Functional characterization of Bmcap in uric acid metabolism in the silkworm

After a millennium of domestication,numerous silkworm mutants have emerged that exhibit transparent epidermis,which is caused by abnormally low levels of uric acid.We identified the Bombyx mori gene Bmcap(BMSK0003832)as the homolog of cappuccino,a subunit of the biogenesis of lysosome-related organelles complex-1(BLOC-1)that has been extensively characterized in human,mouse,and insect species,by analyzing the amino acid sequences of putative purine metabolism genes.Using the clustered regularly interspaced palindromic repeats(CRISPR)/CRISPR-associated protein 9 system,we disrupted Bmcap,resulting in decreased uric acid levels in the silkworm epidermis and a translucent skin phenotype.In the Bmcap mutant,the purine metabolism,nitrogen metabolism,pyrimidine metabolism,and membrane system were altered compared to the wild type.Biogenesis of lysosome-related organelle complex genes play a role in the pigmentation and biogenesis of lysosome-related organelles(LROs)in platelets,melanocytes,and megakaryocytes.LROs exhibit unique morphologies and functions in various tissues and cells.Investigation of the Bmcap mutant will enhance our understanding of the uric acid metabolic pathway in silkworms,and this mutant offers a valuable silkworm model for LRO studies.

The RNase III enzyme Dicer1 is essential for larval development in Bombyx mori

MicroRNAs(miRNAs)are important regulators of nearly all aspects of biological processes in eukaryotes.During the biogenesis of miRNAs,the RNase III enzyme Dicer processes double-strand precursor miRNAs into mature miRNAs and promotes the assembly of RNA-induced silencing complexes(RISCs).Dicer has been reported to participate in a wide range of physiological processes,including development and immunity,in some insect species.However,the physiological roles of Dicer in lepidopterans remain poorly understood.In this study,we investigated the function of Bombyx mori Dicer1.We first performed sequence alignment and found that the sequence of functional domains of Dicer1 are varied among Lepidoptera,Diptera,Coleoptera,Blattaria,and Orthoptera.Using a binary clustered regularly interspaced palindromic repeats(CRISPR)/CRISPR-associated protein 9 genome editing approach,we showed that BmDicer1 mutants have arrested development from the 3rd instar into the 4th instar.RNA sequencing analysis indicated that the defects in BmDicer1 mutants are due to dysregulation of genes that encode proteins involved in metabolism,protein degradation,absorption,and renin–angiotensin pathways.Analysis using quantitative real-time polymerase chain reaction showed that mutation of BmDicer1 altered expression of miRNAs and their target genes.Therefore,our study demonstrates the critical roles of BmDicer1 in miRNA biogenesis and larval development in silkworm.