Individual tissue-expression of BmorCSP mRNAs.

<p>Agarose gel electrophoresis of BmorCSP1-, BmorCSP2-, BmorCSP4-, BmorCSP14-, PBP1- and Actin4-encoding cDNA PCR products from the antennae (A), legs (L), head (H), pheromone gland (P) and wings (W) from five individual newly-emerged virgin females of the silkworm moth <i>B. mori</i> (Fm1, Fm2, Fm3, Fm4 and Fm5). Genomic DNA (g) was amplified in the same conditions, showing no differences among individuals.</p

Schematic organization of BmorCSP genes.

<p>The four BmorCSPs (BmorCSP1, BmorCSP2, BmorCSP4 and BmorCSP14) matched scaffold nsaf2767 in positions 270K-360K. “BGIBMGA” are gene names identified for BmorCSPs in silkDB. Grey boxes indicate complete genes. The pointing arrow refers to the orientation of the gene (5′ to 3′). Exons are shown as black boxes and intron regions as dotted lines. Repetitive elements are shown as black-bordered white boxes. The numbers on the scale represent the position of the genes in the scaffold. <i>BmorCSP4</i> is the largest gene (1978 bps) and sits distantly from <i>BmorCSP1</i>, <i>BmorCSP2</i> and <i>BmorCSP14</i>.</p

Total number of RDDs on cDNA of <i>B. mori</i> CSP-RNAs depending on gene structure and tissue expression.

<p>Total number of RDDs on cDNA of <i>B. mori</i> CSP-RNAs depending on gene structure and tissue expression.</p

Amino acid mutations on BmorCSPs in the pheromone gland.

<p>Alignment shows the amino acid composition of proteins encoded by genomic DNA. Conserved amino acid residues are shown in grey. Mutations sites on the native protein are underlined. Mutant peptide motifs are shown in bold. The arrowhead indicates amino acid insertion (mainly G). The dash above residues indicates amino acid inversion. The cross (x) indicates amino acid deletion in the motif. The grey circles below the alignment indicate the position of functional elements such as α-helices <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0086932#pone.0086932-Jansen2" target="_blank">[36]</a>.</p

Functional expression of CSP-RDDs.

<p>Electrophoretic separation and Western blot analysis of CSP proteins in the antennae (A), legs (L), heads (H), pheromone gland (P) and wings (W) of female <i>B. mori</i>. Protein extracts corresponding to 10 antennae, 30 legs, 5 heads, 5 pheromone glands and 20 wings equivalent were subjected to 15% SDS-PAGE (A. and D.) and transferred to nitrocellulose membranes (B–C. and E.F.). Nitrocellulose blots were labeled with two different antisera: “anti-CSP1” (B. and E.) and “antiCSP14” (C. and F.). Positions of molecular weight markers are indicated on the left of the gel. Proteins of 9 to 14 kDa are labeled with the two CSP antibodies in the pheromone gland, legs and wings samples.</p

Tissue-specific editing on cDNA of BmorCSP mRNAs.

<p>Sequence analysis of cDNA PCR products encoding BmorCSP1 reveals nucleotide insertion, deletion and substitution at the editing sites (black rectangles) in the antennae, head, legs, pheromone gland (Pgland) and wings. The size of the black rectangle is proportional to the frequency of RDDs at this location. The consensus sequence below the alignment corresponds to the nucleotide composition of the genomic DNA sequence (gDNA) encoding BmorCSP1. The number in brackets next to tissue indicates the number of CSP clones obtained for each tissue cDNA and gDNA. “.” indicates that the base is similar to the consensus sequence on this location. “A”, “T”, “G”, “C” point out a switch to adenosine, thymidine, guanosine and cytosine base in tissue-specific cDNA sequences, respectively. “1⁻” indicates base deletion in one sequence of the tissue group. “n^A” indicates switch to A in n sequences of the tissue group. Number of mismatches seen in only one tissue: 38. Number of mismatches seen in two tissues: 4.</p