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Production in Human NK Cells
PRDM1/Blimp-1 Controls Effector Cytokine
Djeu, Esteban Celis, Michael A. Caligiuri and Kenneth L.
Becknell, Aharon G. Freud, Jianhua Yu, Sheng Wei, Julie
Matthew A. Smith, Michelle Maurin, Hyun Il Cho, Brian
2010; 185:6058-6067; Prepublished online 13
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The Journal of Immunology
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The Journal of Immunology
PRDM1/Blimp-1 Controls Effector Cytokine Production in
Human NK Cells
Matthew A. Smith,*,†Michelle Maurin,* Hyun Il Cho,* Brian Becknell,‡Aharon G. Freud,‡
Jianhua Yu,‡Sheng Wei,* Julie Djeu,* Esteban Celis,* Michael A. Caligiuri,‡
and Kenneth L. Wright*,†
NK cells are major effectors of the innate immune response through cytolysis and bridge to the adaptive immune response
through cytokine release. The mediators of activation are well studied; however, little is known about the mechanisms that re-
strain activation. In this report, we demonstrate that the transcriptional repressor PRDM1 (also known as Blimp-1 or PRDI-BF1)
is a critical negative regulator of NK function. Three distinct PRDM1 isoforms are selectively induced in the CD56dimNK
population in response to activation. PRDM1 coordinately suppresses the release of IFN-g, TNF-a, and TNF-b through direct
binding to multiple conserved regulatory regions. Ablation of PRDM1 expression leads to enhanced production of IFN-g and
TNF-a but does not alter cytotoxicity, whereas overexpression blocks cytokine production. PRDM1 response elements are defined
at the IFNG and TNF loci. Collectively, these data demonstrate a key role for PRDM1 in the negative regulation of NK activation
and position PRDM1 as a common regulator of the adaptive and innate immune response.
in a non-MHC–restricted manner, subsequent studies highlighted
their role in cytokine production. In response to activating stimuli,
NK cells proliferate, increase cytotoxicity, and produce cytokines,
such as IFN-g, TNF-a, and GM-CSF (1). IL-2 upregulates the ex-
pression of effector molecules and enhances natural cytotoxicity
against a variety of targets. Furthermore, IL-2 and IL-15 signal
through the common gcR to control proliferation, with IL-15 be-
ing uniquely required for survival in vivo (2). IL-12 and IL-18
signal through distinct heterodimeric receptor complexes to elicit
increases in IFN-g via several mechanisms, including increased
transcription, message stability, and nuclear retention (3–5). Syn-
ergistic increases in cytotoxicity and IFN-g production are ob-
served in response to costimulation with IL-12 and IL-18 (6, 7).
Cytokine-mediated activation of NK cells proceeds through
several well-characterized nuclear transcription factors, many of
which are functionally conserved between T and NK lineages (8).
STAT4 is induced in response to IL-12 and is required for optimal
The Journal of Immunology, 2010,
atural killer cells play critical functions in innate and
adaptive immunity. Although these lymphocytes were
initially identified by their ability to lyse leukemia cells
IFN-g production and increased cytotoxicity (9). IL-18 induces nu-
clear localization of NF-kB p50/p65 which, cooperatively with
AP-1, increases IFN-g and cytotoxicity (10). Furthermore, NFAT
induces transcription of GM-CSF and TNF-a in NK cells (11).
Conversely, relatively few negative regulators of activation-induced
transcription have been identified in NK cells. ATF3 was recently
shown to downregulate IFN-g levels, and ATF2/2mice exhibit in-
creased resistance to murine CMV infection (12). The transcription
factor H2.0-like homeobox negatively regulates IFN-g production,
primarily through degradation of phosphorylated STAT4 not
direct DNA-binding activity (13).
PRDM1 (also known as Blimp-1 or PRDI-BF1) is a transcrip-
tional repressor encoded by the PRDM1 gene on chromosome
6q21. It was originally identified as a postinduction suppressor of
IFNB in virally infected osteosarcoma cells (14). Subsequent
work revealed a pivotal role in the terminal differentiation of Ab-
producing plasma cells (15). We and other investigators previously
showed that PRDM1 exerts its repressive functions through re-
cruitment of histone-modifying enzymes (HDAC2, G9a, PRMT5,
and LSD1) and Groucho corepressors (16–18). Through silencing
of direct (cMyc, CIITA, Pax5) and indirect targets, PRDM1 is a
master regulator of terminal differentiation of B lymphocytes, me-
diating cell cycle exit, repression of early B cell factors, and in-
duction of Ig secretion (19, 20).
