Skip to main content

Main menu

  • Home
  • Articles
    • Current Issue
    • Next in The JI
    • Archive
    • Brief Reviews
    • Pillars of Immunology
    • Translating Immunology
    • Most Read
    • Top Downloads
    • Annual Meeting Abstracts
  • COVID-19/SARS/MERS Articles
  • Info
    • About the Journal
    • For Authors
    • Journal Policies
    • Influence Statement
    • For Advertisers
  • Editors
  • Submit
    • Submit a Manuscript
    • Instructions for Authors
    • Journal Policies
  • Subscribe
    • Journal Subscriptions
    • Email Alerts
    • RSS Feeds
    • ImmunoCasts
  • More
    • Most Read
    • Most Cited
    • ImmunoCasts
    • AAI Disclaimer
    • Feedback
    • Help
    • Accessibility Statement
  • Other Publications
    • American Association of Immunologists
    • ImmunoHorizons

User menu

  • Subscribe
  • Log in

Search

  • Advanced search
The Journal of Immunology
  • Other Publications
    • American Association of Immunologists
    • ImmunoHorizons
  • Subscribe
  • Log in
The Journal of Immunology

Advanced Search

  • Home
  • Articles
    • Current Issue
    • Next in The JI
    • Archive
    • Brief Reviews
    • Pillars of Immunology
    • Translating Immunology
    • Most Read
    • Top Downloads
    • Annual Meeting Abstracts
  • COVID-19/SARS/MERS Articles
  • Info
    • About the Journal
    • For Authors
    • Journal Policies
    • Influence Statement
    • For Advertisers
  • Editors
  • Submit
    • Submit a Manuscript
    • Instructions for Authors
    • Journal Policies
  • Subscribe
    • Journal Subscriptions
    • Email Alerts
    • RSS Feeds
    • ImmunoCasts
  • More
    • Most Read
    • Most Cited
    • ImmunoCasts
    • AAI Disclaimer
    • Feedback
    • Help
    • Accessibility Statement
  • Follow The Journal of Immunology on Twitter
  • Follow The Journal of Immunology on RSS

A Role for NF-κB Activation in Perforin Expression of NK Cells Upon IL-2 Receptor Signaling

Jun Zhou, Jin Zhang, Mathias G. Lichtenheld and Gary G. Meadows
J Immunol August 1, 2002, 169 (3) 1319-1325; DOI: https://doi.org/10.4049/jimmunol.169.3.1319
Jun Zhou
*Cancer Prevention and Research Center and Department of Pharmaceutical Sciences, College of Pharmacy, Washington State University, Pullman, WA 99164; and
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Jin Zhang
†Department of Microbiology and Immunology, University of Miami School of Medicine, Miami, FL 33101
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Mathias G. Lichtenheld
†Department of Microbiology and Immunology, University of Miami School of Medicine, Miami, FL 33101
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Gary G. Meadows
*Cancer Prevention and Research Center and Department of Pharmaceutical Sciences, College of Pharmacy, Washington State University, Pullman, WA 99164; and
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • Article
  • Figures & Data
  • Info & Metrics
  • PDF
Loading

Article Figures & Data

Figures

  • FIGURE 1.
    • Download figure
    • Open in new tab
    • Download powerpoint
    FIGURE 1.

    Perforin gene regulation in primary NK cells. Purified murine NK cells were stimulated with 100 U/ml IL-2 for 4 h in the presence or absence of 100 nM PDTC or 20 μM TPCK. RNA was extracted and analyzed by RT-PCR for perforin and cyclophilin A as an internal standard.

  • FIGURE 2.
    • Download figure
    • Open in new tab
    • Download powerpoint
    FIGURE 2.

