Serpin 2a is induced in activated macrophages and conjugates to a ubiquitin homolog.
After i.p. infection of mice with the intracellular bacterium Mycobacterium bovis bacillus Calmette-Guérin, macrophages recovered from the peritoneal cavity display classical signs of immune activation. We have identified a member of the serine protease inhibitor (serpin) family which is highly induced in macrophages during bacillus Calmette-Guérin infection. Serpin 2a (spi2a) expression is also induced in macrophages in vivo during infection with Salmonella typhimurium and Listeria monocytogenes, and in vitro by a variety of bacteria and bacterial products. The cytokine IFN-gamma also induces spi2a expression in macrophages, and this induction is synergistic with bacterial products. We also demonstrate here that a ubiquitin homolog, IFN-stimulated gene of 15-kDa (ISG15), is strongly induced during in vitro and in vivo activation of macrophages and that it conjugates to spi2a in activated macrophages. The ISG15-spi2a conjugates were identified by tandem mass spectrometry and contained spi2a conjugated to either one or two molecules of ISG15. Whereas spi2a was induced by either bacterial products or IFN-gamma, ISG15 was induced only by bacterial products. Although many protein targets have been described for ubiquitin conjugation, spi2a is the first ISG15-modified protein to be reported. Macrophage activation is accompanied by the activation of a variety of proteases. It is of interest that a member of the serine protease inhibitor family is concomitantly induced and modified by a ubiquitin-like protein.
Hamerman
Jessica A
JA
Department of Immunology, University of Washington, Seattle, WA 98185, USA. jah@itsa.ucsf.edu
Hayashi
Fumitaka
F
Schroeder
Lea A
LA
Gygi
Steven P
SP
Haas
Arthur L
AL
Hampson
Lynne
L
Coughlin
Paul
P
Aebersold
Ruedi
R
Aderem
Alan
A
D000595:Amino Acid Sequence; D000818:Animals; D001424:Bacterial Infections / Q000235:genetics / Q000276:immunology; D002460:Cell Line; D016207:Cytokines / Q000378:metabolism*; D007371:Interferon-gamma / Q000494:pharmacology; D046911:Macromolecular Substances; D008262:Macrophage Activation; D008264:Macrophages / Q000187:drug effects / Q000276:immunology*; D051379:Mice; D008810:Mice, Inbred C57BL; D008813:Mice, Inbred ICR; D008969:Molecular Sequence Data; D011135:Polysaccharides, Bacterial / Q000494:pharmacology; D012333:RNA, Messenger / Q000096:biosynthesis; D015843:Serpins / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism*; D015533:Transcriptional Activation; D025801:Ubiquitin / Q000235:genetics; D014452:Ubiquitins; D015854:Up-Regulation
D016207:Cytokines; C489550:G1p2 protein, mouse; D046911:Macromolecular Substances; D011135:Polysaccharides, Bacterial; D012333:RNA, Messenger; C490400:Serpina3k protein, mouse; D015843:Serpins; D025801:Ubiquitin; D014452:Ubiquitins; D007371:Interferon-gamma