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Expression and protease targets of the twelve serpin‐1 isoforms in the caterpillar Manduca sexta
Author(s) -
Ragan Emily Jane,
Kanost Michael R
Publication year - 2007
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/fasebj.21.5.a648-c
Subject(s) - serpin , proteases , protease , serine protease , masp1 , manduca sexta , biology , manduca , gene isoform , biochemistry , protease inhibitor (pharmacology) , microbiology and biotechnology , enzyme , immunology , virus , gene , botany , larva , antiretroviral therapy , viral load
There are 25 known serine proteases in the hemolymph of the caterpillar Manduca sexta, some of which are involved in aspects of innate immunity. Serine protease inhibitors (serpins) contribute to regulation of proteases by covalently binding to specific active proteases and forming serpin‐protease complexes. Serpin‐1 has an alternatively spliced ninth exon that codes for 12 serpin‐1 isoforms, which differ in inhibitory selectivity. Our goals are to understand which serpin‐1 isoforms are present in plasma, at what levels, and to identify protease targets of serpin‐1. Reverse‐transcriptase (RT) PCR showed that all 12 isoforms are expressed in hemocytes and fat body. We are using real‐time RT PCR to compare expression levels of the individual isoforms. We used antibodies to serpin‐1 to construct an affinity column for serpin‐1 isolate two serpin‐1‐protease complexes that formed after challenging plasma with bacterial lipopolysaccharide. MALDI‐TOF/TOF analysis of these serpin‐1‐protease complexes identified the digestive enzyme chymotrypsin as a specific target of serpin‐1K. Surprisingly chymotrypsin is expressed at low levels in hemocytes in addition to higher levels in midgut. Discovering the serpin‐1 isoforms that inhibit specific proteases enhances our understanding of proteolytic cascades in Manduca. Research supported by NIH R01‐GM41247