Uncharacterized protein with amino acid deletions from Bombyx mori illustrates divergence from Bombyx mandarina hemocytin and showed modulated gene expression after infection by Nosema bombycis

Genome wide gene expression analysis, transcription analysis and mass spectrometry have revealed several uncharacterized genes and proteins from commercial silkworm Bombyx mori . Infection of B. mori larvae with microsporidian Nosema bombycis induced appearance of exclusive proteins in hemocytes of B. mori showing host response. Mass spectrometry of the exclusive proteins revealed abundance of an uncharacterized protein H9JAZ8 that displayed conserved domains of Von Willebrand Factor type D domain (VWF), C8 domain, trypsin inhibitor-like cysteine rich domain and conserved epidermal growth factor like. Protein model of H9JAZ8 confirmed presence of VWF domains, cysteine rich domains and disulphide bonds. Expression of H9JAZ8 gene showed upregulation on day 2 after the infection followed by significant down regulated expression till day 10 exhibiting infection- associated modulation of the gene. BLASTP of H9JAZ8 showed 99% amino acid sequence similarity with B. mori BGIBMGA006693 demonstrating H9JAZ8 as a paralog of B. mori hemocytin. BLASTP of H9JAZ8 revealed 18 most similar hemocytin homologues from different insect species with 99% similarity for B. mandarina hemocytin. A Maximum Parsimony phylogeny tree showed three clusters where H9JAZ8 branched from the hemocytin of B. mandarina with 100% bootstrap value. Global BLAST analysis of H9JAZ8 with hemocytin of B. mandarina (XP_028038271.1) showed deletion of two regions, a 27 amino acid stretch, RRIFFSFILIYCTFNCEAGYGIPASSD at 2–28th position and a 21 amino acid stretch, DVVVISPNHHNNSSYYSYSQG at 317-338th position. Togteher, B. mori H9JAZ8 is diverged from B. mandarina hemocytin as part of speciation and immune evolution during domestication from the wild silkworm.