Serine/arginine-rich proteins are a class of highly conserved splicing factor proteins involved in constitutive and alternative splicing. We screened a low molecular weight serine/arginine rich protein from silkworms and named it BmUP. Temporal and spatial expression analysis indicated that the BmUP gene was specifically expressed in the silkworm testis, and the highest expression occurred in the pre-pupa stage from the fifth instar to the moth stages. Here, we generated BmUP knockout individuals with the CRISPR/Cas9 system. Both the internal and external genitalia of knockout individuals were abnormal in knockout compared with wild-type male silkworms. In transgenic silkworms overexpressing BmUP, male silkworms showed a phenotype similar to that of the knockout individuals, whereas female individuals showed no significant differences from the wild type. In addition, by conducting promoter analysis, we identified Bmachi, a transcription factor that regulates the BmUP gene. Gel migration experiments revealed that BmAchi specifically binds the BmUP promoter. Quantitative real-time PCR showed that an increase in Bmachi expression up-regulated the expression of BmUP. In contrast, when the expression of Bmachi decreased, the expression of BmUP also downregulated in the experimental group compared with the control group. These results provide new insights for studying the effects of serine/arginine-rich proteins on the development of silkworm genitals.
Keywords: BmUP; Bombyx mori; CRISPR/Cas9; serine/arginine-rich; transgene overexpression.