Abstract

Silk is a protein of interest to both biological and industrial sciences. The silkworm, Bombyx mori, forms this protein into strong threads starting from soluble silk proteins using a number of biochemical and physical cues to allow the transition from liquid to fibrous silk. A pH gradient has been measured along the gland, but the methodology employed was not able to precisely determine the pH at specific regions of interest in the silk gland. Furthermore, the physiological mechanisms responsible for the generation of this pH gradient are unknown. In this study, concentric ion selective microelectrodes were used to determine the luminal pH of B. mori silk glands. A gradient from pH 8.2 to 7.2 was measured in the posterior silk gland, with a pH 7 throughout the middle silk gland, and a gradient from pH 6.8 to 6.2 in the beginning of the anterior silk gland where silk processing into fibers occurs. The small diameter of the most anterior region of the anterior silk gland prevented microelectrode access in this region. Using a histochemical method, the presence of active carbonic anhydrase was identified in the funnel and anterior silk gland of fifth instar larvae. The observed pH gradient collapsed upon addition of the carbonic anhydrase inhibitor meth- azolamide, confirming an essential role for this enzyme in pH regulation in the B. mori silk gland. Plastic embedding of whole silk glands allowed clear visualization of the morphology, including the identifi- cation of four distinct epithelial cell types in the gland and allowed correlations between silk gland morphology and silk stages of assembly related to the pH gradient. B. mori silk glands have four different epithelial cell types, one of which produces carbonic anhydrase. Carbonic anhydrase is necessary for the mechanism that generates an intraluminal pH gradient, which likely regulates the assembly of silk proteins and then the formation of fibers from soluble silk proteins. These new insights into native silk formation may lead to a more efficient production of artificial or regenerated silkworm silk fibers.

Keywords

Fibroin; Silk gland; Ion-selective microelectrodes; Morphology; Carbonic anhydrase

Published in

Insect Biochemistry and Molecular Biology
2015, Volume: 65, pages: 100-106

SLU Authors

• Andersson, Marlene

  • Department of Animal Biosciences, Swedish University of Agricultural Sciences
    

• Johansson, Jan

  • Department of Animal Biosciences, Swedish University of Agricultural Sciences
    
  • Karolinska Institute
  • Tallinn University

• Rising, Anna

  • Department of Animal Biosciences, Swedish University of Agricultural Sciences
    
  • Karolinska Institute

UKÄ Subject classification

Other Chemistry Topics


Publication identifier

DOI: https://doi.org/10.1016/j.ibmb.2015.09.001

Permanent link to this page (URI)

https://res.slu.se/id/publ/68672