BACKGROUND: In astrocytes, the inflammatory induction of Nitric Oxide Synthase 
type 2 (NOS2) is inhibited by noradrenaline (NA) at the transcriptional level 
however its effects on specific transcription factors are not fully known. 
Recent studies show that the activity of several transcription factors including 
C/EBPbeta, which is needed for maximal NOS2 expression, is modulated by 
conjugation of the small molecular weight protein SUMO. We examined whether the 
expression of SUMO Related Genes (SRGs: SUMO-1, the conjugating enzyme Ubc9, and 
the protease SENP1) are affected by inflammatory conditions or NA and whether 
SUMO-1 regulates NOS2 through interaction with C/EBPbeta.
METHODS: Bacterial endotoxin lipopolysaccharide (LPS) was used to induce 
inflammatory responses including NOS2 expression in primary astrocytes. The mRNA 
levels of SRGs were determined by QPCR. A functional role for SUMOylation was 
evaluated by determining effects of over-expressing SRGs on NOS2 promoter and 
NFkappaB binding-element reporter constructs. Interactions of SUMO-1 and 
C/EBPbeta with the NOS2 promoter were examined by chromatin immunoprecipitation 
assays. Interactions of SUMO-1 with C/EBPbeta were examined by 
immunoprecipitation and Western blot analysis and by fluorescence resonance 
energy transfer (FRET) assays.
RESULTS: LPS decreased mRNA levels of SUMO-1, Ubc9 and SENP1 in primary 
astrocytes and a similar decrease occurred during normal aging in brain. NA 
attenuated the LPS-induced reductions and increased SUMO-1 above basal levels. 
Over-expression of SUMO-1, Ubc9, or SENP1 reduced the activation of a NOS2 
promoter, whereas activation of a 4 x NFkappaB binding-element reporter was only 
reduced by SUMO-1. ChIP studies revealed interactions of SUMO-1 and C/EBPbeta 
with C/EBP binding sites on the NOS2 promoter that were modulated by LPS and NA. 
SUMO-1 co-precipitated with C/EBPbeta and a close proximity was confirmed by 
FRET analysis.
CONCLUSION: Our results demonstrate that SUMOylation regulates NOS2 expression 
in astrocytes, and point to modification of C/EBPbeta as a possible mechanism of 
action. Targeting the SUMOylation pathway may therefore offer a novel means to 
regulate inflammatory NOS2 expression in neurological conditions and diseases.