Production of enhanced silk fibers through genetic integration of Antheraea yamamai (Lepidoptera: Saturniidae) fibroin gene into Bombyx mori (Lepidoptera: Bombycidae)

Abstract Numerous insect species are capable of producing silk fibers with superior properties. For example, Antheraea yamamai Guérin-Méneville (Lepidoptera: Saturniidae) produces silk fibers with a range of compelling attributes, including notable gloss, durability, compressive elasticity, chemical resistance, and favorable mechanical properties. However, the practical utilization of their silk fibers remains limited due to the challenges associated with their large-scale rearing and fiber extraction. In contrast, the silkworm, Bombyx mori Linnaeus (Lepidoptera: Bombycidae), has been reared in large-scale from thousands of years as the major silk contributor. Furthermore, B. mori has been developed as an excellent bioreactor to express exogenous proteins through genetic manipulation. In this study, we molecularly cloned the 8,565 bp full length of the silk fibroin heavy chain gene (AyFibH) from A. yamamai and successfully generated transgenic silkworms expressing the 257 kDa fusion AyFibH protein specifically in their silk glands and cocoons. This was accomplished using a transposon-mediated transgenic strategy driven by the promoter of the B. mori fibroin heavy chain gene (BmFibH). As a result, the produced silk contained approximately 6% (w/w) of the heterologously expressed A. yamamai fibroin relative to the total silk proteins, contributing to a marked enhancement in the mechanical properties. According to structural analysis, the presence of unique polyalanines within the repetitive sequences of AyFibH is primarily responsible for a considerable rise in fiber crystallinity of approximately 44%. These findings demonstrate the feasibility of improving silk performance through the genetic integration of foreign fibroin genes, providing a potential strategy for biomaterial innovation.