Unbiased discovery of interactions at a control locus driving expression of the cancer-specific therapeutic and diagnostic target, mesothelin

Yunzhao R. Ren, Raghothama Chaerkady, Shaohui Hu, Jun Wan, Jiang Qian, Heng Zhu, Akhilesh Pandey, Scott E. Kern

Research output: Contribution to journalArticle

4 Citations (Scopus)

Abstract

Although significant effort is expended on identifying transcripts/proteins that are up-regulated in cancer, there are few reports on systematic elucidation of transcriptional mechanisms underlying such druggable cancer-specific targets. The mesothelin (MSLN) gene offers a promising subject, being expressed in a restricted pattern normally, yet highly overexpressed in almost one-third of human malignancies and a target of cancer immunotherapeutic trials. CanScript, a cis promoter element, appears to control MSLN cancer-specific expression; its related genomic sequences may up-regulate other cancer markers. CanScript is a 20-nt bipartite element consisting of an SP1-like motif and a consensus MCAT sequence. The latter recruits TEAD (TEA domain) family members, which are universally expressed. Exploration of the active CanScript element, especially the proteins binding to the SP1-like motif, thus could reveal cancer-specific features having diagnostic or therapeutic interest. The efficient identification of sequence-specific DNA-binding proteins at a given locus, however, has lagged in biomarker explorations. We used two orthogonal proteomics approaches-unbiased SILAC (stable isotope labeling by amino acids in cell culture)/DNA affinity-capture/mass spectrometry survey (SD-MS) and a large transcription factor protein microarray (TFM)-and functional validation to explore systematically the CanScript interactome. SD-MS produced nine candidates, and TFM, 18. The screens agreed in confirming binding by TEAD proteins and by newly identified NAB1 and NFATc. Among other identified candidates, we found functional roles for ZNF24, NAB1 and RFX1 in MSLN expression by cancer cells. Combined interactome screens yield an efficient, reproducible, sensitive, and unbiased approach to identify sequence-specific DNA-binding proteins and other participants in disease-specific DNA elements.

Original languageEnglish (US)
Pages (from-to)5301-5310
Number of pages10
JournalJournal of Proteome Research
Volume11
Issue number11
DOIs
StatePublished - Nov 2 2012
Externally publishedYes

Fingerprint

Internal-External Control
DNA-Binding Proteins
Microarrays
Neoplasms
Proteins
Transcription Factors
Protein Array Analysis
DNA
Biomarkers
Therapeutics
Cell culture
Isotopes
Labeling
Mass spectrometry
Genes
Cells
Isotope Labeling
Amino Acids
mesothelin
Consensus Sequence

Keywords

  • mesothelin
  • NAB1
  • protein microarray
  • SILAC
  • ZNF24

ASJC Scopus subject areas

  • Biochemistry
  • Chemistry(all)

Cite this

Unbiased discovery of interactions at a control locus driving expression of the cancer-specific therapeutic and diagnostic target, mesothelin. / Ren, Yunzhao R.; Chaerkady, Raghothama; Hu, Shaohui; Wan, Jun; Qian, Jiang; Zhu, Heng; Pandey, Akhilesh; Kern, Scott E.

In: Journal of Proteome Research, Vol. 11, No. 11, 02.11.2012, p. 5301-5310.

Research output: Contribution to journalArticle

Ren, Yunzhao R. ; Chaerkady, Raghothama ; Hu, Shaohui ; Wan, Jun ; Qian, Jiang ; Zhu, Heng ; Pandey, Akhilesh ; Kern, Scott E. / Unbiased discovery of interactions at a control locus driving expression of the cancer-specific therapeutic and diagnostic target, mesothelin. In: Journal of Proteome Research. 2012 ; Vol. 11, No. 11. pp. 5301-5310.
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AU - Kern, Scott E.

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