A cell-specific glycosylated silk protein from Chironomus thummi salivary glands. Cloning, chromosomal localization, and characterization of cDNA.
Chironomid salivary glands contain 40 cells dedicated to the synthesis of a relatively small ensemble of silk proteins. Glands in some species contain a special lobe composed of 4 cells distinguishable from the others. We have cloned a special lobe-specific cDNA from Chironomus thummi salivary glands. Northern blots of salivary gland RNA demonstrated that the cDNA hybridizes to a 2.5-kilobase transcript present only in the special lobe. In situ hybridization mapped the gene encoding this cDNA to region A2b on polytene chromosome IV, the locus of the special lobe-specific Balbiani ring a. The deduced amino acid sequence encodes a protein with a calculated molecular mass of 77 kDa and numerous potential glycosylation sites; it appears unrelated to other known chironomid silk proteins. Polyclonal antibody, raised against a cDNA-encoded fusion protein, reacted exclusively with a special lobe-specific 160-kDa silk protein. Lectin binding studies indicate that the immunoreactive 160-kDa protein contains both N- and O-linked glycan moieties. We conclude that glycosylation most likely contributes to the difference between calculated and apparent molecular masses and that this cDNA encodes the special lobe-specific silk protein previously described as ssp160 (Kolesnikov, N. N., Karakin, E. I., Sebeleva, T. E., Meyer, L., and Serfling, E. (1981) Chromosoma 83, 661-677).
Hoffman
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Department of Biochemistry, University of Mississippi Medical Center, Jackson, Mississippi 39216-4505, USA.
Schmidt
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Case
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D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D015153:Blotting, Western; D002683:Chironomidae / Q000235:genetics* / Q000378:metabolism*; D002874:Chromosome Mapping; D003001:Cloning, Molecular; D018076:DNA, Complementary; D006023:Glycoproteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics*; D006031:Glycosylation; D017403:In Situ Hybridization; D019476:Insect Proteins; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D012091:Repetitive Sequences, Nucleic Acid; D012469:Salivary Glands / Q000166:cytology / Q000378:metabolism*; D012471:Salivary Proteins and Peptides / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics*; D047011:Silk
D018076:DNA, Complementary; D006023:Glycoproteins; D019476:Insect Proteins; D011994:Recombinant Proteins; D012471:Salivary Proteins and Peptides; D047011:Silk; C099286:ssp160 protein, silk