Complex regulation of tartrate-resistant acid phosphatase (TRAP) expression by interleukin 4 (IL-4): IL-4 indirectly suppresses receptor activator of NF-kappaB ligand (RANKL)-mediated TRAP expression but modestly induces its expression directly.
Interleukin 4 (IL-4) inhibits receptor activator of NF-kappaB ligand (RANKL)-induced osteoclast formation and functional activity in a STAT6-dependent manner. IL-4 down-regulates expression of tartrate-resistant acid phosphatase (TRAP) in mature osteoclasts. To determine whether IL-4 regulates TRAP promoter activity, RAW264.7 cells were transfected with a TRAP promoter-luciferase reporter. Treatment with IL-4 alone modestly enhanced TRAP luciferase activity. However, IL-4 suppressed the ability of RANKL to up-regulate TRAP-luciferase activity, suggesting that IL-4 has multiple effects on TRAP transcription. IL-4 also reduced the RANKL-induced association of RNA polymerase II with the TRAP gene in osteoclasts. The TRAP promoter contains a STAT6-binding motif, and STAT6 bound to the endogenous TRAP promoter after IL-4 treatment. To determine the impact of STAT6 binding, we transfected cells with STAT6VT, a constitutively active STAT6 mutant. STAT6VT alone up-regulated TRAP-luciferase activity; this effect was abrogated by mutating the STAT6 binding site in the minimal TRAP promoter. STAT6VT did not inhibit the potent up-regulation of TRAP promoter activity caused by overexpression of NFATc1, PU.1, and microphthalmia transcription factor, downstream targets of macrophage colony-stimulating factor and RANKL. IL-4 down-regulated the expression of c-Fos and NFATc1 in mature osteoclasts. Knockdown of NFATc1 by short interfering RNA caused TRAP expression to be down-regulated, and ectopic expression of NFATc1 abrogated the IL-4-induced down-regulation of TRAP. These results suggest that STAT6 plays two distinct roles in TRAP expression. The IL-4-induced activation of STAT6 mediates suppression of the RANKL-induced TRAP promoter activity indirectly by inhibiting NFATc1 expression. However, in the absence of RANKL and osteoclast differentiation, STAT6 binds the TRAP promoter after IL-4 treatment and directly enhances TRAP expression.
Yu
Minjun
M
Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, Maryland 21201, USA.
Moreno
Jose L
JL
Stains
Joseph P
JP
Keegan
Achsah D
AD
D000135:Acid Phosphatase / Q000378:metabolism*; D000818:Animals; D001483:Base Sequence; D015536:Down-Regulation; D005260:Female; D015971:Gene Expression Regulation, Enzymologic; D015847:Interleukin-4 / Q000378:metabolism*; D007527:Isoenzymes / Q000378:metabolism*; D016173:Macrophage Colony-Stimulating Factor / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D009154:Mutation; D010010:Osteoclasts / Q000166:cytology / Q000378:metabolism; D011401:Promoter Regions, Genetic; D053245:RANK Ligand / Q000378:metabolism*; D012689:Sequence Homology, Nucleic Acid; D000071681:Tartrate-Resistant Acid Phosphatase
D007527:Isoenzymes; D053245:RANK Ligand; C505688:TNFSF11 protein, human; D015847:Interleukin-4; D016173:Macrophage Colony-Stimulating Factor; D000135:Acid Phosphatase; C000605511:Acp5 protein, mouse; D000071681:Tartrate-Resistant Acid Phosphatase