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Conservation of silk genes in Trichoptera and Lepidoptera.

Yonemura N, Mita K, Tamura T, Sehnal F - J. Mol. Evol. (2009)

Bottom Line: They are extremely uniform in R. obliterata.The trichopteran H-fibroins further contain charged amphiphilic motifs but lack the strings of alanines or alanine-glycine dipeptides that are typical lepidopteran motifs.On the other hand, sequences composed of a motif similar to ERIVAPTVITR surrounded by the (SX)(4-6) strings and modifications of the GRRGWGRRG motif occur in Trichoptera and not in Lepidoptera.

View Article: PubMed Central - PubMed

Affiliation: National Institute of Agrobiological Sciences, Tsukuba, Ibaraki, 305-8634, Japan.

ABSTRACT
Larvae of the sister orders Trichoptera and Lepidoptera are characterized by silk secretion from a pair of labial glands. In both orders the silk filament consists of heavy (H)- and light (L)-chain fibroins and in Lepidoptera it also includes a P25 glycoprotein. The L-fibroin and H-fibroin genes of Rhyacophila obliterata and Hydropsyche angustipennis caddisflies have exon/intron structuring (seven exons in L-fibroin and two in H-fibroin) similar to that in their counterparts in Lepidoptera. Fibroin cDNAs are also known in Limnephilus decipiens, representing the third caddisfly suborder. Amino acid sequences of deduced L-fibroin proteins and of the terminal H-fibroin regions are about 50% identical among the three caddisfly species but their similarity to lepidopteran fibroins is <25%. Positions of some residues are conserved, including cysteines that were shown to link the L-fibroin and H-fibroin by a disulfide bridge in Lepidoptera. The long internal part of H-fibroins is composed of short motifs arranged in species-specific repeats. They are extremely uniform in R. obliterata. Motifs (SX)(n), GGX, and GPGXX occur in both Trichoptera and Lepidoptera. The trichopteran H-fibroins further contain charged amphiphilic motifs but lack the strings of alanines or alanine-glycine dipeptides that are typical lepidopteran motifs. On the other hand, sequences composed of a motif similar to ERIVAPTVITR surrounded by the (SX)(4-6) strings and modifications of the GRRGWGRRG motif occur in Trichoptera and not in Lepidoptera.

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Similarities in the H-fibroin gene among Trichoptera and Lepidoptera (nucleotides conserved in at least two caddisfly species are highlighted in gray); no sequence homologies were detected upstream or downstream from the depicted regions. Exons are in capital letters. a The H-fibroin 5′ region in R. obliterata (Ro; GenBank accession no. AB354689), H. angustipennis (Ha; AB354591), G. mellonella (Gm; AF095239), and B. mori (Bm; AF226688). Sequences are numbered from the transcription start, the TATA box is underlined with a straight line, the initiator consensus sequence (AyCAGyyy) with a wavy line, and the translation initiation codon is double-underlined. Intron donor (gtaagt) and acceptor (tcttattttag) sites are underlined in the Ha sequence. b Alignment of the H-fibroin 3′ region in R. obliterata (Ro; GenBank accession no. AB354588), H. angustipennis (Ha; AB214507), L. decipiens (Ld; AB214509), Y. evonymella (Ye; AB195978), G. mellonella (Gm; AF095240), and B. mori (Bm; AF226688\0. The termination codon is in boldface
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Fig5: Similarities in the H-fibroin gene among Trichoptera and Lepidoptera (nucleotides conserved in at least two caddisfly species are highlighted in gray); no sequence homologies were detected upstream or downstream from the depicted regions. Exons are in capital letters. a The H-fibroin 5′ region in R. obliterata (Ro; GenBank accession no. AB354689), H. angustipennis (Ha; AB354591), G. mellonella (Gm; AF095239), and B. mori (Bm; AF226688). Sequences are numbered from the transcription start, the TATA box is underlined with a straight line, the initiator consensus sequence (AyCAGyyy) with a wavy line, and the translation initiation codon is double-underlined. Intron donor (gtaagt) and acceptor (tcttattttag) sites are underlined in the Ha sequence. b Alignment of the H-fibroin 3′ region in R. obliterata (Ro; GenBank accession no. AB354588), H. angustipennis (Ha; AB214507), L. decipiens (Ld; AB214509), Y. evonymella (Ye; AB195978), G. mellonella (Gm; AF095240), and B. mori (Bm; AF226688\0. The termination codon is in boldface

