Limits...
HIF-2α mediates hypoxia-induced LIF expression in human colorectal cancer cells.

Wu L, Yu H, Zhao Y, Zhang C, Wang J, Yue X, Yang Q, Hu W - Oncotarget (2015)

Bottom Line: LIF is frequently overexpressed in a variety of human tumors and its overexpression is often associated with poor prognosis of patients.Knockdown of endogenous HIF-2α but not HIF-1α by siRNA largely abolished the induction of LIF by hypoxia in cells.Furthermore, there is a strong association of HIF-2α overexpression with LIF overexpression in human colorectal cancer specimens.

View Article: PubMed Central - PubMed

Affiliation: Rutgers Cancer Institute of New Jersey, Rutgers the State University of New Jersey, New Brunswick, NJ, USA.

ABSTRACT
Leukemia inhibitory factor (LIF), a multi-functional cytokine, has a complex role in cancer. While LIF induces the differentiation of several myeloid leukemia cells and inhibits their growth, it also promotes tumor progression, metastasis and chemoresistance in many solid tumors. LIF is frequently overexpressed in a variety of human tumors and its overexpression is often associated with poor prognosis of patients. Currently, the mechanism for LIF overexpression in tumor cells is not well-understood. Here, we report that hypoxia, a hallmark of solid tumors, induced LIF mRNA expression in human colorectal cancer cells. Analysis of LIF promoter revealed several hypoxia-responsive elements (HREs) that can specifically interact with and be transactivated by HIF-2α but not HIF-1α. Consistently, ectopic expression of HIF-2α but not HIF-1α transcriptionally induced LIF expression levels in cells. Knockdown of endogenous HIF-2α but not HIF-1α by siRNA largely abolished the induction of LIF by hypoxia in cells. Furthermore, there is a strong association of HIF-2α overexpression with LIF overexpression in human colorectal cancer specimens. In summary, results from this study demonstrate that hypoxia induces LIF expression in human cancer cells mainly through HIF-2α, which could be an important underlying mechanism for LIF overexpression in human cancers.

Show MeSH

Related in: MedlinePlus

Hypoxia transactivates hypoxia-responsive elements (HREs) in the LIF promoter through HIF-2α(A) The human LIF gene contains 4 putative HREs in its promoter region. (B) Hypoxia activates the luciferase activity of reporter vectors containing HRE-C or HRE-D sites in the LIF promoter. RKO and HCT116 cells were transfected with the luciferase reporter vectors, and then subjected to hypoxia treatment for 36 h before measuring luciferase activities. Luciferase reporter vectors containing the HRE site in the VEGF promoter was included as a positive control. (C) and (D) HIF-2α but not HIF-1α binds to HRE-C and HRE-D sites in the LIF promoter under the hypoxic condition in RKO cells as determined by ChIP assays. Cells were cultured under the hypoxic or normoxic conditions for 36 h before assays. The HRE site in the VEGF promoter serves as a positive control. The amount of DNA fragments pulled-down was determined by real-time PCR (C) or conventional PCR (D). Data are presented as mean ± SD (n = 3). *:p < 0.01 (Student's t-test).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4414199&req=5

Figure 2: Hypoxia transactivates hypoxia-responsive elements (HREs) in the LIF promoter through HIF-2α(A) The human LIF gene contains 4 putative HREs in its promoter region. (B) Hypoxia activates the luciferase activity of reporter vectors containing HRE-C or HRE-D sites in the LIF promoter. RKO and HCT116 cells were transfected with the luciferase reporter vectors, and then subjected to hypoxia treatment for 36 h before measuring luciferase activities. Luciferase reporter vectors containing the HRE site in the VEGF promoter was included as a positive control. (C) and (D) HIF-2α but not HIF-1α binds to HRE-C and HRE-D sites in the LIF promoter under the hypoxic condition in RKO cells as determined by ChIP assays. Cells were cultured under the hypoxic or normoxic conditions for 36 h before assays. The HRE site in the VEGF promoter serves as a positive control. The amount of DNA fragments pulled-down was determined by real-time PCR (C) or conventional PCR (D). Data are presented as mean ± SD (n = 3). *:p < 0.01 (Student's t-test).

Mentions: The transcriptional response to hypoxia in cells is largely mediated by HIFs. HIFs are heterodimeric transcription factors that are composed of an α-subunit and a β-subunit of helix-loop-helix-PAS family proteins. HIFs bind to DNA containing a hypoxia-responsive element (HRE; 5′-G/ACGTG-3′) [23]. Under the hypoxic condition, HIFs are stabilized and the transcriptional activities of HIFs increase. HIF-1α and HIF-2α are two major α-subunits of HIFs. HIF-1α and HIF-2α induce a number of common genes, and at the same time they each have their unique target genes [23]. To investigate whether the transcriptional induction of LIF by hypoxia is mediated by HIFs, we searched for the HRE consensus sequence in the promoter region of the LIF gene from 3 kb upstream of transcriptional site to exon 1. Four putative HRE sites (HRE-A, HRE-B, HRE-C and HRE-D) were identified in the promoter region of the LIF gene (Figure 2A).


