Epigenetic changes and alternate promoter usage by human colon cancers for expressing DCLK1-isoforms: Clinical Implications

Sci Rep. 2015 Oct 8;5:14983. doi: 10.1038/srep14983.

Abstract

DCLK1 specifically marks colon/pancreatic cancers in mice, and is expressed by human colon adenocarcinomas (hCRCs). Down-regulation of DCLK1 results in loss of cancer-stem-cells (CSCs), and inhibits spheroidal/xenograft growths from hCRC-cells. The 5'-promoter of DCLK1-gene is reportedly hypermethylated in hCRCs, resulting in loss of expression of DCLK1-transcripts, originating from 5'(α)-promoter (termed DCLK1-L, in here). However, in mouse colon-tumors, 5'-promoter of DCLK1-gene remains unchanged, and DCLK1-L, originating from 5'(α)-promoter, is expressed. We hypothesized that elevated levels of DCLK1-protein in hCRC-cells, may be transcribed/translated from an alternate-promoter. Several in silico and molecular biology approaches were used to test our hypothesis. We report for the first time that majority of hCRCs express short-transcripts of DCLK1 (termed DCLK1-S, in here) from an alternate β-promoter in IntronV of the gene, while normal-colons mainly express DCLK1-L from 5'(α)-promoter. We additionally report an important role of β-catenin and TCF4/LEF binding-sites for activating (α)-promoter, while activated NF-κBp65 (bound to NF-κB-cis-element), activates (β)-promoter in cancer-cells. DCLK1-S expression was examined in a cohort of 92 CRC patients; high-expressors had significantly worse overall-survival compared to low-expressors. Our novel findings' regarding usage of alternate (β)-promoter by hCRCs, suggests that DCLK1-S may represent an important target for preventing/inhibiting colon-cancers, and for eliminating colon-CSCs.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Aged
  • Amino Acid Sequence
  • Animals
  • Cell Line
  • Cell Line, Tumor
  • Colonic Neoplasms / genetics*
  • Colonic Neoplasms / metabolism
  • Colonic Neoplasms / pathology
  • DNA Methylation
  • Epigenesis, Genetic*
  • Female
  • Gene Expression Regulation, Neoplastic
  • HCT116 Cells
  • HEK293 Cells
  • Humans
  • Immunoblotting
  • Intracellular Signaling Peptides and Proteins / genetics*
  • Intracellular Signaling Peptides and Proteins / metabolism
  • Isoenzymes / genetics
  • Isoenzymes / metabolism
  • Male
  • Mice
  • Molecular Sequence Data
  • Neoplastic Stem Cells / metabolism
  • Neoplastic Stem Cells / pathology
  • Promoter Regions, Genetic / genetics*
  • Protein-Serine-Threonine Kinases / genetics*
  • Protein-Serine-Threonine Kinases / metabolism
  • RNA Interference
  • Reverse Transcriptase Polymerase Chain Reaction
  • Sequence Homology, Amino Acid

Substances

  • Intracellular Signaling Peptides and Proteins
  • Isoenzymes
  • DCLK1 protein, human
  • Protein-Serine-Threonine Kinases