Premium
Novel 5‐lipoxygenase isoforms affect the biosynthesis of 5‐lipoxygenase products
Author(s) -
Boudreau Luc H.,
Bertin Jonathan,
Robichaud Philippe P.,
Laflamme Mark,
Ouellette Rodney J.,
Flamand Nicolas,
Surette Marc E.
Publication year - 2011
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fj.10-173856
Subject(s) - gene isoform , alternative splicing , exon , arachidonate 5 lipoxygenase , lipoxygenase , biosynthesis , biology , microbiology and biotechnology , rna splicing , biochemistry , enzyme , gene , rna , arachidonic acid
5‐Lipoxygenase (5‐LO) is the essential enzyme for the biosynthesis of leukotrienes, important mediators of inflammation. This study investigated whether variants of 5‐LO exist in human leukocytes. 5‐LO mRNA isoforms that are consistent with alternative splicing were identified by RT‐PCR in a cell line or cell type‐specific pattern. All evaluated cells expressed mRNA containing all 14 exons of 5‐LO with the expected splicing sites. Individual isoforms that retained intron 10 (α‐10), lacked exon 13 (Δ‐13), and lacked exons 10 and 13 (Δ‐10,13) or that lacked the first 96 base pairs of exon 10 (Δ‐p10) were identified. Immunoreactive bands coeluting with the cloned α‐10 and Δ‐13 isoforms were measured in primary neutrophils and in Raji cells. When expressed in HEK293 cells, alternative proteins were without catalytic activity. However, when coexpressed with the active full‐length 5‐LO, alternative isoforms significantly decreased the biosynthesis of 5‐LO products by up to 44%, as assessed by reverse‐phase HPLC analysis. Additionally, in stimulated neutrophils the full‐length active 5‐LO was detected by immunoblot in both nuclear and non‐nuclear compartments, while the Δ‐13 isoform was only detected in the nuclear fraction. These alternative 5‐LO isoforms may represent a new mechanism for the regulation of the 5‐LO pathway and lipid mediator biosynthesis.—Boudreau, L. H., Bertin, J., Robichaud, P. P., Laflamme, M., Ouellette, R. J., Flamand, N., Surette, M. E. Novel 5‐lipoxygenase isoforms affect the biosynthesis of 5‐lipoxygenase products. FASEB J. 25, 1097–1105 (2011). www.fasebj.org