Epigenetic regulation of HYAL-1 hyaluronidase expression

Identification of HYAL-1 promoter

Vinata B. Lokeshwar, Pablo Gomez, Mario Kramer, Judith Knapp, Melissa A. McCornack, Luis E. Lopez, Nevis L. Fregien, Neetika Dhir, Steve Scherer, David J. Klumpp, Murugesan Manoharan, Mark S. Soloway, Bal L. Lokeshwar

Research output: Contribution to journalArticle

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Abstract

HYAL-1 (hyaluronoglucosaminidase-1) belongs to the hyaluronidase family of enzymes that degrade hyaluronic acid. HYAL-1 is a marker for cancer diagnosis and a molecular determinant of tumor growth, invasion, and angiogenesis. The regulation of HYAL-1 expression is unknown. Real time reverse transcription-PCR using 11 bladder and prostate cancer cells and 69 bladder tissues showed that HYAL-1 mRNA levels are elevated 10-30-fold in cells/tissues that express high hyaluronidase activity. Although multiple transcription start sites (TSS) for HYAL-1 mRNA were detected in various tissues, the major TSS in many tissues, including bladder and prostate, was at nucleotide 27274 in the cosmid clone LUCA13 (AC002455). By analyzing the 1532 base sequence 5′ to this TSS, using cloning and luciferase reporter assays, we identified a TACAAA sequence at position -31 and the minimal promoter region between nucleotides -93 and -38. Mutational analysis identified that nucleotides -73 to -50 (which include overlapping binding consensus sites for SP1, Egr-1, and AP-2), bases C -71 and C-59, and an NFκB-binding site (at position -15) are necessary for promoter activity. The chromatin immunoprecipitation assay identified that Egr-1, AP-2, and NFκB bind to the promoter in HYAL-1-expressing cells, whereas SP1 binds to the promoter in non-HYAL-1-expressing cells. 5-Aza-2′-deoxycytidine treatment, bisulfite DNA sequencing, and methylation-specific PCR revealed that HYAL-1 expression is regulated by methylation at C-71 and C-59; both Cs are part of the SP1/Egr-1-binding sites. Thus, HYAL-1 expression is epigenetically regulated by the binding of different transcription factors to the methylated and unmethylated HYAL-1 promoter.

Original languageEnglish
Pages (from-to)29215-29227
Number of pages13
JournalJournal of Biological Chemistry
Volume283
Issue number43
DOIs
StatePublished - Oct 24 2008

Fingerprint

Hyaluronoglucosaminidase
Epigenomics
Transcription Initiation Site
Tissue
Methylation
Nucleotides
Binding Sites
decitabine
Transcription Factor AP-1
Assays
Urinary Bladder
Polymerase Chain Reaction
Cosmids
Messenger RNA
Chromatin Immunoprecipitation
Cloning
Binding sites
DNA Methylation
Hyaluronic Acid
Transcription

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Cite this

Lokeshwar, V. B., Gomez, P., Kramer, M., Knapp, J., McCornack, M. A., Lopez, L. E., ... Lokeshwar, B. L. (2008). Epigenetic regulation of HYAL-1 hyaluronidase expression: Identification of HYAL-1 promoter. Journal of Biological Chemistry, 283(43), 29215-29227. https://doi.org/10.1074/jbc.M801101200

Epigenetic regulation of HYAL-1 hyaluronidase expression : Identification of HYAL-1 promoter. / Lokeshwar, Vinata B.; Gomez, Pablo; Kramer, Mario; Knapp, Judith; McCornack, Melissa A.; Lopez, Luis E.; Fregien, Nevis L.; Dhir, Neetika; Scherer, Steve; Klumpp, David J.; Manoharan, Murugesan; Soloway, Mark S.; Lokeshwar, Bal L.

In: Journal of Biological Chemistry, Vol. 283, No. 43, 24.10.2008, p. 29215-29227.

Research output: Contribution to journalArticle

Lokeshwar, VB, Gomez, P, Kramer, M, Knapp, J, McCornack, MA, Lopez, LE, Fregien, NL, Dhir, N, Scherer, S, Klumpp, DJ, Manoharan, M, Soloway, MS & Lokeshwar, BL 2008, 'Epigenetic regulation of HYAL-1 hyaluronidase expression: Identification of HYAL-1 promoter', Journal of Biological Chemistry, vol. 283, no. 43, pp. 29215-29227. https://doi.org/10.1074/jbc.M801101200
Lokeshwar VB, Gomez P, Kramer M, Knapp J, McCornack MA, Lopez LE et al. Epigenetic regulation of HYAL-1 hyaluronidase expression: Identification of HYAL-1 promoter. Journal of Biological Chemistry. 2008 Oct 24;283(43):29215-29227. https://doi.org/10.1074/jbc.M801101200
Lokeshwar, Vinata B. ; Gomez, Pablo ; Kramer, Mario ; Knapp, Judith ; McCornack, Melissa A. ; Lopez, Luis E. ; Fregien, Nevis L. ; Dhir, Neetika ; Scherer, Steve ; Klumpp, David J. ; Manoharan, Murugesan ; Soloway, Mark S. ; Lokeshwar, Bal L. / Epigenetic regulation of HYAL-1 hyaluronidase expression : Identification of HYAL-1 promoter. In: Journal of Biological Chemistry. 2008 ; Vol. 283, No. 43. pp. 29215-29227.
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abstract = "HYAL-1 (hyaluronoglucosaminidase-1) belongs to the hyaluronidase family of enzymes that degrade hyaluronic acid. HYAL-1 is a marker for cancer diagnosis and a molecular determinant of tumor growth, invasion, and angiogenesis. The regulation of HYAL-1 expression is unknown. Real time reverse transcription-PCR using 11 bladder and prostate cancer cells and 69 bladder tissues showed that HYAL-1 mRNA levels are elevated 10-30-fold in cells/tissues that express high hyaluronidase activity. Although multiple transcription start sites (TSS) for HYAL-1 mRNA were detected in various tissues, the major TSS in many tissues, including bladder and prostate, was at nucleotide 27274 in the cosmid clone LUCA13 (AC002455). By analyzing the 1532 base sequence 5′ to this TSS, using cloning and luciferase reporter assays, we identified a TACAAA sequence at position -31 and the minimal promoter region between nucleotides -93 and -38. Mutational analysis identified that nucleotides -73 to -50 (which include overlapping binding consensus sites for SP1, Egr-1, and AP-2), bases C -71 and C-59, and an NFκB-binding site (at position -15) are necessary for promoter activity. The chromatin immunoprecipitation assay identified that Egr-1, AP-2, and NFκB bind to the promoter in HYAL-1-expressing cells, whereas SP1 binds to the promoter in non-HYAL-1-expressing cells. 5-Aza-2′-deoxycytidine treatment, bisulfite DNA sequencing, and methylation-specific PCR revealed that HYAL-1 expression is regulated by methylation at C-71 and C-59; both Cs are part of the SP1/Egr-1-binding sites. Thus, HYAL-1 expression is epigenetically regulated by the binding of different transcription factors to the methylated and unmethylated HYAL-1 promoter.",
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