authors.initials: A
authors.lastname: Pawlak
authors.forename: Janusz
authors.initials: J
authors.lastname: Rak
authors.forename: Leon
authors.initials: L
authors.lastname: Strzadala
mesh_terms: D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology; D015687:Cell Hypoxia; D015179:Colorectal Neoplasms / Q000235:genetics*; D019175:DNA Methylation; D019008:Drug Resistance, Neoplasm / Q000235:genetics; D044127:Epigenesis, Genetic; D005472:Fluorouracil / Q000494:pharmacology; D015972:Gene Expression Regulation, Neoplastic; D020868:Gene Silencing; D019073:HT29 Cells; D006801:Humans; D051793:Hypoxia-Inducible Factor 1 / Q000235:genetics / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics*; D053858:Metabolic Networks and Pathways; D011518:Proto-Oncogene Proteins / Q000235:genetics* / Q000378:metabolism*; D016283:Proto-Oncogene Proteins p21(ras); D018631:ras Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D000964:Antimetabolites, Antineoplastic; C090121:BNIP3 protein, human; D051793:Hypoxia-Inducible Factor 1; C117307:KRAS protein, human; D008565:Membrane Proteins; D011518:Proto-Oncogene Proteins; D016283:Proto-Oncogene Proteins p21(ras); D018631:ras Proteins; D005472:Fluorouracil
keywords: 5-Aza-2′-deoxycytidine; BNIP3; Colorectal cancer; HIF-1; Hypoxia; KRas