More recently, a role for PRDM1 in T lymphocytes has emerged.
PRDM1 is expressed in CD4 and CD8 T cell lineages and is critical
for maintenance of homeostasis. Conditional knockout in T lym-
phocytes leads to increased effector populations, resulting in severe
colitis (21, 22). Upon activation, an autoregulatory loop exists,
whereby IL-2 induces PRDM1 expression, which, in turn, nega-
tively regulates IL-2 transcription (23, 24). During CD4 polariza-
tion, PRDM1 is preferentially expressed in Th2 cells and reinforces
commitment to this lineage through repression of Ifng, cfos, and
tbx21 (24, 25). Within the CD8 lineage, PRDM1 is expressed at
higher levels in exhausted subsets and promotes acquisition of
the effector phenotype through suppression of memory potential
(26–28). Thus, in addition to well-characterized B cell-specific
*Immunology Program, H. Lee Moffitt Cancer Center and Research Institute;
†Department of Molecular Medicine, University of South Florida, Tampa, FL
33612; and‡Ohio State Comprehensive Cancer Center, Columbus, OH 43210
Received for publication May 20, 2010. Accepted for publication September 11,
This work was supported by the James and Ester King Biomedical Research Program
The sequences presented in this article have been submitted to the Gene Expression
Omnibus under accession number GE22919.
Address correspondence and reprint requests to Dr. Kenneth L. Wright, H. Lee
Moffitt Cancer Center, MRC4E, 12902 Magnolia Drive, Tampa, FL 33612. E-mail
The online version of this article contains supplemental material.
Abbreviations used in this paper: ChIP, chromatin immunoprecipitation; Ct, threshold
cycle; KD, knockdown; NT, nontargeting; PARP, poly(ADP-ribose) polymerase;
poly-IC, polyinosinic-polycytidylic acid; siRNA, small interfering RNA.
by guest on December 28, 2015
interleukin-15 in the regulation of human natural killer cell survival. J. Clin.
Invest. 99: 937–943.
3. Robertson, M. J., R. J. Soiffer, S. F. Wolf, T. J. Manley, C. Donahue, D. Young,
S. H. Herrmann, and J. Ritz. 1992. Response of human natural killer (NK) cells
to NK cell stimulatory factor (NKSF): cytolytic activity and proliferation of
NK cells are differentially regulated by NKSF. J. Exp. Med. 175: 779–788.
4. Chan, S. H., M. Kobayashi, D. Santoli, B. Perussia, and G. Trinchieri. 1992.
Mechanisms of IFN-gamma induction by natural killer cell stimulatory factor
(NKSF/IL-12). Role of transcription and mRNA stability in the synergistic in-
teraction between NKSF and IL-2. J. Immunol. 148: 92–98.
5. Mavropoulos, A., G. Sully, A. P. Cope, and A. R. Clark. 2005. Stabilization of
IFN-gammamRNA byMAPK p38
human NK cells. Blood 105: 282–288.
6. Lapaque, N., T. Walzer, S. Me ´resse, E. Vivier, and J. Trowsdale. 2009. Inter-
actions between human NK cells and macrophages in response to Salmonella
infection. J. Immunol. 182: 4339–4348.
7. Ortaldo, J. R., R. Winkler-Pickett, J. Wigginton, M. Horner, E. W. Bere,
A. T. Mason, N. Bhat, J. Cherry, M. Sanford, D. L. Hodge, and H. A. Young.
2006. Regulation of ITAM-positive receptors: role of IL-12 and IL-18. Blood
8. Glimcher, L. H., M. J. Townsend, B. M. Sullivan, and G. M. Lord. 2004. Recent
developments in the transcriptional regulation of cytolytic effector cells. Nat.
Rev. Immunol. 4: 900–911.
9. Thierfelder, W. E., J. M. van Deursen, K. Yamamoto, R. A. Tripp, S. R. Sarawar,
R. T. Carson, M. Y. Sangster, D. A. Vignali, P. C. Doherty, G. C. Grosveld, and
J. N. Ihle. 1996. Requirement for Stat4 in interleukin-12-mediated responses of
natural killer and T cells. Nature 382: 171–174.