Mentions: The H-fibroin gene of Lepidoptera contains a large intron not far from the start of the coding sequence. The existence of a similar arrangement in the H-fibroin of caddisflies was probed by PCR based on primers from the nonrepetitive 5′ cDNA region. A 3-kb fragment was amplified from the genomic DNA of R. obliterata larva designated Ro01 with primers RoH-F21 and RoH-R11 (Table 1). The sequence of this fragment revealed overlap with the cDNA sequence and disclosed the presence of an intron 2645 bp long (Fig. 5a). To determine the upstream gene region, the Ro01 DNA was digested with a mix of EcoRI and MunI, self-ligated, and used for inverse PCR with primers RoH-R31 and RoH-F31 (Table 1). A 3.5-kb PCR product extended from a MunI restriction site in the intron (+425 from the transcription start) to a MunI site at –3550. A combination of this sequence with that of the 3-kb PCR product yielded a contig that spanned from –3550 to +3017 and. in the regions 1–42 (first exon) and 2713–3655 (part of the second exon), matched precisely the cDNA sequence established with 5′ RACE. Since the cDNA contained an additional 638 nt, the 5′ part of the H-fibroin gene (Ro03HF5′) deposited in GenBank under accession no. AB354689 covers the region from –3550 to +3655. The 3′ end of the R. obliterataH-fibroin gene (Fig. 5b) was analyzed only at the cDNA level.Fig. 5


Conservation of silk genes in Trichoptera and Lepidoptera.

Yonemura N, Mita K, Tamura T, Sehnal F - J. Mol. Evol. (2009)

Similarities in the H-fibroin gene among Trichoptera and Lepidoptera (nucleotides conserved in at least two caddisfly species are highlighted in gray); no sequence homologies were detected upstream or downstream from the depicted regions. Exons are in capital letters. a The H-fibroin 5′ region in R. obliterata (Ro; GenBank accession no. AB354689), H. angustipennis (Ha; AB354591), G. mellonella (Gm; AF095239), and B. mori (Bm; AF226688). Sequences are numbered from the transcription start, the TATA box is underlined with a straight line, the initiator consensus sequence (AyCAGyyy) with a wavy line, and the translation initiation codon is double-underlined. Intron donor (gtaagt) and acceptor (tcttattttag) sites are underlined in the Ha sequence. b Alignment of the H-fibroin 3′ region in R. obliterata (Ro; GenBank accession no. AB354588), H. angustipennis (Ha; AB214507), L. decipiens (Ld; AB214509), Y. evonymella (Ye; AB195978), G. mellonella (Gm; AF095240), and B. mori (Bm; AF226688\0. The termination codon is in boldface
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Related In: Results  -  Collection

