Human Dicer C-terminus functions as a 5-lipoxygenase binding domain.
Dicer is a multidomain ribonuclease III enzyme involved in the biogenesis of microRNAs (miRNAs) in the vast majority of eukaryotes. In human, Dicer has been shown to interact with cellular proteins via its N-terminal domain. Here, we demonstrate the ability of Dicer C-terminus to interact with 5-lipoxygenase (5LO), an enzyme involved in the biosynthesis of inflammatory mediators, in vitro and in cultured human cells. Yeast two-hybrid and GST binding assays delineated the smallest 5-lipoxygenase binding domain (5LObd) of Dicer to its C-terminal 140 amino acids comprising the double-stranded RNA (dsRNA) binding domain (dsRBD). The Dicer 5LObd-5LO association was disrupted upon Ala substitution of Trp residues 13, 75 and 102 in 5LO, suggesting that the Dicer 5LObd may recognize 5LO via its N-terminal C2-like domain. Whereas a catalytically active 5LObd-containing Dicer fragment was found to enhance 5LO enzymatic activity in vitro, human 5LO modified the miRNA precursor processing activity of Dicer. Providing a link between miRNA-mediated regulation of gene expression and inflammation, our results suggest that the formation of miRNAs may be regulated by 5LO in leukocytes and cancer cells expressing this lipoxygenase.
Dincbas-Renqvist
Vildan
V
Department of Medical Biochemistry and Biophysics, Karolinska Institutet, S-171 77, Stockholm, Sweden.
Pépin
Geneviève
G
Rakonjac
Marija
M
Plante
Isabelle
I
Ouellet
Dominique L
DL
Hermansson
Andreas
A
Goulet
Isabelle
I
Doucet
Johanne
J
Samuelsson
Bengt
B
Rådmark
Olof
O
Provost
Patrick
P
D001094:Arachidonate 5-Lipoxygenase / Q000378:metabolism*; D002478:Cells, Cultured; D053487:DEAD-box RNA Helicases / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding / Q000235:genetics; D017434:Protein Structure, Tertiary / Q000235:genetics; D020798:Two-Hybrid System Techniques
D001094:Arachidonate 5-Lipoxygenase; D053487:DEAD-box RNA Helicases