10. Robinson, D., K. Shibuya, A. Mui, F. Zonin, E. Murphy, T. Sana, S. B. Hartley,
S. Menon, R. Kastelein, F. Bazan, and A. O’Garra. 1997. IGIF does not drive
Th1 development but synergizes with IL-12 for interferon-gamma production
and activates IRAK and NFkappaB. Immunity 7: 571–581.
11. Aramburu, J., L. Azzoni, A. Rao, and B. Perussia. 1995. Activation and ex-
pression of the nuclear factors of activated T cells, NFATp and NFATc, in human
natural killer cells: regulation upon CD16 ligand binding. J. Exp. Med. 182: 801–
12. Rosenberger, C. M., A. E. Clark, P. M. Treuting, C. D. Johnson, and A. Aderem.
2008. ATF3 regulates MCMV infection in mice by modulating IFN-gamma
expression in natural killer cells. Proc. Natl. Acad. Sci. USA 105: 2544–2549.
13. Becknell, B., T. L. Hughes, A. G. Freud, B. W. Blaser, J. Yu, R. Trotta,
H. C. Mao, M. L. Caligiuri de Jesu ´s, M. Alghothani, D. M. Benson, Jr., et al.
2007. Hlx homeobox transcription factor negatively regulates interferon-gamma
production in monokine-activated natural killer cells. Blood 109: 2481–2487.
14. Keller, A. D., and T. Maniatis. 1991. Identification and characterization of
a novel repressor of beta-interferon gene expression. Genes Dev. 5: 868–879.
15. Turner, C. A., Jr., D. H. Mack, and M. M. Davis. 1994. Blimp-1, a novel zinc
finger-containing protein that can drive the maturation of B lymphocytes into
immunoglobulin-secreting cells. Cell 77: 297–306.
16. Gyory, I., J. Wu, G. Feje ´r, E. Seto, and K. L. Wright. 2004. PRDI-BF1 recruits
the histone H3 methyltransferase G9a in transcriptional silencing. Nat. Immunol.
17. Martins, G., and K. Calame. 2008. Regulation and functions of Blimp-1 in Tand
B lymphocytes. Annu. Rev. Immunol. 26: 133–169.
18. Su, S. T., H. Y. Ying, Y. K. Chiu, F. R. Lin, M. Y. Chen, and K. I. Lin. 2009.
Involvement of histone demethylase LSD1 in Blimp-1-mediated gene repression
during plasma cell differentiation. Mol. Cell. Biol. 29: 1421–1431.
19. Shaffer, A. L., K. I. Lin, T. C. Kuo, X. Yu, E. M. Hurt, A. Rosenwald,
J. M. Giltnane, L. Yang, H. Zhao, K. Calame, and L. M. Staudt. 2002. Blimp-1
orchestrates plasma cell differentiation by extinguishing the mature B cell gene
expression program. Immunity 17: 51–62.
20. Sciammas, R., and M. M. Davis. 2004. Modular nature of Blimp-1 in the reg-
ulation of gene expression during B cell maturation. J. Immunol. 172: 5427–
21. Kallies, A., E. D. Hawkins, G. T. Belz, D. Metcalf, M. Hommel, L. M. Corcoran,
P. D. Hodgkin, and S. L. Nutt. 2006. Transcriptional repressor Blimp-1 is es-
sential for T cell homeostasis and self-tolerance. Nat. Immunol. 7: 466–474.
22. Martins, G. A., L. Cimmino, M. Shapiro-Shelef, M. Szabolcs, A. Herron,
E. Magnusdottir, and K. Calame. 2006. Transcriptional repressor Blimp-1 reg-
ulates T cell homeostasis and function. Nat. Immunol. 7: 457–465.
23. Gong, D., and T. R. Malek. 2007. Cytokine-dependent Blimp-1 expression in
activated T cells inhibits IL-2 production. J. Immunol. 178: 242–252.
24. Martins, G. A., L. Cimmino, J. Liao, E. Magnusdottir, and K. Calame. 2008.
Blimp-1 directly represses Il2 and the Il2 activator Fos, attenuating T cell pro-
liferation and survival. J. Exp. Med. 205: 1959–1965.