    Regulation of perforin expression in NK3.3 cells stimulated with IL-2 in the presence or absence of 50 μM PDTC or 40 μM TPCK. A, The cell viability was assessed by the annexin V and PI staining after 5 h. B, RT-PCR analysis of perforin and β2m mRNA levels after 5 h. C, Nuclear run-on analysis of the perforin gene locus and β2m gene locus after 2 h of stimulation.

  • FIGURE 3.
    • Download figure
    • Open in new tab
    • Download powerpoint
    FIGURE 3.

    IL-2 stimulation of NK3.3 cells activates an NF-κB signaling pathway. A, Kinase assay analysis for IL-2-induced IKKα activity. NK3.3 cells were stimulated for the indicated times with IL-2, and IKKα immunoprecipitates were applied to in vitro kinase assays using rIκBα as substrate. The shown autorad is representative of two different experiments. B, IL-2-induced degradation of IκBα as determined by Western blot analysis of NK3.3 cells stimulated with IL-2. The result shown is representative of three independent experiments. C, EMSA of NF-κB DNA binding activity induced by IL-2 in NK3.3 cell nuclear extracts. NK3.3 cells treated with IL-2 for 4 h in the presence or absence of either 50 μM PDTC or 40 μM TPCK. The gel shift shown is representative of two different experiments. D, DNA binding specificities and identities of the IL-2-induced NF-κB-like binding activity. Cold competition EMSA and supershift assays with indicated probes and Abs were used to investigate the IL-2-induced NF-κB binding complex. The result shown is representative of three independent experiments.

  • FIGURE 4.
    • Download figure
    • Open in new tab
    • Download powerpoint
    FIGURE 4.

    NF-κB p50 binds to the upstream enhancer of the perforin gene. A, EMSA of the full-length perforin upstream enhancer and human recombinant p50 NF-κB. The binding reactions were conducted in the presence of the indicated amounts of NF-κB and the radiolabeled probe in the presence or absence of the indicated cold competitors. B, In vitro DNase I footprinting analysis of the perforin upstream enhancer for NF-κB p50 binding sites. A 32P 5′ end-labeled 117-bp DNA fragment containing the human perforin upstream enhancer was incubated with human recombinant p50, and subjected to DNase I footprinting analysis. C, Summary of the footprinting analysis of B. The sequence of upstream enhancer, three footprints of B (I, II, and III), three 2-bp mutations analyzed in Fig. 5, and the previously characterized tandem STAT element (10 ) are indicated. Probe I depicts the sequence used for EMSA in Fig. 6.

  • FIGURE 5.
    • Download figure
    • Open in new tab
    • Download powerpoint
    FIGURE 5.

    An NF-κB-like element of the perforin upstream enhancer is required for full enhancer activation by IL-2R signaling. A, Luciferase activity of the perforin upstream enhancer and three mutants (see Fig. 4) within the context of the SV40 promoter was determined upon cotransfection with pRL-CMV and standardization according to its activity in presence or absence of 500 IU/ml IL-2 in NK3.3 cells (n = 3). The first two bars represent the activity of the promotorless pGL3 vector in the absence and presence of IL-2. B, IL-2 induction (500 IU/ml) of the perforin upstream enhancer and the promotorless pGL3 vector in YT cells. The YT cells were transfected and analyzed as described for A (n = 3). The first two bars represent the activity of the promotorless pGL3 vector in the absence and presence of IL-2. C, Luciferase activity of the perforin upstream enhancer and mutant I (see Fig. 4) within the context of the SV40 promoter was determined upon cotransfection with pRL-CMV and standardization according to its activity in presence or absence of 500 IU/ml IL-2 in YT cells (n = 3).

  • FIGURE 6.
    • Download figure
    • Open in new tab
    • Download powerpoint
    FIGURE 6.

    IL-2-dependent and PDTC and TPCK sensitive NF-κB binding to the NF-κB consensus element of the perforin upstream enhancer (EMSA). Left panel, Effects of IL-2, PDTC, and TPCK on the binding activity of NK3.3 cell nuclear extracts to the perforin NF-κB element. Middle panel, Binding specificity of the IL-2-induced complex. The indicated wild-type or mutant NF-κB elements are used as probe. Right panel, The IL-2 induced perforin NF-κB consensus binding activity is specific in cold competition analyses.

