Comprehensive Transcriptome Analysis in the Testis of the Silkworm, Bombyx mori

SIMPLE SUMMARY: Spermatogenesis, an important process in reproduction, is conserved across species but has unique aspects in silkworms, such as the maintenance of the spermatogonium by a somatic single cell and coordinated fertilization by nucleated and anucleated sperm. Despite some histological research, the molecular mechanisms of silkworm spermatogenesis remain largely unexplored. Taking advantage of advances in RNA-seq analysis technology, this study characterizes genes expressed in silkworm testis via a comprehensive transcriptome analysis, contrasting different tissues and regions within the testis and making comparisons with fruit flies. Our investigation revealed extensive gene expression (86.5% of all genes) in the silkworm testis, the highest number of tissue-specific genes among examined tissues in silkworm testis. We also identified region-specific enriched genes in undifferentiated and differentiated germ cells. Moreover, cross-species transcriptome comparisons confirmed conserved gene expression patterns. Further analysis of region-specific enriched genes in the silkworm testis revealed the enrichment of genes associated with the biological process, corroborating findings in silkworms and other organisms. This study extends our understanding of silkworm spermatogenesis and fills an important gap for future investigations. ABSTRACT: Spermatogenesis is an important process in reproduction and is conserved across species, but in Bombyx mori, it shows peculiarities, such as the maintenance of spermatogonia by apical cells and fertilization by dimorphic spermatozoa. In this study, we attempted to characterize the genes expressed in the testis of B. mori, focusing on aspects of expression patterns and gene function by transcriptome comparisons between different tissues, internal testis regions, and Drosophila melanogaster. The transcriptome analysis of 12 tissues of B. mori, including those of testis, revealed the widespread gene expression of 20,962 genes and 1705 testis-specific genes. A comparative analysis of the stem region (SR) and differentiated regions (DR) of the testis revealed 4554 and 3980 specific-enriched genes, respectively. In addition, comparisons with D. melanogaster testis transcriptome revealed homologs of 1204 SR and 389 DR specific-enriched genes that were similarly expressed in equivalent regions of Drosophila testis. Moreover, gene ontology (GO) enrichment analysis was performed for SR-specific enriched genes and DR-specific enriched genes, and the GO terms of several biological processes were enriched, confirming previous findings. This study advances our understanding of spermatogenesis in B. mori and provides an important basis for future research, filling a knowledge gap between fly and mammalian studies.