25. Cimmino, L., G. A. Martins, J. Liao, E. Magnusdottir, G. Grunig, R. K. Perez,
and K. L. Calame. 2008. Blimp-1 attenuates Th1 differentiation by repression of
ifng, tbx21, and bcl6 gene expression. J. Immunol. 181: 2338–2347.
26. Rutishauser, R. L., G. A. Martins, S. Kalachikov, A. Chandele, I. A. Parish,
E. Meffre, J. Jacob, K. Calame, and S. M. Kaech. 2009. Transcriptional repressor
Blimp-1 promotes CD8(+) T cell terminal differentiation and represses the ac-
quisition of central memory T cell properties. Immunity 31: 296–308.
27. Kallies, A., A. Xin, G. T. Belz, and S. L. Nutt. 2009. Blimp-1 transcription factor
is required for the differentiation of effector CD8(+) T cells and memory
responses. Immunity 31: 283–295.
28. Shin, H., S. D. Blackburn, A. M. Intlekofer, C. Kao, J. M. Angelosanto,
S. L. Reiner, and E. J. Wherry. 2009. A role for the transcriptional repressor
in IL-12-and IL-18-stimulated
Blimp-1 in CD8(+) T cell exhaustion during chronic viral infection. Immunity
29. Van Gelder, R. N., M. E. von Zastrow, A. Yool, W. C. Dement, J. D. Barchas, and
J. H. Eberwine. 1990. Amplified RNA synthesized from limited quantities of
heterogeneous cDNA. Proc. Natl. Acad. Sci. USA 87: 1663–1667.
30. Desai, S., S. C. Bolick, M. Maurin, and K. L. Wright. 2009. PU.1 regulates
positive regulatory domain I-binding factor 1/Blimp-1 transcription in
lymphoma cells. J. Immunol. 183: 5778–5787.
31. Nilsson, M., J. Ljungberg, J. Richter, T. Kiefer, M. Magnusson, A. Lieber,
B. Widegren, S. Karlsson, and X. Fan. 2004. Development of an adenoviral
vector system with adenovirus serotype 35 tropism; efficient transient gene
transfer into primary malignant hematopoietic cells. J. Gene Med. 6: 631–641.
32. Gyo ¨ry, I., G. Feje ´r, N. Ghosh, E. Seto, and K. L. Wright. 2003. Identification of
a functionally impaired positive regulatory domain I binding factor 1 tran-
scription repressor in myeloma cell lines. J. Immunol. 170: 3125–3133.
33. Schmidt, D., A. Nayak, J. E. Schumann, A. Schimpl, I. Berberich, and
F. Berberich-Siebelt. 2008. Blimp-1Deltaexon7: a naturally occurring Blimp-1
deletion mutant with auto-regulatory potential. Exp. Cell Res. 314: 3614–3627.
34. Chehimi, J., S. E. Starr, I. Frank, M. Rengaraju, S. J. Jackson, C. Llanes,
M. Kobayashi, B. Perussia, D. Young, E. Nickbarg, et al. 1992. Natural killer
(NK) cell stimulatory factor increases the cytotoxic activity of NK cells from
both healthy donors and human immunodeficiency virus-infected patients. J.
Exp. Med. 175: 789–796.
35. Huntington, N. D., C. A. J. Vosshenrich, and J. P. Di Santo. 2007. Developmental
pathways that generate natural-killer-cell diversity in mice and humans. Nat.
Rev. Immunol. 7: 703–714.
36. Schoenborn, J. R., M. O. Dorschner, M. Sekimata, D. M. Santer, M. Shnyreva,
D. R. Fitzpatrick, J. A. Stamatoyannopoulos, and C. B. Wilson. 2007. Com-
prehensive epigenetic profiling identifies multiple distal regulatory elements
directing transcription of the gene encoding interferon-gamma. [Published errata
appear in 2008 Nat. Immunol. 9: 105 and 2007 Nat Immunol. 8: 893.] Nat.
Immunol. 8: 732–742.
37. Taylor, J. M., K. Wicks, C. Vandiedonck, and J. C. Knight. 2008. Chromatin
profiling across the human tumour necrosis factor gene locus reveals a com-
plex, cell type-specific landscape with novel regulatory elements. Nucleic Acids
Res. 36: 4845–4862.
