Acidification of the Silk Gland Lumen in Bombyx mori and Samia cynthia ricini and Localization of H⁺-Translocating Vacuolar-Type ATPase

Abstract

The silk gland of Bombyx mori and Samia cynthia ricini produces vast amounts of silk proteins and stores them in the glandular lumen as a liquid silk during the larval growth and development. We have explored the system regulating pH in the silk gland, because the gelation of fibroin is pH-dependent. By injecting the pH-sensitive dye (phenol red) into silkworm larvae, we have estimated the pH in the glandular lumen. Although the entry of dye was unsuccessful in the anterior silk gland (ASG) of Bombyx, the lumen of the middle silk gland (MSG: major reservoir for fibroin) and that of the posterior silk gland (PSG: the fibroin factory) were colored with phenol red. The coloration by phenol red indicated that the MSG was acidic (pH 5~6) in the vigorously feeding larvae leading to gelation of silk proteins at the MSG and that the PSG was neutral (pH 7~8). When the larvae started spinning, the lumen in the MSG became neutral. A similar pattern in the luminal pH shift was obtained in the silk gland of Samia cynthia ricini (Eri-silkworm) with a dye-injection experiment. In Samia, the H⁺-translocating vacuolar-type ATPase (V-ATPase) locates at the apical surface of PSG, where the fibroin is produced, secreted and temporarily stored. The V-ATPase was also distributed at the apical surface of the anterior MSG as well as that in ASG. These V-ATPases became undetectable after the onset of spinning. The V-ATPase at the plasma membrane of silk gland cells regulates the physico-chemical state of liquid silk in the glandular lumen, in particular at the MSG of Bombyx and at the PSG of Samia, respectively.