Effect of BmPriL on the development of silk glands in silkworm

Silk proteins have important economic implications. Silk glands are the only organs in silkworms that synthesize and secrete silk proteins. Ser1 (Ser1P) and Fib-H promoters (FibHP) are expressed in the middle (MSG) and posterior silk glands (PSG), respectively. The primase large subunit (PriL) is involved in DNA replication. However, its role in regulating silk gland development and the underlying mechanisms remain unclear. In this study, we constructed transgenic vectors, pb-MOE and pb-POE, in which BmPriL was overexpressed under the control of Ser1P and FibHP, respectively. The corresponding transgenic lines, M-OE and P-OE, were generated using embryo microinjections. The BmPriL content, length and weight of the silk glands, and surface area and DNA content of the silk gland cells were significantly increased in M-OE and P-OE silkworms than in the control silkworms. Using CRISPR/Cas9, BmPriL was knocked out of MSG and PSG cells. The corresponding lines were referred to as M-KO and P-KO. The phenotypes observed in M-KO and P-KO were contrary to those observed in M-OE and P-OE. Transcriptome analysis and qPCR verification revealed no apparent differences in silk protein gene expression levels between knockout strains and control silkworms. The H, F, and G subunits of eukaryotic translation initiation factor 3 (eIF3) were significantly downregulated in M-KO and P-KO. These results indicate that the BmPriL knockout decreases in eIF3 expression, thereby inhibiting silk protein synthesis and reducing silk production.