2007. Activation-dependent intrachromosomal interactions formed by the TNF gene
promoter and two distal enhancers. Proc. Natl. Acad. Sci. USA 104: 16850–16855.
39. Dybkaer, K., J. Iqbal, G. Zhou, H. Geng, L. Xiao, A. Schmitz, F. d’Amore, and
W. C. Chan. 2007. Genome wide transcriptional analysis of resting and IL2
activated human natural killer cells: gene expression signatures indicative of
novel molecular signaling pathways. BMC Genomics 8: 230–246.
40. Hanna, J., P. Bechtel, Y. Zhai, F. Youssef, K. McLachlan, and O. Mandelboim.
2004. Novel insights on human NK cells’ immunological modalities revealed by
gene expression profiling. J. Immunol. 173: 6547–6563.
41. Koopman, L. A., H. D. Kopcow, B. Rybalov, J. E. Boyson, J. S. Orange,
F. Schatz, R. Masch, C. J. Lockwood, A. D. Schachter, P. J. Park, and
J. L. Strominger. 2003. Human decidual natural killer cells are a unique NK cell
subset with immunomodulatory potential. J. Exp. Med. 198: 1201–1212.
42. Wendt, K., E. Wilk, S. Buyny, J. Buer, R. E. Schmidt, and R. Jacobs. 2006. Gene
and protein characteristics reflect functional diversity of CD56dim and
CD56bright NK cells. J. Leukoc. Biol. 80: 1529–1541.
43. Chang, S., and T. M. Aune. 2005. Histone hyperacetylated domains across the
Ifng gene region in natural killer cells and T cells. Proc. Natl. Acad. Sci. USA
44. Fehniger, T. A., M. A. Cooper, G. J. Nuovo, M. Cella, F. Facchetti, M. Colonna,
and M. A. Caligiuri. 2003. CD56bright natural killer cells are present in human
lymph nodes and are activated by T cell-derived IL-2: a potential new link be-
tween adaptive and innate immunity. Blood 101: 3052–3057.
45. Chan, A., D. L. Hong, A. Atzberger, S. Kollnberger, A. D. Filer, C. D. Buckley,
A. McMichael, T. Enver, and P. Bowness. 2007. CD56bright human NK cells
differentiate into CD56dim cells: role of contact with peripheral fibroblasts. J.
Immunol. 179: 89–94.
46. Yu, J., H. C. Mao, M. Wei, T. Hughes, J. Zhang, I. K. Park, S. Liu, S. McClory,
G. Marcucci, R. Trotta, and M. A. Caligiuri. 2010. CD94 surface density
identifies a functional intermediary between the CD56bright and CD56dim hu-
man NK-cell subsets. Blood 115: 274–281.
47. Zhao, W. L., Y. Y. Liu, Q. L. Zhang, L. Wang, C. Leboeuf, Y. W. Zhang, J. Ma,
J. F. Garcia, Y. P. Song, J. M. Li, et al. 2008. PRDM1 is involved in chemo-
resistance of T-cell lymphoma and down-regulated by the proteasome inhibitor.
Blood 111: 3867–3871.
48. Keller, A. D., and T. Maniatis. 1992. Only two of the five zinc fingers of the
eukaryotic transcriptional repressor PRDI-BF1 are required for sequence-
specific DNA binding. Mol. Cell. Biol. 12: 1940–1949.
49. Iqbal, J., C. Kucuk, R. J. Deleeuw, G. Srivastava, W. Tam, H. Geng,
D. Klinkebiel, J. K. Christman, K. Patel, K. Cao, et al. 2009. Genomic analyses
reveal global functional alterations that promote tumor growth and novel tumor
suppressor genes in natural killer-cell malignancies. Leukemia 23: 1139–1151.
50. Huang, Y., A. de Reynie `s, L. de Leval, B. Ghazi, N. Martin-Garcia, M. Travert,
J. Bosq, J. Brie `re, B. Petit, E. Thomas, et al. 2010. Gene expression profiling
identifies emerging oncogenic pathways operating in extranodal NK/T-cell
lymphoma, nasal type. Blood 115: 1226–1237.
51. Sun, J. C., J. N. Beilke, and L. L. Lanier. 2009. Adaptive immune features of
natural killer cells. Nature 457: 557–561.
The Journal of Immunology6067
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