Abstract
Cyclophilins (CYPs) are a group of ubiquitous proteins characterized by their ability to bind to the immunosuppressive drug cyclosporin A. The CYP family occurs in a wide range of organisms and contains a conserved peptidyl-prolyl cis/trans isomerase domain. In addition to fulfilling a basic role in protein folding, CYPs may also play diverse important roles, e.g. in protein degradation, mRNA processing, development, and stress responses. We performed a genome-wide database survey and identified a total of 94 CYP genes encoding 91 distinct proteins. Sequence alignment analysis of the putative BnCYP cyclophilin-like domains revealed highly conserved motifs. By using RNA-Seq, we could verify the presence of 77 BnCYP genes under control conditions. To identify phloem-specific BnCYP proteins in a complementary approach, we used LC-MS/MS to determine protein abundances in leaf and phloem extracts. We detected 26 BnCYPs in total with 12 being unique to phloem sap. Our analysis provides the basis for future studies concentrating on the functional characterization of individual members of this gene family in a plant of dual importance: as a crop and a model system for polyploidization and long-distance signalling.
Highlights
As one of the most important crops for nutritional oil, fodder, biodiesel, chemical and pharmaceutical products, Brassica napus is widespread in agriculture, especially in the European Union, China, and Canada
Putative CYPs of B. napus containing full length or partial CYP-like domains were identified by BLASTp of A. thaliana CYPs47, 48
The 94 B. napus gene sequences determined by this approach resulted in 91 distinct proteins which were subjected to additional analysis like sequence alignments with the respective Arabidopsis homologs and the verification of cyclophilin-like domain (CLD)
Summary
As one of the most important crops for nutritional oil, fodder, biodiesel, chemical and pharmaceutical products, Brassica napus (oilseed rape or canola) is widespread in agriculture, especially in the European Union, China, and Canada. Despite the lack of sequence and structure similarity, FKBPs and CYPs each possess a conserved domain responsible for their common peptidyl-prolyl cis/trans isomerase (PPIase) activity, catalyzing the rate-limiting rotation of X-proline peptide bonds from a cis to a trans conformation[14] These domains are called the FKBP and cyclophilin-like domain (CLD), respectively. The drugs of CYPs and FKBPs are cyclosporin A (CsA) and FK506/rapamycin, respectively, that bind to the catalytic pocket of the PPIase domain[16], thereby inhibiting its activity and forming a high-affinity binding site for the interaction with calcineurin[17] These drugs do not occur naturally in cells, the consequences of drug treatment have clinical but no physiological relevance[18]. To identify CYP proteins occurring in phloem sap, we performed complementary protein analyses by LC-MS/MS on leaf and phloem extracts and found 26 different BnCYP proteins in total, 12 only present in phloem sap
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