1: J Immunol. 2005 May 15;174(10):6238-49. 

Transcriptional regulation of the human CD3 gamma gene: the TATA-less CD3 gamma
promoter functions via an initiator and contiguous Sp-binding elements.

Badran BM, Kunstman K, Stanton J, Moschitta M, Zerghe A, Akl H, Burny A,
Wolinsky SM, Willard-Gallo KE.

Laboratory of Experimental Hematology, Bordet Institute, Faculty of Medicine,
University of Brussels, Brussels, Belgium.

Growing evidence that the CD3gamma gene is specifically targeted in some T cell
diseases focused our attention on the need to identify and characterize the
elusive elements involved in CD3gamma transcriptional control. In this study, we
show that while the human CD3gamma and CD3delta genes are oriented head-to-head
and separated by only 1.6 kb, the CD3gamma gene is transcribed from an
independent but weak, lymphoid-specific TATA-less proximal promoter. Using RNA
ligase-mediated rapid amplification of cDNA ends, we demonstrate that a cluster
of transcription initiation sites is present in the vicinity of the primary core
promoter, and the major start site is situated in a classical initiator
sequence. A GT box immediately upstream of the initiator binds Sp family
proteins and the general transcription machinery, with the activity of these
adjacent elements enhanced by the presence of a second GC box 10 nt further
upstream. The primary core promoter is limited to a sequence that extends
upstream to -15 and contains the initiator and GT box. An identical GT box
located approximately 50 nt from the initiator functions as a weak secondary
core promoter and likely generates transcripts originating upstream from the +1.
Finally, we show that two previously identified NFAT motifs in the proximal
promoter positively (NFATgamma(1)) or negatively (NFATgamma(1) and NFATgamma(2))
regulate expression of the human CD3gamma gene by their differential binding of
NFATc1 plus NF-kappaB p50 or NFATc2 containing complexes, respectively. These
data elucidate some of the mechanisms controlling expression of the CD3gamma
gene as a step toward furthering our understanding of how its transcription is
targeted in human disease.

PMID: 15879122 [PubMed - indexed for MEDLINE]


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2: J Biol Chem. 2005 May 27;280(21):20867-78. Epub 2005 Mar 2. 

c-Jun N-terminal kinase (JNK) positively regulates NFATc2 transactivation
through phosphorylation within the N-terminal regulatory domain.

Ortega-Perez I, Cano E, Were F, Villar M, Vazquez J, Redondo JM.

Centro Nacional de Investigaciones Cardiovasculares (CNIC), Ronda de Poniente 5,
Tres Cantos, Madrid 28760, Spain.

The nuclear factor of activated T cells (NFAT) family of transcription factors
regulates the transcription of cytokine genes and other genes involved in the
regulation and function of the immune system. NFAT activity is regulated by the
phosphatase calcineurin, which binds and dephosphorylates the NFAT N-terminal
regulatory domain, a critical step required for nuclear translocation and
transcriptional activity. Here we show that the mitogen-activated protein kinase
(MAPK) JNK activates NFATc2-dependent transcription. Mass spectrometry revealed
that JNK phosphorylates at least six residues within the NFATc2 regulatory
domain in vitro. Transfection of cells with a chimeric construct encoding the
GAL-4 DNA binding domain linked to wild-type NFATc2 showed that JNK stimulates
the NFATc2 transactivation domain in activated Jurkat T lymphocytes, an effect
that is inhibited by dominant-negative versions of JNK. Likewise, the mutation
of the phosphorylation sites identified revealed that Thr(116) and Ser(170) are
critical for the transactivation of NFATc2 by JNK. In addition, clustered
mutation of the SP-conserved motifs of NFATc2 showed that SP1 and SP2, but not
SP3, are also important for the inducible transactivation of NFATc2.
Furthermore, mass spectrometry analysis of NFATc2-transfected cells indicated
that the activation of the JNK pathway results in the in vivo phosphorylation of
Thr(116). Our results indicate that, unlike other NFAT members, the
transcriptional activity of NFATc2 is up-regulated by JNK. JNK-mediated
phosphorylation of NFATs thus appears to play a differential physiological role
among NFAT family members.

PMID: 15743762 [PubMed - indexed for MEDLINE]


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