Reduced Level of Secretion and Absence Combination for the Fibroin Synthesized Silkworm, Nd(2) of Subunit by a Mutant

Fibroin is normally composed of one H chain (350 kd) and one L chain (25 kd) which are connected by disulfide bond(s). However, the small amount of fibroin secreted into the lumen of the posterior silk gland of the Nd(2) (naked pupa) mutant does not contain L chain, although L chain mRNA is present and L chain is synthesized in the posterior silk gland cells of the mutant. In a hybrid silkworm, Nd(2)/Tamanashikasuri, where Tamanashikasuri is a normal producer of fibroin, L chain from the two alleles are distinguishable electrophoretically. It is demonstrated using this system that the L chain from the Nd(2) allele can combine normally with the H chain from Tamanashikasuri and the H-L complex is secreted normally. In another hybrid system, Nd(2)[J-131, where J-131 is a normal producer of fibroin, fibroin derived from the two alleles are distinguishable due to the different electrophoretic mobility of H chain. The fibroin derived from the J-131 allele is composed of H chain and L chain, while the fibroin derived from the Nd(2) allele is devoid of L chain, and its secretion is greatly reduced. We present evidence suggesting that the H chain derived from the Nd(2) allele is structurally abnormal and discuss how the H-L subunit structure is advantageous in the secretion of fibroin. The silk gland of silkworm, Bombyx mori, secretes two kinds of proteins, fibroin from the posterior silk gland and sericin from the middle silk gland. It has been shown that the native fibroin extracted from the lumen of posterior or middle silk gland is composed of a large and a small component connected by disulfide bond(s) (1-3). The molar ratio of the two polypeptides is one to one (3). The large component (350 kd) has an amino acid composition of so-called "fibroin" type, while the small component (25 kd) has a markedly different amino acid composition (4). In this study, we name the large component as heavy chain (H chain) and the small component as light chain (L chain). The H chain has been studied extensively from various aspects (5-8), while the L chain is poorly characterized. Recently mRNA for the L chain was identified in an in vitro translation system coupled with immunological detection with an antiserum against the L chain (9). In the present study, we compared subunit structures of fibroin synthesized by a mutant silkworm, Nd(2), and by strains producing normal levels of fibroin (normal producers). The "naked pupa" strain (symbolized as Nd) was found as a naturally occurring mutant (10). Nd(2), one of the Nd-type strains, produces a thin cocoon that consists mostly ofsericin. The posterior silk gland of Nd(2) (Fig. I A) is extremely short in size, undeveloped, and produces little fibroin. It has been shown, however, that the number of cells present in the posterior silk gland of Nd(2) is the same as that of a normal producer (10). The middle silk gland of Nd(2) is normal and secretes a normal level of sericin. The structural gene of the H chain and the locus responsible for the Nd(2) mutation have been shown to be linked and both are present on the 25th chromosome (11). ~ The H chain gene was originally assigned to the 23rd chromosome because of its linkage with the Nd(2) locus (11). However, location of Nd(2) has recently been corrected to be on the 25th chromosome (12). Accordingly, the H chain gene is assigned to the 25th chromosome. THE JOURNAL OF CELL BIOLOGY VOLUME 99 DECEMBER 198,4 2005-2010 © The Rockefeller University Press • 0021-9525/84/12/2005106 $1.0