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During miRNA biosynthesis, miRNA precursors are cleaved by Dicer at their loop structure to generate mature miRNAs. However, the functional consequences of sequence alterations in the loop region remain poorly understood. The silkworm\n                    <jats:italic>Bombyx mori<\/jats:italic>\n                    is a lepidopteran insect with many genetic strains. In a\n                    <jats:italic>B. mori<\/jats:italic>\n                    strain exhibiting translucent larval skin, we unexpectedly identified a mutant form of\n                    <jats:italic>miR-3260<\/jats:italic>\n                    in which part of the predicted loop structure was deleted. Here, we aimed to analyze the function of wild-type\n                    <jats:italic>miR-3260<\/jats:italic>\n                    and the impact of this loop structure mutation in\n                    <jats:italic>B. mori.<\/jats:italic>\n                    First, we predicted potential binding partners for wild-type\n                    <jats:italic>miR-3260<\/jats:italic>\n                    using the RNAhybrid tool and identified eight candidates, including two juvenile hormone (JH)-related genes. Expression analysis revealed high\n                    <jats:italic>miR-3260<\/jats:italic>\n                    expression in the corpora allata. Furthermore, treatment of BmN cells with JH led to upregulated\n                    <jats:italic>miR-3260<\/jats:italic>\n                    expression, suggesting a relationship between\n                    <jats:italic>miR-3260<\/jats:italic>\n                    and JH signaling. Although\n                    <jats:italic>miR-3260<\/jats:italic>\n                    mimics and inhibitors altered the expression of two JH-related candidate genes, the observed changes were not consistent with the predicted regulatory effects of\n                    <jats:italic>miR-3260<\/jats:italic>\n                    , and no observable larval phenotype was detected. Additionally, analysis of Dicer processing showed that neither the wild-type nor mutant\n                    <jats:italic>miR-3260<\/jats:italic>\n                    precursor underwent Dicer-mediated cleavage. Although\n                    <jats:italic>miR-3260<\/jats:italic>\n                    responds to JH, its function as a conventional miRNA was not supported. These findings suggest that\n                    <jats:italic>miR-3260<\/jats:italic>\n                    may act as a JH-responsive non-coding RNA or represent a novel atypical RNA distinct from canonical miRNAs.\n                  <\/jats:p>","DOI":"10.1371\/journal.pone.0346656","type":"journal-article","created":{"date-parts":[[2026,10,6]],"date-time":"2026-10-06T17:39:06Z","timestamp":1791308346000},"page":"e0346656","update-policy":"https:\/\/doi.org\/10.1371\/journal.pone.corrections_policy","source":"Crossref","is-referenced-by-count":0,"title":["Identification of an atypical microRNA with sequence variation in the silkworm Bombyx mori"],"prefix":"10.1371","volume":"21","author":[{"given":"Mayuko","family":"Harada","sequence":"first","affiliation":[{"name":"Graduate School of Agriculture, Tokyo University of Agriculture and Technology","place":["Fuchu, Tokyo, Japan"],"department":["Department of Science of Biological 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