Germline and somatic DNA methylation and epigenetic regulation of KILLIN in renal cell carcinoma

Kristi L. Bennett, Rebecca Campbell, Shireen Ganapathi, Ming Zhou, Brian Rini, Ram Ganapathi, Hartmut P.H. Neumann, Charis Eng

Research output: Contribution to journalArticle

25 Citations (Scopus)

Abstract

We recently identified germline methylation of KILLIN, a novel p53-regulated tumor suppressor proximal to PTEN, in >1/3 Cowden or Cowden syndrome-like (CS/CSL) individuals who are PTEN mutation negative. Individuals with germline KILLIN methylation had increased risks of renal cell carcinoma (RCC) over those with PTEN mutations. Therefore, we tested the hypothesis that KILLIN may be a RCC susceptibility gene, silenced by germline methylation. We found germline hypermethylation by combined bisulfite restriction analysis in at least one of the four CpG-rich regions in 23/41 (56%) RCC patients compared to 0/50 controls (P < 0.0001). Of the 23, 11 (48%) demonstrated methylation in the -598 to -890 bp region in respect to the KILLIN transcription start site. Furthermore, 19 of 20 advanced RCC showed somatic hypermethylation upstream of KILLIN, with the majority hypermethylated at more than one CpG island (13/19 vs. 3/23 with germline methylation, P < 0.0001). qRT-PCR revealed that methylation significantly downregulates KILLIN expression (P = 0.05), and demethylation treatment by 5-aza-2'deoxycytidine significantly increased KILLIN expression in all RCC cell lines while only increasing PTEN expression in one line. Furthermore, targeted in vitro methylation revealed a significant decrease in KILLIN promoter activity only. These data reveal differential epigenetic regulation by DNA promoter methylation of this bidirectional promoter. In summary, we have identified KILLIN as a potential novel cancer predisposition gene for nonsyndromic clear-cell RCC, and the epigenetic mechanism of KILLIN inactivation in both the germline and somatic setting suggests the potential for treatment with demethylating agents.

Original languageEnglish (US)
Pages (from-to)654-661
Number of pages8
JournalGenes Chromosomes and Cancer
Volume50
Issue number8
DOIs
StatePublished - Aug 1 2011

Fingerprint

DNA Methylation
Renal Cell Carcinoma
Epigenomics
Methylation
decitabine
Mutation
CpG Islands
Neoplasm Genes
Transcription Initiation Site
Down-Regulation
Cell Line
Polymerase Chain Reaction
Therapeutics
Genes
Neoplasms

ASJC Scopus subject areas

  • Genetics
  • Cancer Research

Cite this

Bennett, K. L., Campbell, R., Ganapathi, S., Zhou, M., Rini, B., Ganapathi, R., ... Eng, C. (2011). Germline and somatic DNA methylation and epigenetic regulation of KILLIN in renal cell carcinoma. Genes Chromosomes and Cancer, 50(8), 654-661. https://doi.org/10.1002/gcc.20887

Germline and somatic DNA methylation and epigenetic regulation of KILLIN in renal cell carcinoma. / Bennett, Kristi L.; Campbell, Rebecca; Ganapathi, Shireen; Zhou, Ming; Rini, Brian; Ganapathi, Ram; Neumann, Hartmut P.H.; Eng, Charis.

In: Genes Chromosomes and Cancer, Vol. 50, No. 8, 01.08.2011, p. 654-661.

Research output: Contribution to journalArticle

Bennett, KL, Campbell, R, Ganapathi, S, Zhou, M, Rini, B, Ganapathi, R, Neumann, HPH & Eng, C 2011, 'Germline and somatic DNA methylation and epigenetic regulation of KILLIN in renal cell carcinoma', Genes Chromosomes and Cancer, vol. 50, no. 8, pp. 654-661. https://doi.org/10.1002/gcc.20887
Bennett, Kristi L. ; Campbell, Rebecca ; Ganapathi, Shireen ; Zhou, Ming ; Rini, Brian ; Ganapathi, Ram ; Neumann, Hartmut P.H. ; Eng, Charis. / Germline and somatic DNA methylation and epigenetic regulation of KILLIN in renal cell carcinoma. In: Genes Chromosomes and Cancer. 2011 ; Vol. 50, No. 8. pp. 654-661.
@article{863a6128fe1e4861b45d9dcf774a972c,
title = "Germline and somatic DNA methylation and epigenetic regulation of KILLIN in renal cell carcinoma",
abstract = "We recently identified germline methylation of KILLIN, a novel p53-regulated tumor suppressor proximal to PTEN, in >1/3 Cowden or Cowden syndrome-like (CS/CSL) individuals who are PTEN mutation negative. Individuals with germline KILLIN methylation had increased risks of renal cell carcinoma (RCC) over those with PTEN mutations. Therefore, we tested the hypothesis that KILLIN may be a RCC susceptibility gene, silenced by germline methylation. We found germline hypermethylation by combined bisulfite restriction analysis in at least one of the four CpG-rich regions in 23/41 (56{\%}) RCC patients compared to 0/50 controls (P < 0.0001). Of the 23, 11 (48{\%}) demonstrated methylation in the -598 to -890 bp region in respect to the KILLIN transcription start site. Furthermore, 19 of 20 advanced RCC showed somatic hypermethylation upstream of KILLIN, with the majority hypermethylated at more than one CpG island (13/19 vs. 3/23 with germline methylation, P < 0.0001). qRT-PCR revealed that methylation significantly downregulates KILLIN expression (P = 0.05), and demethylation treatment by 5-aza-2'deoxycytidine significantly increased KILLIN expression in all RCC cell lines while only increasing PTEN expression in one line. Furthermore, targeted in vitro methylation revealed a significant decrease in KILLIN promoter activity only. These data reveal differential epigenetic regulation by DNA promoter methylation of this bidirectional promoter. In summary, we have identified KILLIN as a potential novel cancer predisposition gene for nonsyndromic clear-cell RCC, and the epigenetic mechanism of KILLIN inactivation in both the germline and somatic setting suggests the potential for treatment with demethylating agents.",
author = "Bennett, {Kristi L.} and Rebecca Campbell and Shireen Ganapathi and Ming Zhou and Brian Rini and Ram Ganapathi and Neumann, {Hartmut P.H.} and Charis Eng",
year = "2011",
month = "8",
day = "1",
doi = "10.1002/gcc.20887",
language = "English (US)",
volume = "50",
pages = "654--661",
journal = "Genes Chromosomes and Cancer",
issn = "1045-2257",
publisher = "Wiley-Liss Inc.",
number = "8",