HIF-2α mediates hypoxia-induced LIF expression in human colorectal cancer cells.

Wu L, Yu H, Zhao Y, Zhang C, Wang J, Yue X, Yang Q, Hu W - Oncotarget (2015)

Hypoxia transactivates hypoxia-responsive elements (HREs) in the LIF promoter through HIF-2α(A) The human LIF gene contains 4 putative HREs in its promoter region. (B) Hypoxia activates the luciferase activity of reporter vectors containing HRE-C or HRE-D sites in the LIF promoter. RKO and HCT116 cells were transfected with the luciferase reporter vectors, and then subjected to hypoxia treatment for 36 h before measuring luciferase activities. Luciferase reporter vectors containing the HRE site in the VEGF promoter was included as a positive control. (C) and (D) HIF-2α but not HIF-1α binds to HRE-C and HRE-D sites in the LIF promoter under the hypoxic condition in RKO cells as determined by ChIP assays. Cells were cultured under the hypoxic or normoxic conditions for 36 h before assays. The HRE site in the VEGF promoter serves as a positive control. The amount of DNA fragments pulled-down was determined by real-time PCR (C) or conventional PCR (D). Data are presented as mean ± SD (n = 3). *:p < 0.01 (Student's t-test).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4414199&req=5

Figure 2: Hypoxia transactivates hypoxia-responsive elements (HREs) in the LIF promoter through HIF-2α(A) The human LIF gene contains 4 putative HREs in its promoter region. (B) Hypoxia activates the luciferase activity of reporter vectors containing HRE-C or HRE-D sites in the LIF promoter. RKO and HCT116 cells were transfected with the luciferase reporter vectors, and then subjected to hypoxia treatment for 36 h before measuring luciferase activities. Luciferase reporter vectors containing the HRE site in the VEGF promoter was included as a positive control. (C) and (D) HIF-2α but not HIF-1α binds to HRE-C and HRE-D sites in the LIF promoter under the hypoxic condition in RKO cells as determined by ChIP assays. Cells were cultured under the hypoxic or normoxic conditions for 36 h before assays. The HRE site in the VEGF promoter serves as a positive control. The amount of DNA fragments pulled-down was determined by real-time PCR (C) or conventional PCR (D). Data are presented as mean ± SD (n = 3). *:p < 0.01 (Student's t-test).
Mentions: The transcriptional response to hypoxia in cells is largely mediated by HIFs. HIFs are heterodimeric transcription factors that are composed of an α-subunit and a β-subunit of helix-loop-helix-PAS family proteins. HIFs bind to DNA containing a hypoxia-responsive element (HRE; 5′-G/ACGTG-3′) [23]. Under the hypoxic condition, HIFs are stabilized and the transcriptional activities of HIFs increase. HIF-1α and HIF-2α are two major α-subunits of HIFs. HIF-1α and HIF-2α induce a number of common genes, and at the same time they each have their unique target genes [23]. To investigate whether the transcriptional induction of LIF by hypoxia is mediated by HIFs, we searched for the HRE consensus sequence in the promoter region of the LIF gene from 3 kb upstream of transcriptional site to exon 1. Four putative HRE sites (HRE-A, HRE-B, HRE-C and HRE-D) were identified in the promoter region of the LIF gene (Figure 2A).

Bottom Line: LIF is frequently overexpressed in a variety of human tumors and its overexpression is often associated with poor prognosis of patients.Knockdown of endogenous HIF-2α but not HIF-1α by siRNA largely abolished the induction of LIF by hypoxia in cells.Furthermore, there is a strong association of HIF-2α overexpression with LIF overexpression in human colorectal cancer specimens.

View Article: PubMed Central - PubMed

Affiliation: Rutgers Cancer Institute of New Jersey, Rutgers the State University of New Jersey, New Brunswick, NJ, USA.

ABSTRACT
Leukemia inhibitory factor (LIF), a multi-functional cytokine, has a complex role in cancer. While LIF induces the differentiation of several myeloid leukemia cells and inhibits their growth, it also promotes tumor progression, metastasis and chemoresistance in many solid tumors. LIF is frequently overexpressed in a variety of human tumors and its overexpression is often associated with poor prognosis of patients. Currently, the mechanism for LIF overexpression in tumor cells is not well-understood. Here, we report that hypoxia, a hallmark of solid tumors, induced LIF mRNA expression in human colorectal cancer cells. Analysis of LIF promoter revealed several hypoxia-responsive elements (HREs) that can specifically interact with and be transactivated by HIF-2α but not HIF-1α. Consistently, ectopic expression of HIF-2α but not HIF-1α transcriptionally induced LIF expression levels in cells. Knockdown of endogenous HIF-2α but not HIF-1α by siRNA largely abolished the induction of LIF by hypoxia in cells. Furthermore, there is a strong association of HIF-2α overexpression with LIF overexpression in human colorectal cancer specimens. In summary, results from this study demonstrate that hypoxia induces LIF expression in human cancer cells mainly through HIF-2α, which could be an important underlying mechanism for LIF overexpression in human cancers.

Show MeSH
Related in: MedlinePlus