Diversity of Coding Sequences and Gene Structures of the Antifungal Peptide Mytimycin (MytM) from the Mediterranean Mussel, Mytilus galloprovincialis

Abstract

Knowledge on antifungal biomolecules is limited compared to antibacterial peptides. A strictly antifungal peptide from the blue mussel, Mytilus edulis named mytimycin (MytM) was reported in 1996 as partial NH(2) 33 amino acid sequence. Using back-translations of the previous sequence, MytM-related nucleotide sequences were identified from a normalized Mytilus galloprovincialis expressed sequence tag library. Primers designed from a consensus sequence have been used to obtain a fragment of 560 nucleotides, including the complete coding sequence of 456 nucleotides. Precursor is constituted by a signal peptide of 23 amino acids, followed by MytM of 54 amino acids (6.2-6.3 kDa, 12 cysteines) and C-terminal extension of 75 amino acids. Only two major amino acid precursor sequences emerged, one shared by M. galloprovincialis from Venice and Vigo, the other belonging to M. galloprovincialis from Palavas, with nine amino acid differences between the two MytM. Predicted disulfide bonds suggested the presence of two constrained domains joined by amino acidic NIFG track. Intriguing was the presence of conserved canonical EF hand-motif located in the C-terminus extension of the precursor. The MytM gene was found interrupted by two introns. Intron 2 existed in two forms, a long (1,112 nucleotides) and a short (716 nucleotides) one resulting from the removal of the central part of the long one. Both the short (GenBank FJ804479) and the long (GenBank FJ804478) genes are simultaneously present in the mussel genome.

Gene expression specificity of the mussel antifungal mytimycin (MytM)

“…In a previous report we analyzed the diversity of MytM mRNA purified from hemocytes [34]. In the present work, we quantified such mRNA in several tissues.…”

“…Recently, we isolated 16 nucleotide sequences related to MytM from a M. galloprovincialis EST library [37] and established the full cds of the precursor [34]. It possesses the same construction as other antibacterial peptides: defensin, mytilin and myticin, i.e.…”

“…In addition, 2 major deduced aa precursor sequences were observed, one shared by the mussels from Venice-Italy and Vigo-Spain, the other belonged to those mussels from Palavas-France. Predicted disulfide bonds of these 12 cysteine peptides suggested the presence of 2 constrained domains [34]. Recently, a partial 47 aa (no cysteine) antifungal peptide, but sharing also a weak effect on bacteria, has been isolated from a native Chilean scallop, Argopecten purpuratus, and used as framework to design substitutes for conventional antibiotics [2].…”

Molecular diversity and evolution of defensins in the manila clam Ruditapes philippinarum

“…The occurrence frequencies of different Rpdefs varied in clams from different geographical locations. The transcript variation was perhaps related to pathogen load in clams and other possible environmental factors of different sites, which may have forced the defensin genes to diversify [43]. The transcript sequences of Rpdef2 and Rpdef3 were almost averagely present in three tissues, while the transcript sequences of Rpdef1 were mainly present in hemocytes and Rpdef4 were mainly detected in gills and digestive gland.…”

MIF from mussel: Coding sequence, phylogeny, polymorphism, 3D model and regulation of expression

“…Among the remaining amino acid sequences, 9 were encountered only once or twice. Several other proteins were previously reported as highly polymorphic in mussels: antimicrobial peptides defensin, mytilin (Parisi et al, 2009) and particularly myticin (Costa et al, 2009;Pallavicini et al, 2008), C1q-like (Gerdol et al, 2011;Gestal et al, 2010), FREPs (Romero et al, 2011) and antifungal mytimycin (Sonthi et al, 2011). Although not understood, no doubt that the existence of numerous variants probably corresponds to as yet undefined functional mechanism(s).…”