}

TY - JOUR

T1 - Germline and somatic DNA methylation and epigenetic regulation of KILLIN in renal cell carcinoma

AU - Bennett, Kristi L.

AU - Campbell, Rebecca

AU - Ganapathi, Shireen

AU - Zhou, Ming

AU - Rini, Brian

AU - Ganapathi, Ram

AU - Neumann, Hartmut P.H.

AU - Eng, Charis

PY - 2011/8/1

Y1 - 2011/8/1

N2 - We recently identified germline methylation of KILLIN, a novel p53-regulated tumor suppressor proximal to PTEN, in >1/3 Cowden or Cowden syndrome-like (CS/CSL) individuals who are PTEN mutation negative. Individuals with germline KILLIN methylation had increased risks of renal cell carcinoma (RCC) over those with PTEN mutations. Therefore, we tested the hypothesis that KILLIN may be a RCC susceptibility gene, silenced by germline methylation. We found germline hypermethylation by combined bisulfite restriction analysis in at least one of the four CpG-rich regions in 23/41 (56%) RCC patients compared to 0/50 controls (P < 0.0001). Of the 23, 11 (48%) demonstrated methylation in the -598 to -890 bp region in respect to the KILLIN transcription start site. Furthermore, 19 of 20 advanced RCC showed somatic hypermethylation upstream of KILLIN, with the majority hypermethylated at more than one CpG island (13/19 vs. 3/23 with germline methylation, P < 0.0001). qRT-PCR revealed that methylation significantly downregulates KILLIN expression (P = 0.05), and demethylation treatment by 5-aza-2'deoxycytidine significantly increased KILLIN expression in all RCC cell lines while only increasing PTEN expression in one line. Furthermore, targeted in vitro methylation revealed a significant decrease in KILLIN promoter activity only. These data reveal differential epigenetic regulation by DNA promoter methylation of this bidirectional promoter. In summary, we have identified KILLIN as a potential novel cancer predisposition gene for nonsyndromic clear-cell RCC, and the epigenetic mechanism of KILLIN inactivation in both the germline and somatic setting suggests the potential for treatment with demethylating agents.

AB - We recently identified germline methylation of KILLIN, a novel p53-regulated tumor suppressor proximal to PTEN, in >1/3 Cowden or Cowden syndrome-like (CS/CSL) individuals who are PTEN mutation negative. Individuals with germline KILLIN methylation had increased risks of renal cell carcinoma (RCC) over those with PTEN mutations. Therefore, we tested the hypothesis that KILLIN may be a RCC susceptibility gene, silenced by germline methylation. We found germline hypermethylation by combined bisulfite restriction analysis in at least one of the four CpG-rich regions in 23/41 (56%) RCC patients compared to 0/50 controls (P < 0.0001). Of the 23, 11 (48%) demonstrated methylation in the -598 to -890 bp region in respect to the KILLIN transcription start site. Furthermore, 19 of 20 advanced RCC showed somatic hypermethylation upstream of KILLIN, with the majority hypermethylated at more than one CpG island (13/19 vs. 3/23 with germline methylation, P < 0.0001). qRT-PCR revealed that methylation significantly downregulates KILLIN expression (P = 0.05), and demethylation treatment by 5-aza-2'deoxycytidine significantly increased KILLIN expression in all RCC cell lines while only increasing PTEN expression in one line. Furthermore, targeted in vitro methylation revealed a significant decrease in KILLIN promoter activity only. These data reveal differential epigenetic regulation by DNA promoter methylation of this bidirectional promoter. In summary, we have identified KILLIN as a potential novel cancer predisposition gene for nonsyndromic clear-cell RCC, and the epigenetic mechanism of KILLIN inactivation in both the germline and somatic setting suggests the potential for treatment with demethylating agents.

UR - http://www.scopus.com/inward/record.url?scp=79958071323&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=79958071323&partnerID=8YFLogxK

U2 - 10.1002/gcc.20887

DO - 10.1002/gcc.20887

M3 - Article

C2 - 21584899

AN - SCOPUS:79958071323

VL - 50

SP - 654

EP - 661

JO - Genes Chromosomes and Cancer

JF - Genes Chromosomes and Cancer

SN - 1045-2257

IS - 8

ER -