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Fig5: Similarities in the H-fibroin gene among Trichoptera and Lepidoptera (nucleotides conserved in at least two caddisfly species are highlighted in gray); no sequence homologies were detected upstream or downstream from the depicted regions. Exons are in capital letters. a The H-fibroin 5′ region in R. obliterata (Ro; GenBank accession no. AB354689), H. angustipennis (Ha; AB354591), G. mellonella (Gm; AF095239), and B. mori (Bm; AF226688). Sequences are numbered from the transcription start, the TATA box is underlined with a straight line, the initiator consensus sequence (AyCAGyyy) with a wavy line, and the translation initiation codon is double-underlined. Intron donor (gtaagt) and acceptor (tcttattttag) sites are underlined in the Ha sequence. b Alignment of the H-fibroin 3′ region in R. obliterata (Ro; GenBank accession no. AB354588), H. angustipennis (Ha; AB214507), L. decipiens (Ld; AB214509), Y. evonymella (Ye; AB195978), G. mellonella (Gm; AF095240), and B. mori (Bm; AF226688\0. The termination codon is in boldface
Mentions: The H-fibroin gene of Lepidoptera contains a large intron not far from the start of the coding sequence. The existence of a similar arrangement in the H-fibroin of caddisflies was probed by PCR based on primers from the nonrepetitive 5′ cDNA region. A 3-kb fragment was amplified from the genomic DNA of R. obliterata larva designated Ro01 with primers RoH-F21 and RoH-R11 (Table 1). The sequence of this fragment revealed overlap with the cDNA sequence and disclosed the presence of an intron 2645 bp long (Fig. 5a). To determine the upstream gene region, the Ro01 DNA was digested with a mix of EcoRI and MunI, self-ligated, and used for inverse PCR with primers RoH-R31 and RoH-F31 (Table 1). A 3.5-kb PCR product extended from a MunI restriction site in the intron (+425 from the transcription start) to a MunI site at –3550. A combination of this sequence with that of the 3-kb PCR product yielded a contig that spanned from –3550 to +3017 and. in the regions 1–42 (first exon) and 2713–3655 (part of the second exon), matched precisely the cDNA sequence established with 5′ RACE. Since the cDNA contained an additional 638 nt, the 5′ part of the H-fibroin gene (Ro03HF5′) deposited in GenBank under accession no. AB354689 covers the region from –3550 to +3655. The 3′ end of the R. obliterataH-fibroin gene (Fig. 5b) was analyzed only at the cDNA level.Fig. 5

Bottom Line: They are extremely uniform in R. obliterata.The trichopteran H-fibroins further contain charged amphiphilic motifs but lack the strings of alanines or alanine-glycine dipeptides that are typical lepidopteran motifs.On the other hand, sequences composed of a motif similar to ERIVAPTVITR surrounded by the (SX)(4-6) strings and modifications of the GRRGWGRRG motif occur in Trichoptera and not in Lepidoptera.

View Article: PubMed Central - PubMed

Affiliation: National Institute of Agrobiological Sciences, Tsukuba, Ibaraki, 305-8634, Japan.

ABSTRACT
Larvae of the sister orders Trichoptera and Lepidoptera are characterized by silk secretion from a pair of labial glands. In both orders the silk filament consists of heavy (H)- and light (L)-chain fibroins and in Lepidoptera it also includes a P25 glycoprotein. The L-fibroin and H-fibroin genes of Rhyacophila obliterata and Hydropsyche angustipennis caddisflies have exon/intron structuring (seven exons in L-fibroin and two in H-fibroin) similar to that in their counterparts in Lepidoptera. Fibroin cDNAs are also known in Limnephilus decipiens, representing the third caddisfly suborder. Amino acid sequences of deduced L-fibroin proteins and of the terminal H-fibroin regions are about 50% identical among the three caddisfly species but their similarity to lepidopteran fibroins is <25%. Positions of some residues are conserved, including cysteines that were shown to link the L-fibroin and H-fibroin by a disulfide bridge in Lepidoptera. The long internal part of H-fibroins is composed of short motifs arranged in species-specific repeats. They are extremely uniform in R. obliterata. Motifs (SX)(n), GGX, and GPGXX occur in both Trichoptera and Lepidoptera. The trichopteran H-fibroins further contain charged amphiphilic motifs but lack the strings of alanines or alanine-glycine dipeptides that are typical lepidopteran motifs. On the other hand, sequences composed of a motif similar to ERIVAPTVITR surrounded by the (SX)(4-6) strings and modifications of the GRRGWGRRG motif occur in Trichoptera and not in Lepidoptera.

Show MeSH