Abstract
Simple SummaryThe mechanism of the polyamine pathway in the lepidopteran silkworm is largely unknown. In the current study, we aimed to characterize the function of polyamines and polyamine pathway genes in silkworm cells as a regulator of cell cycle progression. For the first time, we identified the homologous genes of the polyamine pathway in silkworm, and analyzed their expression characteristics in different tissues and their subcellular localizations in cultured silkworm cells. We measured the abundant levels of polyamines in silkworm cells by HPLC analysis. We found that exogenous supplementation of spermidine in cells promoted DNA replication and cell cycle progression and, in contrast, treatment with polyamine biosynthesis inhibitors DFMO and MGBG prevented DNA replication and cell cycle progression. Indeed, the mechanism studies indicated that spermidine increased the expression of cell cycle-related genes, whereas this increase could be abolished by treatment with inhibitors. Taken together, our findings highlight that appropriate levels of polyamines have beneficial effects on the progression of the cell cycle by regulating cell cycle genes in silkworm.Background: Putrescine, spermidine, and spermine are polyamines that are ubiquitously distributed in prokaryotic and eukaryotic cells, which play important roles in cell proliferation and differentiation. Methods: We investigated the expression profiles of polyamine pathway genes by qRT-PCR in different tissues of the lepidopteran silkworm. The polyamine levels in cultured silkworm cells were measured by HPLC. Spermidine and polyamine biosynthetic inhibitors were used for treating the cultured silkworm cells in order to clarify their effects on cell cycle progression. Results: We identified the anabolic and catabolic enzymes that are involved in the polyamine biosynthetic pathway in silkworm. Transcriptional expression showed at least seven genes that were expressed in different silkworm tissues. Treatments of the cultured silkworm cells with spermidine or inhibitor mixtures of DFMO and MGBG induced or inhibited the expression of cell cycle-related genes, respectively, and thus led to changed progression of the cell cycle. Conclusions: The present study is the first to identify the polyamine pathway genes and to demonstrate the roles of polyamines on cell cycle progression via regulation of the expression of cell cycle genes in silkworm.
Highlights
Polyamines are low-molecular-weight polycationic aliphatic amines derived from the metabolism of arginine, which mainly include putrescine, spermidine, and spermine, and are found in prokaryotic and eukaryotic cells [1,2]
In order to explore the function of this polyamine pathway in silkworm, we identified the genes in silkworm
We referred to proteins from the annotated species, including D. melanogaster and humans, and performed BLASTP analysis against the silkworm genome database, and named BmODC, BmSAMDC, BmSPDS, BmSPMS, BmSSAT, BmSMO, and BmAPAO (Figure 1B)
Summary
Polyamines are low-molecular-weight polycationic aliphatic amines derived from the metabolism of arginine, which mainly include putrescine, spermidine, and spermine, and are found in prokaryotic and eukaryotic cells [1,2]. Due to their polycationic nature, polyamines are fully protonated under physiological pH and ionic strength conditions. The biosynthesis of polyamines is regulated by multiple feedback loops that maintain the homeostasis of their metabolites in cells [7] It begins with the conversion of putrescine from the amino acid ornithine by the enzyme ornithine decarboxylase (ODC), a rate-limiting enzyme in polyamine synthesis [9,10]. Spermidine and polyamine biosynthetic inhibitors were used for treating the cultured silkworm cells in order to clarify their effects on cell cycle progression
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