PreviousNext
Back to top

In this issue

The Journal of Immunology: 169 (3)
The Journal of Immunology
Vol. 169, Issue 3
1 Aug 2002
  • Table of Contents
  • About the Cover
Print
Download PDF
Article Alerts
Sign In to Email Alerts with your Email Address
Email Article

Thank you for your interest in spreading the word about The Journal of Immunology.

NOTE: We only request your email address so that the person you are recommending the page to knows that you wanted them to see it, and that it is not junk mail. We do not capture any email address.

Enter multiple addresses on separate lines or separate them with commas.
A Role for NF-κB Activation in Perforin Expression of NK Cells Upon IL-2 Receptor Signaling
(Your Name) has forwarded a page to you from The Journal of Immunology
(Your Name) thought you would like to see this page from the The Journal of Immunology web site.
CAPTCHA
This question is for testing whether or not you are a human visitor and to prevent automated spam submissions.
Citation Tools
A Role for NF-κB Activation in Perforin Expression of NK Cells Upon IL-2 Receptor Signaling
Jun Zhou, Jin Zhang, Mathias G. Lichtenheld, Gary G. Meadows
The Journal of Immunology August 1, 2002, 169 (3) 1319-1325; DOI: 10.4049/jimmunol.169.3.1319

Citation Manager Formats

  • BibTeX
  • Bookends
  • EasyBib
  • EndNote (tagged)
  • EndNote 8 (xml)
  • Medlars
  • Mendeley
  • Papers
  • RefWorks Tagged
  • Ref Manager
  • RIS
  • Zotero
Share
A Role for NF-κB Activation in Perforin Expression of NK Cells Upon IL-2 Receptor Signaling
Jun Zhou, Jin Zhang, Mathias G. Lichtenheld, Gary G. Meadows
The Journal of Immunology August 1, 2002, 169 (3) 1319-1325; DOI: 10.4049/jimmunol.169.3.1319
del.icio.us logo Digg logo Reddit logo Twitter logo CiteULike logo Facebook logo Google logo Mendeley logo
  • Tweet Widget
  • Facebook Like

Jump to section

  • Article
    • Abstract
    • Materials and Methods
    • Results and Discussion
    • Footnotes
    • References
  • Figures & Data
  • Info & Metrics
  • PDF

Related Articles

Cited By...

More in this TOC Section

  • Loss of the Transfer RNA Wobble Uridine–Modifying Enzyme Elp3 Delays T Cell Cycle Entry and Impairs T Follicular Helper Cell Responses through Deregulation of Atf4
  • Structure and Functional Characterization of a Humanized Anti-CCL20 Antibody following Exposure to Serum Reveals the Formation of Immune Complex That Leads to Toxicity
  • The Role of the HLA Class I α2 Helix in Determining Ligand Hierarchy for the Killer Cell Ig-like Receptor 3DL1
Show more MOLECULAR AND STRUCTURAL IMMUNOLOGY

Similar Articles

Navigate

  • Home
  • Current Issue
  • Next in The JI
  • Archive
  • Brief Reviews
  • Pillars of Immunology
  • Translating Immunology

For Authors

  • Submit a Manuscript
  • Instructions for Authors
  • About the Journal
  • Journal Policies
  • Editors

General Information

  • Advertisers
  • Subscribers
  • Rights and Permissions
  • Accessibility Statement
  • Public Access
  • Privacy Policy
  • Disclaimer

Journal Services

  • Email Alerts
  • RSS Feeds
  • ImmunoCasts
  • Twitter

Copyright © 2021 by The American Association of Immunologists, Inc.

Print ISSN 0022-1767        Online ISSN 1550-6606